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Sample records for novo designed peptides

  1. De-novo design of antimicrobial peptides for plant protection.

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    Benjamin Zeitler

    Full Text Available This work describes the de-novo design of peptides that inhibit a broad range of plant pathogens. Four structurally different groups of peptides were developed that differ in size and position of their charged and hydrophobic clusters and were assayed for their ability to inhibit bacterial growth and fungal spore germination. Several peptides are highly active at concentrations between 0,1 and 1 µg/ml against plant pathogenic bacteria, such as Pseudomonas syringae, Pectobacterium carotovorum, and Xanthomonas vesicatoria. Importantly, no hemolytic activity could be detected for these peptides at concentrations up to 200 µg/ml. Moreover, the peptides are also active after spraying on the plant surface demonstrating a possible way of application. In sum, our designed peptides represent new antimicrobial agents and with the increasing demand for antimicrobial compounds for production of "healthy" food, these peptides might serve as templates for novel antibacterial and antifungal agents.

  2. De novo peptide design and experimental validation of histone methyltransferase inhibitors.

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    James Smadbeck

    Full Text Available Histones are small proteins critical to the efficient packaging of DNA in the nucleus. DNA–protein complexes, known as nucleosomes, are formed when the DNA winds itself around the surface of the histones. The methylation of histone residues by enhancer of zeste homolog 2 (EZH2 maintains gene repression over successive cell generations. Overexpression of EZH2 can silence important tumor suppressor genes leading to increased invasiveness of many types of cancers. This makes the inhibition of EZH2 an important target in the development of cancer therapeutics. We employed a three-stage computational de novo peptide design method to design inhibitory peptides of EZH2. The method consists of a sequence selection stage and two validation stages for fold specificity and approximate binding affinity. The sequence selection stage consists of an integer linear optimization model that was solved to produce a rank-ordered list of amino acid sequences with increased stability in the bound peptide-EZH2 structure. These sequences were validated through the calculation of the fold specificity and approximate binding affinity of the designed peptides. Here we report the discovery of novel EZH2 inhibitory peptides using the de novo peptide design method. The computationally discovered peptides were experimentally validated in vitro using dose titrations and mechanism of action enzymatic assays. The peptide with the highest in vitro response, SQ037, was validated in nucleo using quantitative mass spectrometry-based proteomics. This peptide had an IC50 of 13.5 mM, demonstrated greater potency as an inhibitor when compared to the native and K27A mutant control peptides, and demonstrated competitive inhibition versus the peptide substrate. Additionally, this peptide demonstrated high specificity to the EZH2 target in comparison to other histone methyltransferases. The validated peptides are the first computationally designed peptides that directly inhibit EZH2

  3. De novo peptide design and experimental validation of histone methyltransferase inhibitors.

    Directory of Open Access Journals (Sweden)

    James Smadbeck

    Full Text Available Histones are small proteins critical to the efficient packaging of DNA in the nucleus. DNA-protein complexes, known as nucleosomes, are formed when the DNA winds itself around the surface of the histones. The methylation of histone residues by enhancer of zeste homolog 2 (EZH2 maintains gene repression over successive cell generations. Overexpression of EZH2 can silence important tumor suppressor genes leading to increased invasiveness of many types of cancers. This makes the inhibition of EZH2 an important target in the development of cancer therapeutics. We employed a three-stage computational de novo peptide design method to design inhibitory peptides of EZH2. The method consists of a sequence selection stage and two validation stages for fold specificity and approximate binding affinity. The sequence selection stage consists of an integer linear optimization model that was solved to produce a rank-ordered list of amino acid sequences with increased stability in the bound peptide-EZH2 structure. These sequences were validated through the calculation of the fold specificity and approximate binding affinity of the designed peptides. Here we report the discovery of novel EZH2 inhibitory peptides using the de novo peptide design method. The computationally discovered peptides were experimentally validated in vitro using dose titrations and mechanism of action enzymatic assays. The peptide with the highest in vitro response, SQ037, was validated in nucleo using quantitative mass spectrometry-based proteomics. This peptide had an IC50 of 13.5 [Formula: see text]M, demonstrated greater potency as an inhibitor when compared to the native and K27A mutant control peptides, and demonstrated competitive inhibition versus the peptide substrate. Additionally, this peptide demonstrated high specificity to the EZH2 target in comparison to other histone methyltransferases. The validated peptides are the first computationally designed peptides that directly

  4. Antimicrobial peptide capsids of de novo design.

    Science.gov (United States)

    De Santis, Emiliana; Alkassem, Hasan; Lamarre, Baptiste; Faruqui, Nilofar; Bella, Angelo; Noble, James E; Micale, Nicola; Ray, Santanu; Burns, Jonathan R; Yon, Alexander R; Hoogenboom, Bart W; Ryadnov, Maxim G

    2017-12-22

    The spread of bacterial resistance to antibiotics poses the need for antimicrobial discovery. With traditional search paradigms being exhausted, approaches that are altogether different from antibiotics may offer promising and creative solutions. Here, we introduce a de novo peptide topology that-by emulating the virus architecture-assembles into discrete antimicrobial capsids. Using the combination of high-resolution and real-time imaging, we demonstrate that these artificial capsids assemble as 20-nm hollow shells that attack bacterial membranes and upon landing on phospholipid bilayers instantaneously (seconds) convert into rapidly expanding pores causing membrane lysis (minutes). The designed capsids show broad antimicrobial activities, thus executing one primary function-they destroy bacteria on contact.

  5. UniNovo: a universal tool for de novo peptide sequencing.

    Science.gov (United States)

    Jeong, Kyowon; Kim, Sangtae; Pevzner, Pavel A

    2013-08-15

    Mass spectrometry (MS) instruments and experimental protocols are rapidly advancing, but de novo peptide sequencing algorithms to analyze tandem mass (MS/MS) spectra are lagging behind. Although existing de novo sequencing tools perform well on certain types of spectra [e.g. Collision Induced Dissociation (CID) spectra of tryptic peptides], their performance often deteriorates on other types of spectra, such as Electron Transfer Dissociation (ETD), Higher-energy Collisional Dissociation (HCD) spectra or spectra of non-tryptic digests. Thus, rather than developing a new algorithm for each type of spectra, we develop a universal de novo sequencing algorithm called UniNovo that works well for all types of spectra or even for spectral pairs (e.g. CID/ETD spectral pairs). UniNovo uses an improved scoring function that captures the dependences between different ion types, where such dependencies are learned automatically using a modified offset frequency function. The performance of UniNovo is compared with PepNovo+, PEAKS and pNovo using various types of spectra. The results show that the performance of UniNovo is superior to other tools for ETD spectra and superior or comparable with others for CID and HCD spectra. UniNovo also estimates the probability that each reported reconstruction is correct, using simple statistics that are readily obtained from a small training dataset. We demonstrate that the estimation is accurate for all tested types of spectra (including CID, HCD, ETD, CID/ETD and HCD/ETD spectra of trypsin, LysC or AspN digested peptides). UniNovo is implemented in JAVA and tested on Windows, Ubuntu and OS X machines. UniNovo is available at http://proteomics.ucsd.edu/Software/UniNovo.html along with the manual.

  6. MRUniNovo: an efficient tool for de novo peptide sequencing utilizing the hadoop distributed computing framework.

    Science.gov (United States)

    Li, Chuang; Chen, Tao; He, Qiang; Zhu, Yunping; Li, Kenli

    2017-03-15

    Tandem mass spectrometry-based de novo peptide sequencing is a complex and time-consuming process. The current algorithms for de novo peptide sequencing cannot rapidly and thoroughly process large mass spectrometry datasets. In this paper, we propose MRUniNovo, a novel tool for parallel de novo peptide sequencing. MRUniNovo parallelizes UniNovo based on the Hadoop compute platform. Our experimental results demonstrate that MRUniNovo significantly reduces the computation time of de novo peptide sequencing without sacrificing the correctness and accuracy of the results, and thus can process very large datasets that UniNovo cannot. MRUniNovo is an open source software tool implemented in java. The source code and the parameter settings are available at http://bioinfo.hupo.org.cn/MRUniNovo/index.php. s131020002@hnu.edu.cn ; taochen1019@163.com. Supplementary data are available at Bioinformatics online. © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com

  7. Recurrent Neural Network Model for Constructive Peptide Design.

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    Müller, Alex T; Hiss, Jan A; Schneider, Gisbert

    2018-02-26

    We present a generative long short-term memory (LSTM) recurrent neural network (RNN) for combinatorial de novo peptide design. RNN models capture patterns in sequential data and generate new data instances from the learned context. Amino acid sequences represent a suitable input for these machine-learning models. Generative models trained on peptide sequences could therefore facilitate the design of bespoke peptide libraries. We trained RNNs with LSTM units on pattern recognition of helical antimicrobial peptides and used the resulting model for de novo sequence generation. Of these sequences, 82% were predicted to be active antimicrobial peptides compared to 65% of randomly sampled sequences with the same amino acid distribution as the training set. The generated sequences also lie closer to the training data than manually designed amphipathic helices. The results of this study showcase the ability of LSTM RNNs to construct new amino acid sequences within the applicability domain of the model and motivate their prospective application to peptide and protein design without the need for the exhaustive enumeration of sequence libraries.

  8. Identification of a novel Plasmopara halstedii elicitor protein combining de novo peptide sequencing algorithms and RACE-PCR

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    Madlung Johannes

    2010-05-01

    Full Text Available Abstract Background Often high-quality MS/MS spectra of tryptic peptides do not match to any database entry because of only partially sequenced genomes and therefore, protein identification requires de novo peptide sequencing. To achieve protein identification of the economically important but still unsequenced plant pathogenic oomycete Plasmopara halstedii, we first evaluated the performance of three different de novo peptide sequencing algorithms applied to a protein digests of standard proteins using a quadrupole TOF (QStar Pulsar i. Results The performance order of the algorithms was PEAKS online > PepNovo > CompNovo. In summary, PEAKS online correctly predicted 45% of measured peptides for a protein test data set. All three de novo peptide sequencing algorithms were used to identify MS/MS spectra of tryptic peptides of an unknown 57 kDa protein of P. halstedii. We found ten de novo sequenced peptides that showed homology to a Phytophthora infestans protein, a closely related organism of P. halstedii. Employing a second complementary approach, verification of peptide prediction and protein identification was performed by creation of degenerate primers for RACE-PCR and led to an ORF of 1,589 bp for a hypothetical phosphoenolpyruvate carboxykinase. Conclusions Our study demonstrated that identification of proteins within minute amounts of sample material improved significantly by combining sensitive LC-MS methods with different de novo peptide sequencing algorithms. In addition, this is the first study that verified protein prediction from MS data by also employing a second complementary approach, in which RACE-PCR led to identification of a novel elicitor protein in P. halstedii.

  9. Designing anticancer peptides by constructive machine learning.

    Science.gov (United States)

    Grisoni, Francesca; Neuhaus, Claudia; Gabernet, Gisela; Müller, Alex; Hiss, Jan; Schneider, Gisbert

    2018-04-21

    Constructive machine learning enables the automated generation of novel chemical structures without the need for explicit molecular design rules. This study presents the experimental application of such a generative model to design membranolytic anticancer peptides (ACPs) de novo. A recurrent neural network with long short-term memory cells was trained on alpha-helical cationic amphipathic peptide sequences and then fine-tuned with 26 known ACPs. This optimized model was used to generate unique and novel amino acid sequences. Twelve of the peptides were synthesized and tested for their activity on MCF7 human breast adenocarcinoma cells and selectivity against human erythrocytes. Ten of these peptides were active against cancer cells. Six of the active peptides killed MCF7 cancer cells without affecting human erythrocytes with at least threefold selectivity. These results advocate constructive machine learning for the automated design of peptides with desired biological activities. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  10. Peptide de novo sequencing of mixture tandem mass spectra

    DEFF Research Database (Denmark)

    Gorshkov, Vladimir; Hotta, Stéphanie Yuki Kolbeck; Braga, Thiago Verano

    2016-01-01

    they decrease the identification performance using database search engines. De novo sequencing approaches are expected to be even more sensitive to the reduction in mass spectrum quality resulting from peptide precursor co-isolation and thus prone to false identifications. The deconvolution approach matched...... complementary b-, y-ions to each precursor peptide mass, which allowed the creation of virtual spectra containing sequence specific fragment ions of each co-isolated peptide. Deconvolution processing resulted in equally efficient identification rates but increased the absolute number of correctly sequenced...... peptides. The improvement was in the range of 20–35% additional peptide identifications for a HeLa lysate sample. Some correct sequences were identified only using unprocessed spectra; however, the number of these was lower than those where improvement was obtained by mass spectral deconvolution. Tight...

  11. Anti-plasmodial action of de novo-designed, cationic, lysine-branched, amphipathic, helical peptides

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    Kaushik Naveen K

    2012-08-01

    Full Text Available Abstract Background A lack of vaccine and rampant drug resistance demands new anti-malarials. Methods In vitro blood stage anti-plasmodial properties of several de novo-designed, chemically synthesized, cationic, amphipathic, helical, antibiotic peptides were examined against Plasmodium falciparum using SYBR Green assay. Mechanistic details of anti-plasmodial action were examined by optical/fluorescence microscopy and FACS analysis. Results Unlike the monomeric decapeptides {(Ac-GXRKXHKXWA-NH2 (X = F,ΔF (Fm, ΔFm IC50 >100 μM}, the lysine-branched,dimeric versions showed far greater potency {IC50 (μM Fd 1.5 , ΔFd 1.39}. The more helical and proteolytically stable ΔFd was studied for mechanistic details. ΔFq, a K-K2 dendrimer of ΔFm and (ΔFm2 a linear dimer of ΔFm showed IC50 (μM of 0.25 and 2.4 respectively. The healthy/infected red cell selectivity indices were >35 (ΔFd, >20 (ΔFm2 and 10 (ΔFq. FITC-ΔFd showed rapid and selective accumulation in parasitized red cells. Overlaying DAPI and FITC florescence suggested that ΔFd binds DNA. Trophozoites and schizonts incubated with ΔFd (2.5 μM egressed anomalously and Band-3 immunostaining revealed them not to be associated with RBC membrane. Prematurely egressed merozoites from peptide-treated cultures were found to be invasion incompetent. Conclusion Good selectivity (>35, good resistance index (1.1 and low cytotoxicity indicate the promise of ΔFd against malaria.

  12. Library Design-Facilitated High-Throughput Sequencing of Synthetic Peptide Libraries.

    Science.gov (United States)

    Vinogradov, Alexander A; Gates, Zachary P; Zhang, Chi; Quartararo, Anthony J; Halloran, Kathryn H; Pentelute, Bradley L

    2017-11-13

    A methodology to achieve high-throughput de novo sequencing of synthetic peptide mixtures is reported. The approach leverages shotgun nanoliquid chromatography coupled with tandem mass spectrometry-based de novo sequencing of library mixtures (up to 2000 peptides) as well as automated data analysis protocols to filter away incorrect assignments, noise, and synthetic side-products. For increasing the confidence in the sequencing results, mass spectrometry-friendly library designs were developed that enabled unambiguous decoding of up to 600 peptide sequences per hour while maintaining greater than 85% sequence identification rates in most cases. The reliability of the reported decoding strategy was additionally confirmed by matching fragmentation spectra for select authentic peptides identified from library sequencing samples. The methods reported here are directly applicable to screening techniques that yield mixtures of active compounds, including particle sorting of one-bead one-compound libraries and affinity enrichment of synthetic library mixtures performed in solution.

  13. Metal Stabilization of Collagen and de Novo Designed Mimetic Peptides.

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    Parmar, Avanish S; Xu, Fei; Pike, Douglas H; Belure, Sandeep V; Hasan, Nida F; Drzewiecki, Kathryn E; Shreiber, David I; Nanda, Vikas

    2015-08-18

    We explore the design of metal binding sites to modulate triple-helix stability of collagen and collagen-mimetic peptides. Globular proteins commonly utilize metals to connect tertiary structural elements that are well separated in sequence, constraining structure and enhancing stability. It is more challenging to engineer structural metals into fibrous protein scaffolds, which lack the extensive tertiary contacts seen in globular proteins. In the collagen triple helix, the structural adjacency of the carboxy-termini of the three chains makes this region an attractive target for introducing metal binding sites. We engineered His3 sites based on structural modeling constraints into a series of designed homotrimeric and heterotrimeric peptides, assessing the capacity of metal binding to improve stability and in the case of heterotrimers, affect specificity of assembly. Notable enhancements in stability for both homo- and heteromeric systems were observed upon addition of zinc(II) and several other metal ions only when all three histidine ligands were present. Metal binding affinities were consistent with the expected Irving-Williams series for imidazole. Unlike other metals tested, copper(II) also bound to peptides lacking histidine ligands. Acetylation of the peptide N-termini prevented copper binding, indicating proline backbone amide metal-coordination at this site. Copper similarly stabilized animal extracted Type I collagen in a metal-specific fashion, highlighting the potential importance of metal homeostasis within the extracellular matrix.

  14. Inducible Expression of the De-Novo Designed Antimicrobial Peptide SP1-1 in Tomato Confers Resistance to Xanthomonas campestris pv. vesicatoria.

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    Areli Herrera Diaz

    Full Text Available Antimicrobial peptides (AMPs are small peptides with less than 50 amino acids and are part of the innate immune response in almost all organisms, including bacteria, vertebrates, invertebrates and plants. AMPs are active against a broad-spectrum of pathogens. The inducible expression of AMPs in plants is a promising approach to combat plant pathogens with minimal negative side effects, such as phytotoxicity or infertility. In this study, inducible expression of the de-novo designed AMP SP1-1 in Micro Tom tomato protected tomato fruits against bacterial spot disease caused by Xanthomonas campestris pv. vesicatoria. The peptide SP1-1 was targeted to the apoplast which is the primary infection site for plant pathogens, by fusing SP1-1 peptide to the signal peptide RsAFP1 of radish (Raphanus sativus. The pathogen inducibility of the expression was enabled by using an optimized inducible 4XW2/4XS promoter. As a result, the tomato fruits of independently generated SP1-1 transgenic lines were significantly more resistant to X. campestris pv. vesicatoria than WT tomato fruits. In transgenic lines, bacterial infection was reduced up to 65% in comparison to the infection of WT plants. Our study demonstrates that the combination of the 4XW2/4XS cis-element from parsley with the synthetic antimicrobial peptide SP1-1 is a good alternative to protect tomato fruits against infections with X. campestris pv. vesicatoria.

  15. Shape-specific nanostructured protein mimics from de novo designed chimeric peptides.

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    Jiang, Linhai; Yang, Su; Lund, Reidar; Dong, He

    2018-01-30

    Natural proteins self-assemble into highly-ordered nanoscaled architectures to perform specific functions. The intricate functions of proteins have provided great impetus for researchers to develop strategies for designing and engineering synthetic nanostructures as protein mimics. Compared to the success in engineering fibrous protein mimetics, the design of discrete globular protein-like nanostructures has been challenging mainly due to the lack of precise control over geometric packing and intermolecular interactions among synthetic building blocks. In this contribution, we report an effective strategy to construct shape-specific nanostructures based on the self-assembly of chimeric peptides consisting of a coiled coil dimer and a collagen triple helix folding motif. Under salt-free conditions, we showed spontaneous self-assembly of the chimeric peptides into monodisperse, trigonal bipyramidal-like nanoparticles with precise control over the stoichiometry of two folding motifs and the geometrical arrangements relative to one another. Three coiled coil dimers are interdigitated on the equatorial plane while the two collagen triple helices are located in the axial position, perpendicular to the coiled coil plane. A detailed molecular model was proposed and further validated by small angle X-ray scattering experiments and molecular dynamics (MD) simulation. The results from this study indicated that the molecular folding of each motif within the chimeric peptides and their geometric packing played important roles in the formation of discrete protein-like nanoparticles. The peptide design and self-assembly mechanism may open up new routes for the construction of highly organized, discrete self-assembling protein-like nanostructures with greater levels of control over assembly accuracy.

  16. Apoprotein Structure and Metal Binding Characterization of a de Novo Designed Peptide, α3DIV, that Sequesters Toxic Heavy Metals.

    Science.gov (United States)

    Plegaria, Jefferson S; Dzul, Stephen P; Zuiderweg, Erik R P; Stemmler, Timothy L; Pecoraro, Vincent L

    2015-05-12

    De novo protein design is a biologically relevant approach that provides a novel process in elucidating protein folding and modeling the metal centers of metalloproteins in a completely unrelated or simplified fold. An integral step in de novo protein design is the establishment of a well-folded scaffold with one conformation, which is a fundamental characteristic of many native proteins. Here, we report the NMR solution structure of apo α3DIV at pH 7.0, a de novo designed three-helix bundle peptide containing a triscysteine motif (Cys18, Cys28, and Cys67) that binds toxic heavy metals. The structure comprises 1067 NOE restraints derived from multinuclear multidimensional NOESY, as well as 138 dihedral angles (ψ, φ, and χ1). The backbone and heavy atoms of the 20 lowest energy structures have a root mean square deviation from the mean structure of 0.79 (0.16) Å and 1.31 (0.15) Å, respectively. When compared to the parent structure α3D, the substitution of Leu residues to Cys enhanced the α-helical content of α3DIV while maintaining the same overall topology and fold. In addition, solution studies on the metalated species illustrated metal-induced stability. An increase in the melting temperatures was observed for Hg(II), Pb(II), or Cd(II) bound α3DIV by 18-24 °C compared to its apo counterpart. Further, the extended X-ray absorption fine structure analysis on Hg(II)-α3DIV produced an average Hg(II)-S bond length at 2.36 Å, indicating a trigonal T-shaped coordination environment. Overall, the structure of apo α3DIV reveals an asymmetric distorted triscysteine metal binding site, which offers a model for native metalloregulatory proteins with thiol-rich ligands that function in regulating toxic heavy metals, such as ArsR, CadC, MerR, and PbrR.

  17. Bromine isotopic signature facilitates de novo sequencing of peptides in free-radical-initiated peptide sequencing (FRIPS) mass spectrometry.

    Science.gov (United States)

    Nam, Jungjoo; Kwon, Hyuksu; Jang, Inae; Jeon, Aeran; Moon, Jingyu; Lee, Sun Young; Kang, Dukjin; Han, Sang Yun; Moon, Bongjin; Oh, Han Bin

    2015-02-01

    We recently showed that free-radical-initiated peptide sequencing mass spectrometry (FRIPS MS) assisted by the remarkable thermochemical stability of (2,2,6,6-tetramethyl-piperidin-1-yl)oxyl (TEMPO) is another attractive radical-driven peptide fragmentation MS tool. Facile homolytic cleavage of the bond between the benzylic carbon and the oxygen of the TEMPO moiety in o-TEMPO-Bz-C(O)-peptide and the high reactivity of the benzylic radical species generated in •Bz-C(O)-peptide are key elements leading to extensive radical-driven peptide backbone fragmentation. In the present study, we demonstrate that the incorporation of bromine into the benzene ring, i.e. o-TEMPO-Bz(Br)-C(O)-peptide, allows unambiguous distinction of the N-terminal peptide fragments from the C-terminal fragments through the unique bromine doublet isotopic signature. Furthermore, bromine substitution does not alter the overall radical-driven peptide backbone dissociation pathways of o-TEMPO-Bz-C(O)-peptide. From a practical perspective, the presence of the bromine isotopic signature in the N-terminal peptide fragments in TEMPO-assisted FRIPS MS represents a useful and cost-effective opportunity for de novo peptide sequencing. Copyright © 2015 John Wiley & Sons, Ltd.

  18. Metal stabilization of collagen and de novo designed mimetic peptides

    OpenAIRE

    Parmar, Avanish S.; Xu, Fei; Pike, Douglas H.; Belure, Sandeep V.; Hasan, Nida F.; Drzewiecki, Kathryn E.; Shreiber, David I.; Nanda, Vikas

    2015-01-01

    We explore the design of metal binding sites to modulate triple-helix stability of collagen and collagen-mimetic peptides. Globular proteins commonly utilize metals to connect tertiary structural elements that are well separated in sequence, constraining structure and enhancing stability. It is more challenging to engineer structural metals into fibrous protein scaffolds, which lack the extensive tertiary contacts seen in globular proteins. In the collagen triple helix, the structural adjacen...

  19. LESSONS IN DE NOVO PEPTIDE SEQUENCING BY TANDEM MASS SPECTROMETRY

    Science.gov (United States)

    Medzihradszky, Katalin F.; Chalkley, Robert J.

    2015-01-01

    Mass spectrometry has become the method of choice for the qualitative and quantitative characterization of protein mixtures isolated from all kinds of living organisms. The raw data in these studies are MS/MS spectra, usually of peptides produced by proteolytic digestion of a protein. These spectra are “translated” into peptide sequences, normally with the help of various search engines. Data acquisition and interpretation have both been automated, and most researchers look only at the summary of the identifications without ever viewing the underlying raw data used for assignments. Automated analysis of data is essential due to the volume produced. However, being familiar with the finer intricacies of peptide fragmentation processes, and experiencing the difficulties of manual data interpretation allow a researcher to be able to more critically evaluate key results, particularly because there are many known rules of peptide fragmentation that are not incorporated into search engine scoring. Since the most commonly used MS/MS activation method is collision-induced dissociation (CID), in this article we present a brief review of the history of peptide CID analysis. Next, we provide a detailed tutorial on how to determine peptide sequences from CID data. Although the focus of the tutorial is de novo sequencing, the lessons learned and resources supplied are useful for data interpretation in general. PMID:25667941

  20. De novo design and engineering of non-ribosomal peptide synthetases

    Science.gov (United States)

    Bozhüyük, Kenan A. J.; Fleischhacker, Florian; Linck, Annabell; Wesche, Frank; Tietze, Andreas; Niesert, Claus-Peter; Bode, Helge B.

    2018-03-01

    Peptides derived from non-ribosomal peptide synthetases (NRPSs) represent an important class of pharmaceutically relevant drugs. Methods to generate novel non-ribosomal peptides or to modify peptide natural products in an easy and predictable way are therefore of great interest. However, although the overall modular structure of NRPSs suggests the possibility of adjusting domain specificity and selectivity, only a few examples have been reported and these usually show a severe drop in production titre. Here we report a new strategy for the modification of NRPSs that uses defined exchange units (XUs) and not modules as functional units. XUs are fused at specific positions that connect the condensation and adenylation domains and respect the original specificity of the downstream module to enable the production of the desired peptides. We also present the use of internal condensation domains as an alternative to other peptide-chain-releasing domains for the production of cyclic peptides.

  1. De novo sequencing of two novel peptides homologous to calcitonin-like peptides, from skin secretion of the Chinese Frog, Odorrana schmackeri

    NARCIS (Netherlands)

    Evaristo, Geisa P C; Pinkse, Martijn W H; Chen, Tianbao; Wang, Lei; Mohammed, Shabaz; Heck, Albert J R; Mathes, Isabella; Lottspeich, Friedrich; Shaw, Chris; Albar, Juan Pablo; Verhaert, Peter D E M

    2015-01-01

    An MS/MS based analytical strategy was followed to solve the complete sequence of two new peptides from frog (Odorrana schmackeri) skin secretion. This involved reduction and alkylation with two different alkylating agents followed by high resolution tandem mass spectrometry. De novo sequencing was

  2. Membrane interaction and secondary structure of de novo designed arginine-and tryptophan peptides with dual function

    KAUST Repository

    Rydberg, Hanna A.

    2012-10-01

    Cell-penetrating peptides and antimicrobial peptides are two classes of positively charged membrane active peptides with several properties in common. The challenge is to combine knowledge about the membrane interaction mechanisms and structural properties of the two classes to design peptides with membrane-specific actions, useful either as transporters of cargo or as antibacterial substances. Membrane active peptides are commonly rich in arginine and tryptophan. We have previously designed a series of arg/trp peptides and investigated how the position and number of tryptophans affect cellular uptake. Here we explore the antimicrobial properties and the interaction with lipid model membranes of these peptides, using minimal inhibitory concentrations assay (MIC), circular dichroism (CD) and linear dichroism (LD). The results show that the arg/trp peptides inhibit the growth of the two gram positive strains Staphylococcus aureus and Staphylococcus pyogenes, with some individual variations depending on the position of the tryptophans. No inhibition of the gram negative strains Proteus mirabilis or Pseudomonas aeruginosa was noticed. CD indicated that when bound to lipid vesicles one of the peptides forms an α-helical like structure, whereas the other five exhibited rather random coiled structures. LD indicated that all six peptides were somehow aligned parallel with the membrane surface. Our results do not reveal any obvious connection between membrane interaction and antimicrobial effect for the studied peptides. By contrast cell-penetrating properties can be coupled to both the secondary structure and the degree of order of the peptides. © 2012 Elsevier Inc.

  3. Biological effects of a de novo designed myxoma virus peptide analogue: evaluation of cytotoxicity on tumor cells.

    Directory of Open Access Journals (Sweden)

    Taghrid S Istivan

    Full Text Available BACKGROUND: The Resonant Recognition Model (RRM is a physico-mathematical model that interprets protein sequence linear information using digital signal processing methods. In this study the RRM concept was employed for structure-function analysis of myxoma virus (MV proteins and the design of a short bioactive therapeutic peptide with MV-like antitumor/cytotoxic activity. METHODOLOGY/PRINCIPAL FINDINGS: The analogue RRM-MV was designed by RRM as a linear 18 aa 2.3 kDa peptide. The biological activity of this computationally designed peptide analogue against cancer and normal cell lines was investigated. The cellular cytotoxicity effects were confirmed by confocal immunofluorescence microscopy, by measuring the levels of cytoplasmic lactate dehydrogenase (LDH and by Prestoblue cell viability assay for up to 72 hours in peptide treated and non-treated cell cultures. Our results revealed that RRM-MV induced a significant dose and time-dependent cytotoxic effect on murine and human cancer cell lines. Yet, when normal murine cell lines were similarly treated with RRM-MV, no cytotoxic effects were observed. Furthermore, the non-bioactive RRM designed peptide RRM-C produced negligible cytotoxic effects on these cancer and normal cell lines when used at similar concentrations. The presence/absence of phosphorylated Akt activity in B16F0 mouse melanoma cells was assessed to indicate the possible apoptosis signalling pathway that could be affected by the peptide treatment. So far, Akt activity did not seem to be significantly affected by RRM-MV as is the case for the original viral protein. CONCLUSIONS/SIGNIFICANCE: Our findings indicate the successful application of the RRM concept to design a bioactive peptide analogue (RRM-MV with cytotoxic effects on tumor cells only. This 2.345 kDa peptide analogue to a 49 kDa viral protein may be suitable to be developed as a potential cancer therapeutic. These results also open a new direction to the rational

  4. CycloBranch: De Novo Sequencing of Nonribosomal Peptides from Accurate Product Ion Mass Spectra

    Czech Academy of Sciences Publication Activity Database

    Novák, Jiří; Lemr, Karel; Schug, K. A.; Havlíček, Vladimír

    2015-01-01

    Roč. 26, č. 10 (2015), s. 1780-1786 ISSN 1044-0305 R&D Projects: GA ČR(CZ) GAP206/12/1150 Grant - others:OPPC(XE) CZ.2.16/3.1.00/24023 Institutional support: RVO:61388971 Keywords : De novo sequencing * Nonribosomal peptides * Linear Subject RIV: CE - Biochemistry Impact factor: 3.031, year: 2015

  5. de novo computational enzyme design.

    Science.gov (United States)

    Zanghellini, Alexandre

    2014-10-01

    Recent advances in systems and synthetic biology as well as metabolic engineering are poised to transform industrial biotechnology by allowing us to design cell factories for the sustainable production of valuable fuels and chemicals. To deliver on their promises, such cell factories, as much as their brick-and-mortar counterparts, will require appropriate catalysts, especially for classes of reactions that are not known to be catalyzed by enzymes in natural organisms. A recently developed methodology, de novo computational enzyme design can be used to create enzymes catalyzing novel reactions. Here we review the different classes of chemical reactions for which active protein catalysts have been designed as well as the results of detailed biochemical and structural characterization studies. We also discuss how combining de novo computational enzyme design with more traditional protein engineering techniques can alleviate the shortcomings of state-of-the-art computational design techniques and create novel enzymes with catalytic proficiencies on par with natural enzymes. Copyright © 2014 Elsevier Ltd. All rights reserved.

  6. De novo sequencing of two novel peptides homologous to calcitonin-like peptides, from skin secretion of the Chinese Frog, Odorrana schmackeri

    Directory of Open Access Journals (Sweden)

    Geisa P.C. Evaristo

    2015-09-01

    Full Text Available An MS/MS based analytical strategy was followed to solve the complete sequence of two new peptides from frog (Odorrana schmackeri skin secretion. This involved reduction and alkylation with two different alkylating agents followed by high resolution tandem mass spectrometry. De novo sequencing was achieved by complementary CID and ETD fragmentations of full-length peptides and of selected tryptic fragments. Heavy and light isotope dimethyl labeling assisted with annotation of sequence ion series. The identified primary structures are GCD[I/L]STCATHN[I/L]VNE[I/L]NKFDKSKPSSGGVGPESP-NH2 and SCNLSTCATHNLVNELNKFDKSKPSSGGVGPESF-NH2, i.e. two carboxyamidated 34 residue peptides with an aminoterminal intramolecular ring structure formed by a disulfide bridge between Cys2 and Cys7. Edman degradation analysis of the second peptide positively confirmed the exact sequence, resolving I/L discriminations. Both peptide sequences are novel and share homology with calcitonin, calcitonin gene related peptide (CGRP and adrenomedullin from other vertebrates. Detailed sequence analysis as well as the 34 residue length of both O. schmackeri peptides, suggest they do not fully qualify as either calcitonins (32 residues or CGRPs (37 amino acids and may justify their classification in a novel peptide family within the calcitonin gene related peptide superfamily. Smooth muscle contractility assays with synthetic replicas of the S–S linked peptides on rat tail artery, uterus, bladder and ileum did not reveal myotropic activity.

  7. Cyclic peptide therapeutics: past, present and future.

    Science.gov (United States)

    Zorzi, Alessandro; Deyle, Kaycie; Heinis, Christian

    2017-06-01

    Cyclic peptides combine several favorable properties such as good binding affinity, target selectivity and low toxicity that make them an attractive modality for the development of therapeutics. Over 40 cyclic peptide drugs are currently in clinical use and around one new cyclic peptide drug enters the market every year on average. The vast majority of clinically approved cyclic peptides are derived from natural products, such as antimicrobials or human peptide hormones. New powerful techniques based on rational design and in vitro evolution have enabled the de novo development of cyclic peptide ligands to targets for which nature does not offer solutions. A look at the cyclic peptides currently under clinical evaluation shows that several have been developed using such techniques. This new source for cyclic peptide ligands introduces a freshness to the field, and it is likely that de novo developed cyclic peptides will be in clinical use in the near future. Copyright © 2017 Elsevier Ltd. All rights reserved.

  8. Rational design of peptide affinity ligands for the purification of therapeutic enzymes.

    Science.gov (United States)

    Trasatti, John P; Woo, James; Ladiwala, Asif; Cramer, Steven; Karande, Pankaj

    2018-04-25

    Non-mAb biologics represent a growing class of therapeutics under clinical development. Although affinity chromatography is a potentially attractive approach for purification, the development of platform technologies, such as Protein A for mAbs, has been challenging due to the inherent chemical and structural diversity of these molecules. Here, we present our studies on the rapid development of peptide affinity ligands for the purification of biologics using a prototypical enzyme therapeutic in clinical use. Employing a suite of de novo rational and combinatorial design strategies we designed and screened a library of peptides on microarray platforms for their ability to bind to the target with high affinity and selectivity in cell culture fluid. Lead peptides were evaluated on resin in batch conditions and compared with a commercially available resin to evaluate their efficacy. Two lead candidates identified from microarray studies provided high binding capacity to the target while demonstrating high selectivity against culture contaminants and product variants compared to a commercial resin system. These findings provide a proof-of-concept for developing affinity peptide-based bioseparations processes for a target biologic. Peptide affinity ligand design and screening approaches presented in this work can also be easily translated to other biologics of interest. © 2018 American Institute of Chemical Engineers Biotechnol. Prog., 2018. © 2018 American Institute of Chemical Engineers.

  9. 2-d and 1-d Nanomaterials Construction through Peptide Computational Design and Solution Assembly

    Science.gov (United States)

    Pochan, Darrin

    Self-assembly of molecules is an attractive materials construction strategy due to its simplicity in application. By considering peptidic molecules in the bottom-up materials self-assembly design process, one can take advantage of inherently biomolecular attributes; intramolecular folding events, secondary structure, and electrostatic/H-bonding/hydrophobic interactions to define hierarchical material structure and consequent properties. Importantly, while biomimicry has been a successful strategy for the design of new peptide molecules for intermolecular assembly, computational tools have been developed to de novo design peptide molecules required for construction of pre-determined, desired nanostructures and materials. A new system comprised of coiled coil bundle motifs theoretically designed to assemble into designed, one and two-dimensional nanostructures will be introduced. The strategy provides the opportunity for arbitrary nanostructure formation, i.e. structures not observed in nature, with peptide molecules. Importantly, the desired nanostructure was chosen first while the peptides needed for coiled coil formation and subsequent nanomaterial formation were determined computationally. Different interbundle, two-dimensional nanostructures are stabilized by differences in amino acid composition exposed on the exterior of the coiled coil bundles. Computation was able to determine molecules required for different interbundle symmetries within two-dimensional sheets stabilized by subtle differences in amino acid composition of the inherent peptides. Finally, polymers were also created through covalent interactions between bundles that allowed formation of architectures spanning flexible network forming chains to ultra-stiff polymers, all with the same building block peptides. The success of the computational design strategy is manifested in the nanomaterial results as characterized by electron microscopy, scattering methods, and biophysical techniques. Support

  10. Generative Recurrent Networks for De Novo Drug Design.

    Science.gov (United States)

    Gupta, Anvita; Müller, Alex T; Huisman, Berend J H; Fuchs, Jens A; Schneider, Petra; Schneider, Gisbert

    2018-01-01

    Generative artificial intelligence models present a fresh approach to chemogenomics and de novo drug design, as they provide researchers with the ability to narrow down their search of the chemical space and focus on regions of interest. We present a method for molecular de novo design that utilizes generative recurrent neural networks (RNN) containing long short-term memory (LSTM) cells. This computational model captured the syntax of molecular representation in terms of SMILES strings with close to perfect accuracy. The learned pattern probabilities can be used for de novo SMILES generation. This molecular design concept eliminates the need for virtual compound library enumeration. By employing transfer learning, we fine-tuned the RNN's predictions for specific molecular targets. This approach enables virtual compound design without requiring secondary or external activity prediction, which could introduce error or unwanted bias. The results obtained advocate this generative RNN-LSTM system for high-impact use cases, such as low-data drug discovery, fragment based molecular design, and hit-to-lead optimization for diverse drug targets. © 2017 The Authors. Published by Wiley-VCH Verlag GmbH & Co. KGaA.

  11. DeNovoGUI: an open source graphical user interface for de novo sequencing of tandem mass spectra.

    Science.gov (United States)

    Muth, Thilo; Weilnböck, Lisa; Rapp, Erdmann; Huber, Christian G; Martens, Lennart; Vaudel, Marc; Barsnes, Harald

    2014-02-07

    De novo sequencing is a popular technique in proteomics for identifying peptides from tandem mass spectra without having to rely on a protein sequence database. Despite the strong potential of de novo sequencing algorithms, their adoption threshold remains quite high. We here present a user-friendly and lightweight graphical user interface called DeNovoGUI for running parallelized versions of the freely available de novo sequencing software PepNovo+, greatly simplifying the use of de novo sequencing in proteomics. Our platform-independent software is freely available under the permissible Apache2 open source license. Source code, binaries, and additional documentation are available at http://denovogui.googlecode.com .

  12. Evolving the use of peptides as biomaterials components

    Science.gov (United States)

    Collier, Joel H.; Segura, Tatiana

    2012-01-01

    This manuscript is part of a debate on the statement that “the use of short synthetic adhesion peptides, like RGD, is the best approach in the design of biomaterials that guide cell behavior for regenerative medicine and tissue engineering”. We take the position that although there are some acknowledged disadvantages of using short peptide ligands within biomaterials, it is not necessary to discard the notion of using peptides within biomaterials entirely, but rather to reinvent and evolve their use. Peptides possess advantageous chemical definition, access to non-native chemistries, amenability to de novo design, and applicability within parallel approaches. Biomaterials development programs that require such aspects may benefit from a peptide-based strategy. PMID:21515167

  13. Computer-Aided Design of Antimicrobial Peptides

    DEFF Research Database (Denmark)

    Fjell, Christopher D.; Hancock, Robert E.W.; Jenssen, Håvard

    2010-01-01

    in antimicrobial activity. Consequently, the majority of peptides put into clinical trials have failed at some point, underlining the importance of a thorough peptide optimization. An important tool in peptide design and optimization is quantitative structure-activity relationship (QSAR) analysis, correlating...... chemical parameters with biological activities of the peptide, using statistical methods. In this review we will discuss two different in silico strategies of computer-aided antibacterial peptide design, a linear correlation model build as an extension of traditional principal component analysis (PCA......) and a non-linear artificial neural network model. Studies on structurally diverse peptides, have concluded that the PCA derived model are able to guide the antibacterial peptide design in a meaningful way, however requiring rather a high homology between the peptides in the test-set and the in silico...

  14. In Silico Generation of Peptides by Replica Exchange Monte Carlo: Docking-Based Optimization of Maltose-Binding-Protein Ligands.

    Directory of Open Access Journals (Sweden)

    Anna Russo

    Full Text Available Short peptides can be designed in silico and synthesized through automated techniques, making them advantageous and versatile protein binders. A number of docking-based algorithms allow for a computational screening of peptides as binders. Here we developed ex-novo peptides targeting the maltose site of the Maltose Binding Protein, the prototypical system for the study of protein ligand recognition. We used a Monte Carlo based protocol, to computationally evolve a set of octapeptides starting from a polialanine sequence. We screened in silico the candidate peptides and characterized their binding abilities by surface plasmon resonance, fluorescence and electrospray ionization mass spectrometry assays. These experiments showed the designed binders to recognize their target with micromolar affinity. We finally discuss the obtained results in the light of further improvement in the ex-novo optimization of peptide based binders.

  15. A Pareto Algorithm for Efficient De Novo Design of Multi-functional Molecules.

    Science.gov (United States)

    Daeyaert, Frits; Deem, Micheal W

    2017-01-01

    We have introduced a Pareto sorting algorithm into Synopsis, a de novo design program that generates synthesizable molecules with desirable properties. We give a detailed description of the algorithm and illustrate its working in 2 different de novo design settings: the design of putative dual and selective FGFR and VEGFR inhibitors, and the successful design of organic structure determining agents (OSDAs) for the synthesis of zeolites. We show that the introduction of Pareto sorting not only enables the simultaneous optimization of multiple properties but also greatly improves the performance of the algorithm to generate molecules with hard-to-meet constraints. This in turn allows us to suggest approaches to address the problem of false positive hits in de novo structure based drug design by introducing structural and physicochemical constraints in the designed molecules, and by forcing essential interactions between these molecules and their target receptor. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  16. Design of a minimal protein oligomerization domain by a structural approach.

    Science.gov (United States)

    Burkhard, P; Meier, M; Lustig, A

    2000-12-01

    Because of the simplicity and regularity of the alpha-helical coiled coil relative to other structural motifs, it can be conveniently used to clarify the molecular interactions responsible for protein folding and stability. Here we describe the de novo design and characterization of a two heptad-repeat peptide stabilized by a complex network of inter- and intrahelical salt bridges. Circular dichroism spectroscopy and analytical ultracentrifugation show that this peptide is highly alpha-helical and 100% dimeric tinder physiological buffer conditions. Interestingly, the peptide was shown to switch its oligomerization state from a dimer to a trimer upon increasing ionic strength. The correctness of the rational design principles used here is supported by details of the atomic structure of the peptide deduced from X-ray crystallography. The structure of the peptide shows that it is not a molten globule but assumes a unique, native-like conformation. This de novo peptide thus represents an attractive model system for the design of a molecular recognition system.

  17. Peptide array-based interaction assay of solid-bound peptides and anchorage-dependant cells and its effectiveness in cell-adhesive peptide design.

    Science.gov (United States)

    Kato, Ryuji; Kaga, Chiaki; Kunimatsu, Mitoshi; Kobayashi, Takeshi; Honda, Hiroyuki

    2006-06-01

    Peptide array, the designable peptide library covalently synthesized on cellulose support, was applied to assay peptide-cell interaction, between solid-bound peptides and anchorage-dependant cells, to study objective peptide design. As a model case, cell-adhesive peptides that could enhance cell growth as tissue engineering scaffold material, was studied. On the peptide array, the relative cell-adhesion ratio of NIH/3T3 cells was 2.5-fold higher on the RGDS (Arg-Gly-Asp-Ser) peptide spot as compared to the spot with no peptide, thus indicating integrin-mediated peptide-cell interaction. Such strong cell adhesion mediated by the RGDS peptide was easily disrupted by single residue substitution on the peptide array, thus indicating that the sequence recognition accuracy of cells was strictly conserved in our optimized scheme. The observed cellular morphological extension with active actin stress-fiber on the RGD motif-containing peptide supported our strategy that peptide array-based interaction assay of solid-bound peptide and anchorage-dependant cells (PIASPAC) could provide quantitative data on biological peptide-cell interaction. The analysis of 180 peptides obtained from fibronectin type III domain (no. 1447-1629) yielded 18 novel cell-adhesive peptides without the RGD motif. Taken together with the novel candidates, representative rules of ineffective amino acid usage were obtained from non-effective candidate sequences for the effective designing of cell-adhesive peptides. On comparing the amino acid usage of the top 20 and last 20 peptides from the 180 peptides, the following four brief design rules were indicated: (i) Arg or Lys of positively charged amino acids (except His) could enhance cell adhesion, (ii) small hydrophilic amino acids are favored in cell-adhesion peptides, (iii) negatively charged amino acids and small amino acids (except Gly) could reduce cell adhesion, and (iv) Cys and Met could be excluded from the sequence combination since they have

  18. Discovery of Novel Antimicrobial Peptides from Varanus komodoensis (Komodo Dragon) by Large-Scale Analyses and De-Novo-Assisted Sequencing Using Electron-Transfer Dissociation Mass Spectrometry.

    Science.gov (United States)

    Bishop, Barney M; Juba, Melanie L; Russo, Paul S; Devine, Megan; Barksdale, Stephanie M; Scott, Shaylyn; Settlage, Robert; Michalak, Pawel; Gupta, Kajal; Vliet, Kent; Schnur, Joel M; van Hoek, Monique L

    2017-04-07

    Komodo dragons are the largest living lizards and are the apex predators in their environs. They endure numerous strains of pathogenic bacteria in their saliva and recover from wounds inflicted by other dragons, reflecting the inherent robustness of their innate immune defense. We have employed a custom bioprospecting approach combining partial de novo peptide sequencing with transcriptome assembly to identify cationic antimicrobial peptides from Komodo dragon plasma. Through these analyses, we identified 48 novel potential cationic antimicrobial peptides. All but one of the identified peptides were derived from histone proteins. The antimicrobial effectiveness of eight of these peptides was evaluated against Pseudomonas aeruginosa (ATCC 9027) and Staphylococcus aureus (ATCC 25923), with seven peptides exhibiting antimicrobial activity against both microbes and one only showing significant potency against P. aeruginosa. This study demonstrates the power and promise of our bioprospecting approach to cationic antimicrobial peptide discovery, and it reveals the presence of a plethora of novel histone-derived antimicrobial peptides in the plasma of the Komodo dragon. These findings may have broader implications regarding the role that intact histones and histone-derived peptides play in defending the host from infection. Data are available via ProteomeXChange with identifier PXD005043.

  19. Comprehensive computational design of ordered peptide macrocycles

    Science.gov (United States)

    Hosseinzadeh, Parisa; Bhardwaj, Gaurav; Mulligan, Vikram Khipple; Shortridge, Matthew D.; Craven, Timothy W.; Pardo-Avila, Fátima; Rettie, Stephen A.; Kim, David E.; Silva, Daniel-Adriano; Ibrahim, Yehia M.; Webb, Ian K.; Cort, John R.; Adkins, Joshua N.; Varani, Gabriele; Baker, David

    2018-01-01

    Mixed-chirality peptide macrocycles such as cyclosporine are among the most potent therapeutics identified to date, but there is currently no way to systematically search the structural space spanned by such compounds. Natural proteins do not provide a useful guide: Peptide macrocycles lack regular secondary structures and hydrophobic cores, and can contain local structures not accessible with L-amino acids. Here, we enumerate the stable structures that can be adopted by macrocyclic peptides composed of L- and D-amino acids by near-exhaustive backbone sampling followed by sequence design and energy landscape calculations. We identify more than 200 designs predicted to fold into single stable structures, many times more than the number of currently available unbound peptide macrocycle structures. Nuclear magnetic resonance structures of 9 of 12 designed 7- to 10-residue macrocycles, and three 11- to 14-residue bicyclic designs, are close to the computational models. Our results provide a nearly complete coverage of the rich space of structures possible for short peptide macrocycles and vastly increase the available starting scaffolds for both rational drug design and library selection methods. PMID:29242347

  20. Membrane interaction and secondary structure of de novo designed arginine-and tryptophan peptides with dual function

    KAUST Repository

    Rydberg, Hanna A.; Carlsson, Nils; Nordé n, Bengt

    2012-01-01

    of arg/trp peptides and investigated how the position and number of tryptophans affect cellular uptake. Here we explore the antimicrobial properties and the interaction with lipid model membranes of these peptides, using minimal inhibitory concentrations

  1. Designing Antibacterial Peptides with Enhanced Killing Kinetics

    Directory of Open Access Journals (Sweden)

    Faiza H. Waghu

    2018-02-01

    Full Text Available Antimicrobial peptides (AMPs are gaining attention as substitutes for antibiotics in order to combat the risk posed by multi-drug resistant pathogens. Several research groups are engaged in design of potent anti-infective agents using natural AMPs as templates. In this study, a library of peptides with high sequence similarity to Myeloid Antimicrobial Peptide (MAP family were screened using popular online prediction algorithms. These peptide variants were designed in a manner to retain the conserved residues within the MAP family. The prediction algorithms were found to effectively classify peptides based on their antimicrobial nature. In order to improve the activity of the identified peptides, molecular dynamics (MD simulations, using bilayer and micellar systems could be used to design and predict effect of residue substitution on membranes of microbial and mammalian cells. The inference from MD simulation studies well corroborated with the wet-lab observations indicating that MD-guided rational design could lead to discovery of potent AMPs. The effect of the residue substitution on membrane activity was studied in greater detail using killing kinetic analysis. Killing kinetics studies on Gram-positive, negative and human erythrocytes indicated that a single residue change has a drastic effect on the potency of AMPs. An interesting outcome was a switch from monophasic to biphasic death rate constant of Staphylococcus aureus due to a single residue mutation in the peptide.

  2. De novo design, synthesis and characterisation of MP3, a new catalytic four-helix bundle hemeprotein.

    Science.gov (United States)

    Faiella, Marina; Maglio, Ornella; Nastri, Flavia; Lombardi, Angela; Lista, Liliana; Hagen, Wilfred R; Pavone, Vincenzo

    2012-12-07

    A new artificial metalloenzyme, MP3 (MiniPeroxidase 3), designed by combining the excellent structural properties of four-helix bundle protein scaffolds with the activity of natural peroxidases, was synthesised and characterised. This new hemeprotein model was developed by covalently linking the deuteroporphyrin to two peptide chains of different compositions to obtain an asymmetric helix-loop-helix/heme/helix-loop-helix sandwich arrangement, characterised by 1) a His residue on one chain that acts as an axial ligand to the iron ion; 2) a vacant distal site that is able to accommodate exogenous ligands or substrates; and 3) an Arg residue in the distal site that should assist in hydrogen peroxide activation to give an HRP-like catalytic process. MP3 was synthesised and characterised as its iron complex. CD measurements revealed the high helix-forming propensity of the peptide, confirming the appropriateness of the model procedure; UV/Vis, MCD and EPR experiments gave insights into the coordination geometry and the spin state of the metal. Kinetic experiments showed that Fe(III)-MP3 possesses peroxidase-like activity comparable to R38A-hHRP, highlighting the possibility of mimicking the functional features of natural enzymes. The synergistic application of de novo design methods, synthetic procedures, and spectroscopic characterisation, described herein, demonstrates a method by which to implement and optimise catalytic activity for an enzyme mimetic. Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  3. Adopting De Novo Programming Approach on IC Design Service Firms Resources Integration

    Directory of Open Access Journals (Sweden)

    James K. C. Chen

    2014-01-01

    Full Text Available The semiconductor industry has very important position in computer industry, ICT field, and new electronic technology developing. The IC design service is one of key factor of semiconductor industry development. There are more than 365 IC design service firms have been established around Hsinchu Science Park in Taiwan. Building an efficient planning model for IC design service firm resources integrating is very interest issue. This study aims to construct a planning model for IC design service firm implementation resources integration. This study uses the De Novo programming as an approach of criteria alternative to achieve optimal resource allocation on IC design firm. Results show the IC design service firm should conduct open innovation concept and utilizes design outsourcing obtains cost down and enhance IC design service business performance. This plan model of De Novo programming is not only for IC design service firm and also can apply to the other industrial implementation strategic alliance/integrating resource. This plan model is a universal model for the others industries field.

  4. Comprehensive computational design of ordered peptide macrocycles

    Energy Technology Data Exchange (ETDEWEB)

    Hosseinzadeh, Parisa; Bhardwaj, Gaurav; Mulligan, Vikram K.; Shortridge, Matthew D.; Craven, Timothy W.; Pardo-Avila, Fatima; Rettie, Stephan A.; Kim, David E.; Silva, Daniel A.; Ibrahim, Yehia M.; Webb, Ian K.; Cort, John R.; Adkins, Joshua N.; Varani, Gabriele; Baker, David

    2017-12-14

    Mixed chirality peptide macrocycles such as cyclosporine are among the most potent therapeutics identified to-date, but there is currently no way to systematically search through the structural space spanned by such compounds for new drug candidates. Natural proteins do not provide a useful guide: peptide macrocycles lack regular secondary structures and hydrophobic cores and have different backbone torsional constraints. Hence the development of new peptide macrocycles has been approached by modifying natural products or using library selection methods; the former is limited by the small number of known structures, and the latter by the limited size and diversity accessible through library-based methods. To overcome these limitations, here we enumerate the stable structures that can be adopted by macrocyclic peptides composed of L and D amino acids. We identify more than 200 designs predicted to fold into single stable structures, many times more than the number of currently available unbound peptide macrocycle structures. We synthesize and characterize by NMR twelve 7-10 residue macrocycles, 9 of which have structures very close to the design models in solution. NMR structures of three 11-14 residue bicyclic designs are also very close to the computational models. Our results provide a nearly complete coverage of the rich space of structures possible for short peptide based macrocycles unparalleled for other molecular systems, and vastly increase the available starting scaffolds for both rational drug design and library selection methods.

  5. Designed graphene-peptide nanocomposites for biosensor applications: A review

    International Nuclear Information System (INIS)

    Wang, Li; Zhang, Yujie; Wu, Aiguo; Wei, Gang

    2017-01-01

    The modification of graphene with biomacromolecules like DNA, protein, peptide, and others extends the potential applications of graphene materials in various fields. The bound biomacromolecules could improve the biocompatibility and bio-recognition ability of graphene-based nanocomposites, therefore could greatly enhance their biosensing performances on both selectivity and sensitivity. In this review, we presented a comprehensive introduction and discussion on recent advance in the synthesis and biosensor applications of graphene-peptide nanocomposites. The biofunctionalization of graphene with specifically designed peptides, and the synthesis strategies of graphene-peptide (monomer, nanofibrils, and nanotubes) nanocomposites were demonstrated. On the other hand, the fabrication of graphene-peptide nanocomposite based biosensor architectures for electrochemical, fluorescent, electronic, and spectroscopic biosensing were further presented. This review includes nearly all the studies on the fabrication and applications of graphene-peptide based biosensors recently, which will promote the future developments of graphene-based biosensors in biomedical detection and environmental analysis. - Highlights: • A comprehensive review on the fabrication and application of graphene-peptide nanocomposites was presented. • The design of peptide sequences for biofunctionalization of various graphene materials was presented. • Multi-strategies on the fabrication of biosensors with graphene-peptide nanocomposites were discussed. • Designed graphene-peptide nanocomposites showed wide biosensor applications.

  6. Antimicrobial peptides design by evolutionary multiobjective optimization.

    Directory of Open Access Journals (Sweden)

    Giuseppe Maccari

    Full Text Available Antimicrobial peptides (AMPs are an abundant and wide class of molecules produced by many tissues and cell types in a variety of mammals, plant and animal species. Linear alpha-helical antimicrobial peptides are among the most widespread membrane-disruptive AMPs in nature, representing a particularly successful structural arrangement in innate defense. Recently, AMPs have received increasing attention as potential therapeutic agents, owing to their broad activity spectrum and their reduced tendency to induce resistance. The introduction of non-natural amino acids will be a key requisite in order to contrast host resistance and increase compound's life. In this work, the possibility to design novel AMP sequences with non-natural amino acids was achieved through a flexible computational approach, based on chemophysical profiles of peptide sequences. Quantitative structure-activity relationship (QSAR descriptors were employed to code each peptide and train two statistical models in order to account for structural and functional properties of alpha-helical amphipathic AMPs. These models were then used as fitness functions for a multi-objective evolutional algorithm, together with a set of constraints for the design of a series of candidate AMPs. Two ab-initio natural peptides were synthesized and experimentally validated for antimicrobial activity, together with a series of control peptides. Furthermore, a well-known Cecropin-Mellitin alpha helical antimicrobial hybrid (CM18 was optimized by shortening its amino acid sequence while maintaining its activity and a peptide with non-natural amino acids was designed and tested, demonstrating the higher activity achievable with artificial residues.

  7. PeptideNavigator: An interactive tool for exploring large and complex data sets generated during peptide-based drug design projects.

    Science.gov (United States)

    Diller, Kyle I; Bayden, Alexander S; Audie, Joseph; Diller, David J

    2018-01-01

    There is growing interest in peptide-based drug design and discovery. Due to their relatively large size, polymeric nature, and chemical complexity, the design of peptide-based drugs presents an interesting "big data" challenge. Here, we describe an interactive computational environment, PeptideNavigator, for naturally exploring the tremendous amount of information generated during a peptide drug design project. The purpose of PeptideNavigator is the presentation of large and complex experimental and computational data sets, particularly 3D data, so as to enable multidisciplinary scientists to make optimal decisions during a peptide drug discovery project. PeptideNavigator provides users with numerous viewing options, such as scatter plots, sequence views, and sequence frequency diagrams. These views allow for the collective visualization and exploration of many peptides and their properties, ultimately enabling the user to focus on a small number of peptides of interest. To drill down into the details of individual peptides, PeptideNavigator provides users with a Ramachandran plot viewer and a fully featured 3D visualization tool. Each view is linked, allowing the user to seamlessly navigate from collective views of large peptide data sets to the details of individual peptides with promising property profiles. Two case studies, based on MHC-1A activating peptides and MDM2 scaffold design, are presented to demonstrate the utility of PeptideNavigator in the context of disparate peptide-design projects. Copyright © 2017 Elsevier Ltd. All rights reserved.

  8. Design of non-aggregating variants of Aβ peptide

    Energy Technology Data Exchange (ETDEWEB)

    Caine, Joanne M., E-mail: jo.caine@csiro.au [CSIRO Materials Science and Engineering, 343 Royal Parade, Parkville, Victoria 3052 (Australia); Preventative Health Flagship, 343 Royal Parade, Parkville, Victoria 3052 (Australia); CRC for Mental Health, Level 2, 161 Barry Street, Carlton South, Victoria 3053 (Australia); Churches, Quentin; Waddington, Lynne [CSIRO Materials Science and Engineering, 343 Royal Parade, Parkville, Victoria 3052 (Australia); Preventative Health Flagship, 343 Royal Parade, Parkville, Victoria 3052 (Australia); Nigro, Julie; Breheney, Kerry [CSIRO Materials Science and Engineering, 343 Royal Parade, Parkville, Victoria 3052 (Australia); Preventative Health Flagship, 343 Royal Parade, Parkville, Victoria 3052 (Australia); CRC for Mental Health, Level 2, 161 Barry Street, Carlton South, Victoria 3053 (Australia); Masters, Colin L. [CRC for Mental Health, Level 2, 161 Barry Street, Carlton South, Victoria 3053 (Australia); Florey Institute for Neuroscience and Mental Health, 30 Royal Parade, Parkville, Victoria 3052 (Australia); Nuttall, Stewart D. [CSIRO Materials Science and Engineering, 343 Royal Parade, Parkville, Victoria 3052 (Australia); Preventative Health Flagship, 343 Royal Parade, Parkville, Victoria 3052 (Australia); CRC for Mental Health, Level 2, 161 Barry Street, Carlton South, Victoria 3053 (Australia); Streltsov, Victor A., E-mail: victor.streltsov@csiro.au [CSIRO Materials Science and Engineering, 343 Royal Parade, Parkville, Victoria 3052 (Australia); Preventative Health Flagship, 343 Royal Parade, Parkville, Victoria 3052 (Australia); CRC for Mental Health, Level 2, 161 Barry Street, Carlton South, Victoria 3053 (Australia)

    2014-10-24

    Highlights: • Non-aggregating, non-toxic variants of Aβ peptide were designed using Aβ structure. • Mutations reduce aggregation by stabilising Aβ into small non-toxic oligomers. • Identification of these residues will assist the design of future therapeutic peptides. - Abstract: Self association of the amyloid-β (Aβ{sub 42}) peptide into oligomers, high molecular weight forms, fibrils and ultimately neuritic plaques, has been correlated with progressive cognitive decline in Alzheimer’s disease. Thus, insights into the drivers of the aggregation pathway have the capacity to significantly contribute to our understanding of disease mechanism. Functional assays and a three-dimensional crystal structure of the P3 amyloidogenic region 18–41 of Aβ were used to identify residues important in self-association and to design novel non-aggregating variants of the peptide. Biophysical studies (gel filtration, SDS–PAGE, dynamic light scattering, thioflavin T assay, and electron microscopy) demonstrate that in contrast to wild type Aβ these targeted mutations lose the ability to self-associate. Loss of aggregation also correlates with reduced neuronal toxicity. Our results highlight residues and regions of the Aβ peptide important for future targeting agents aimed at the amelioration of Alzheimer’s disease.

  9. Simultaneous inhibition of key growth pathways in melanoma cells and tumor regression by a designed bidentate constrained helical peptide.

    Science.gov (United States)

    Dhar, Amlanjyoti; Mallick, Shampa; Ghosh, Piya; Maiti, Atanu; Ahmed, Israr; Bhattacharya, Seemana; Mandal, Tapashi; Manna, Asit; Roy, Koushik; Singh, Sandeep; Nayak, Dipak Kumar; Wilder, Paul T; Markowitz, Joseph; Weber, David; Ghosh, Mrinal K; Chattopadhyay, Samit; Guha, Rajdeep; Konar, Aditya; Bandyopadhyay, Santu; Roy, Siddhartha

    2014-07-01

    Protein-protein interactions are part of a large number of signaling networks and potential targets for drug development. However, discovering molecules that can specifically inhibit such interactions is a major challenge. S100B, a calcium-regulated protein, plays a crucial role in the proliferation of melanoma cells through protein-protein interactions. In this article, we report the design and development of a bidentate conformationally constrained peptide against dimeric S100B based on a natural tight-binding peptide, TRTK-12. The helical conformation of the peptide was constrained by the substitution of α-amino isobutyric acid--an amino acid having high helical propensity--in positions which do not interact with S100B. A branched bidentate version of the peptide was bound to S100B tightly with a dissociation constant of 8 nM. When conjugated to a cell-penetrating peptide, it caused growth inhibition and rapid apoptosis in melanoma cells. The molecule exerts antiproliferative action through simultaneous inhibition of key growth pathways, including reactivation of wild-type p53 and inhibition of Akt and STAT3 phosphorylation. The apoptosis induced by the bidentate constrained helix is caused by direct migration of p53 to mitochondria. At moderate intravenous dose, the peptide completely inhibits melanoma growth in a mouse model without any significant observable toxicity. The specificity was shown by lack of ability of a double mutant peptide to cause tumor regression at the same dose level. The methodology described here for direct protein-protein interaction inhibition may be effective for rapid development of inhibitors against relatively weak protein-protein interactions for de novo drug development. © 2014 Wiley Periodicals, Inc.

  10. Massively parallel de novo protein design for targeted therapeutics

    KAUST Repository

    Chevalier, Aaron

    2017-09-26

    De novo protein design holds promise for creating small stable proteins with shapes customized to bind therapeutic targets. We describe a massively parallel approach for designing, manufacturing and screening mini-protein binders, integrating large-scale computational design, oligonucleotide synthesis, yeast display screening and next-generation sequencing. We designed and tested 22,660 mini-proteins of 37-43 residues that target influenza haemagglutinin and botulinum neurotoxin B, along with 6,286 control sequences to probe contributions to folding and binding, and identified 2,618 high-affinity binders. Comparison of the binding and non-binding design sets, which are two orders of magnitude larger than any previously investigated, enabled the evaluation and improvement of the computational model. Biophysical characterization of a subset of the binder designs showed that they are extremely stable and, unlike antibodies, do not lose activity after exposure to high temperatures. The designs elicit little or no immune response and provide potent prophylactic and therapeutic protection against influenza, even after extensive repeated dosing.

  11. Massively parallel de novo protein design for targeted therapeutics

    KAUST Repository

    Chevalier, Aaron; Silva, Daniel-Adriano; Rocklin, Gabriel J.; Hicks, Derrick R.; Vergara, Renan; Murapa, Patience; Bernard, Steffen M.; Zhang, Lu; Lam, Kwok-Ho; Yao, Guorui; Bahl, Christopher D.; Miyashita, Shin-Ichiro; Goreshnik, Inna; Fuller, James T.; Koday, Merika T.; Jenkins, Cody M.; Colvin, Tom; Carter, Lauren; Bohn, Alan; Bryan, Cassie M.; Ferná ndez-Velasco, D. Alejandro; Stewart, Lance; Dong, Min; Huang, Xuhui; Jin, Rongsheng; Wilson, Ian A.; Fuller, Deborah H.; Baker, David

    2017-01-01

    De novo protein design holds promise for creating small stable proteins with shapes customized to bind therapeutic targets. We describe a massively parallel approach for designing, manufacturing and screening mini-protein binders, integrating large-scale computational design, oligonucleotide synthesis, yeast display screening and next-generation sequencing. We designed and tested 22,660 mini-proteins of 37-43 residues that target influenza haemagglutinin and botulinum neurotoxin B, along with 6,286 control sequences to probe contributions to folding and binding, and identified 2,618 high-affinity binders. Comparison of the binding and non-binding design sets, which are two orders of magnitude larger than any previously investigated, enabled the evaluation and improvement of the computational model. Biophysical characterization of a subset of the binder designs showed that they are extremely stable and, unlike antibodies, do not lose activity after exposure to high temperatures. The designs elicit little or no immune response and provide potent prophylactic and therapeutic protection against influenza, even after extensive repeated dosing.

  12. Massively parallel de novo protein design for targeted therapeutics

    Science.gov (United States)

    Chevalier, Aaron; Silva, Daniel-Adriano; Rocklin, Gabriel J.; Hicks, Derrick R.; Vergara, Renan; Murapa, Patience; Bernard, Steffen M.; Zhang, Lu; Lam, Kwok-Ho; Yao, Guorui; Bahl, Christopher D.; Miyashita, Shin-Ichiro; Goreshnik, Inna; Fuller, James T.; Koday, Merika T.; Jenkins, Cody M.; Colvin, Tom; Carter, Lauren; Bohn, Alan; Bryan, Cassie M.; Fernández-Velasco, D. Alejandro; Stewart, Lance; Dong, Min; Huang, Xuhui; Jin, Rongsheng; Wilson, Ian A.; Fuller, Deborah H.; Baker, David

    2018-01-01

    De novo protein design holds promise for creating small stable proteins with shapes customized to bind therapeutic targets. We describe a massively parallel approach for designing, manufacturing and screening mini-protein binders, integrating large-scale computational design, oligonucleotide synthesis, yeast display screening and next-generation sequencing. We designed and tested 22,660 mini-proteins of 37–43 residues that target influenza haemagglutinin and botulinum neurotoxin B, along with 6,286 control sequences to probe contributions to folding and binding, and identified 2,618 high-affinity binders. Comparison of the binding and non-binding design sets, which are two orders of magnitude larger than any previously investigated, enabled the evaluation and improvement of the computational model. Biophysical characterization of a subset of the binder designs showed that they are extremely stable and, unlike antibodies, do not lose activity after exposure to high temperatures. The designs elicit little or no immune response and provide potent prophylactic and therapeutic protection against influenza, even after extensive repeated dosing. PMID:28953867

  13. RECENT ADVANCES TOWARDS THE RATIONAL DESIGN OF PEPTIDE DRUGS

    OpenAIRE

    YEŞİLADA, Akgül; ÖZKANLI, Fügen

    2004-01-01

    In this review, after a short introduction to definition and physiological roles of regulatory peptides, problems faced during the development of peptide drugs, studies directed to solve these problems and rational design of peptide drugs with special emphesis on peptidomimetics are mentioned

  14. Designed β-Boomerang Antiendotoxic and Antimicrobial Peptides

    Science.gov (United States)

    Bhunia, Anirban; Mohanram, Harini; Domadia, Prerna N.; Torres, Jaume; Bhattacharjya, Surajit

    2009-01-01

    Lipopolysaccharide (LPS), an integral part of the outer membrane of Gram-negative bacteria, is involved in a variety of biological processes including inflammation, septic shock, and resistance to host-defense molecules. LPS also provides an environment for folding of outer membrane proteins. In this work, we describe the structure-activity correlation of a series of 12-residue peptides in LPS. NMR structures of the peptides derived in complex with LPS reveal boomerang-like β-strand conformations that are stabilized by intimate packing between the two aromatic residues located at the 4 and 9 positions. This structural feature renders these peptides with a high ability to neutralize endotoxicity, >80% at 10 nm concentration, of LPS. Replacements of these aromatic residues either with Ala or with Leu destabilizes the boomerang structure with the concomitant loss of antiendotoxic and antimicrobial activities. Furthermore, the aromatic packing stabilizing the β-boomerang structure in LPS is found to be maintained even in a truncated octapeptide, defining a structured LPS binding motif. The mode of action of the active designed peptides correlates well with their ability to perturb LPS micelle structures. Fourier transform infrared spectroscopy studies of the peptides delineate β-type conformations and immobilization of phosphate head groups of LPS. Trp fluorescence studies demonstrated selective interactions with LPS and the depth of insertion into the LPS bilayer. Our results demonstrate the requirement of LPS-specific structures of peptides for endotoxin neutralizations. In addition, we propose that structures of these peptides may be employed to design proteins for the outer membrane. PMID:19520860

  15. Database-Guided Discovery of Potent Peptides to Combat HIV-1 or Superbugs

    Directory of Open Access Journals (Sweden)

    Guangshun Wang

    2013-05-01

    Full Text Available Antimicrobial peptides (AMPs, small host defense proteins, are indispensable for the protection of multicellular organisms such as plants and animals from infection. The number of AMPs discovered per year increased steadily since the 1980s. Over 2,000 natural AMPs from bacteria, protozoa, fungi, plants, and animals have been registered into the antimicrobial peptide database (APD. The majority of these AMPs (>86% possess 11–50 amino acids with a net charge from 0 to +7 and hydrophobic percentages between 31–70%. This article summarizes peptide discovery on the basis of the APD. The major methods are the linguistic model, database screening, de novo design, and template-based design. Using these methods, we identified various potent peptides against human immunodeficiency virus type 1 (HIV-1 or methicillin-resistant Staphylococcus aureus (MRSA. While the stepwise designed anti-HIV peptide is disulfide-linked and rich in arginines, the ab initio designed anti-MRSA peptide is linear and rich in leucines. Thus, there are different requirements for antiviral and antibacterial peptides, which could kill pathogens via different molecular targets. The biased amino acid composition in the database-designed peptides, or natural peptides such as θ-defensins, requires the use of the improved two-dimensional NMR method for structural determination to avoid the publication of misleading structure and dynamics. In the case of human cathelicidin LL-37, structural determination requires 3D NMR techniques. The high-quality structure of LL-37 provides a solid basis for understanding its interactions with membranes of bacteria and other pathogens. In conclusion, the APD database is a comprehensive platform for storing, classifying, searching, predicting, and designing potent peptides against pathogenic bacteria, viruses, fungi, parasites, and cancer cells.

  16. PinaColada: peptide-inhibitor ant colony ad-hoc design algorithm.

    Science.gov (United States)

    Zaidman, Daniel; Wolfson, Haim J

    2016-08-01

    Design of protein-protein interaction (PPI) inhibitors is a major challenge in Structural Bioinformatics. Peptides, especially short ones (5-15 amino acid long), are natural candidates for inhibition of protein-protein complexes due to several attractive features such as high structural compatibility with the protein binding site (mimicking the surface of one of the proteins), small size and the ability to form strong hotspot binding connections with the protein surface. Efficient rational peptide design is still a major challenge in computer aided drug design, due to the huge space of possible sequences, which is exponential in the length of the peptide, and the high flexibility of peptide conformations. In this article we present PinaColada, a novel computational method for the design of peptide inhibitors for protein-protein interactions. We employ a version of the ant colony optimization heuristic, which is used to explore the exponential space ([Formula: see text]) of length n peptide sequences, in combination with our fast robotics motivated PepCrawler algorithm, which explores the conformational space for each candidate sequence. PinaColada is being run in parallel, on a DELL PowerEdge 2.8 GHZ computer with 20 cores and 256 GB memory, and takes up to 24 h to design a peptide of 5-15 amino acids length. An online server available at: http://bioinfo3d.cs.tau.ac.il/PinaColada/. danielza@post.tau.ac.il; wolfson@tau.ac.il. © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  17. Machine learning-enabled discovery and design of membrane-active peptides.

    Science.gov (United States)

    Lee, Ernest Y; Wong, Gerard C L; Ferguson, Andrew L

    2017-07-08

    Antimicrobial peptides are a class of membrane-active peptides that form a critical component of innate host immunity and possess a diversity of sequence and structure. Machine learning approaches have been profitably employed to efficiently screen sequence space and guide experiment towards promising candidates with high putative activity. In this mini-review, we provide an introduction to antimicrobial peptides and summarize recent advances in machine learning-enabled antimicrobial peptide discovery and design with a focus on a recent work Lee et al. Proc. Natl. Acad. Sci. USA 2016;113(48):13588-13593. This study reports the development of a support vector machine classifier to aid in the design of membrane active peptides. We use this model to discover membrane activity as a multiplexed function in diverse peptide families and provide interpretable understanding of the physicochemical properties and mechanisms governing membrane activity. Experimental validation of the classifier reveals it to have learned membrane activity as a unifying signature of antimicrobial peptides with diverse modes of action. Some of the discriminating rules by which it performs classification are in line with existing "human learned" understanding, but it also unveils new previously unknown determinants and multidimensional couplings governing membrane activity. Integrating machine learning with targeted experimentation can guide both antimicrobial peptide discovery and design and new understanding of the properties and mechanisms underpinning their modes of action. Copyright © 2017 Elsevier Ltd. All rights reserved.

  18. Generation of Hybrid Peptide-Silver Nanoparticles for Antibacterial and Antifouling Applications

    KAUST Repository

    Seferji, Kholoud

    2018-01-01

    and antifouling agents. Our innovative antibacterial agents are hybrid peptide silver nanoparticles (CH-01-AgNPs) that are created de novo and in situ from a silver nitrate solution (AgNO3) in the presence of ultrashort self-assembling peptides compounds

  19. Designing of peptides with desired half-life in intestine-like environment

    KAUST Repository

    Sharma, Arun

    2014-08-20

    Background: In past, a number of peptides have been reported to possess highly diverse properties ranging from cell penetrating, tumor homing, anticancer, anti-hypertensive, antiviral to antimicrobials. Owing to their excellent specificity, low-toxicity, rich chemical diversity and availability from natural sources, FDA has successfully approved a number of peptide-based drugs and several are in various stages of drug development. Though peptides are proven good drug candidates, their usage is still hindered mainly because of their high susceptibility towards proteases degradation. We have developed an in silico method to predict the half-life of peptides in intestine-like environment and to design better peptides having optimized physicochemical properties and half-life.Results: In this study, we have used 10mer (HL10) and 16mer (HL16) peptides dataset to develop prediction models for peptide half-life in intestine-like environment. First, SVM based models were developed on HL10 dataset which achieved maximum correlation R/R2 of 0.57/0.32, 0.68/0.46, and 0.69/0.47 using amino acid, dipeptide and tripeptide composition, respectively. Secondly, models developed on HL16 dataset showed maximum R/R2 of 0.91/0.82, 0.90/0.39, and 0.90/0.31 using amino acid, dipeptide and tripeptide composition, respectively. Furthermore, models that were developed on selected features, achieved a correlation (R) of 0.70 and 0.98 on HL10 and HL16 dataset, respectively. Preliminary analysis suggests the role of charged residue and amino acid size in peptide half-life/stability. Based on above models, we have developed a web server named HLP (Half Life Prediction), for predicting and designing peptides with desired half-life. The web server provides three facilities; i) half-life prediction, ii) physicochemical properties calculation and iii) designing mutant peptides.Conclusion: In summary, this study describes a web server \\'HLP\\' that has been developed for assisting scientific

  20. Tuning Liposome Membrane Permeability by Competitive Peptide Dimerization and Partitioning-Folding Interactions Regulated by Proteolytic Activity

    Science.gov (United States)

    Lim, Seng Koon; Sandén, Camilla; Selegård, Robert; Liedberg, Bo; Aili, Daniel

    2016-02-01

    Membrane active peptides are of large interest for development of drug delivery vehicles and therapeutics for treatment of multiple drug resistant infections. Lack of specificity can be detrimental and finding routes to tune specificity and activity of membrane active peptides is vital for improving their therapeutic efficacy and minimize harmful side effects. We describe a de novo designed membrane active peptide that partition into lipid membranes only when specifically and covalently anchored to the membrane, resulting in pore-formation. Dimerization with a complementary peptide efficiently inhibits formation of pores. The effect can be regulated by proteolytic digestion of the inhibitory peptide by the matrix metalloproteinase MMP-7, an enzyme upregulated in many malignant tumors. This system thus provides a precise and specific route for tuning the permeability of lipid membranes and a novel strategy for development of recognition based membrane active peptides and indirect enzymatically controlled release of liposomal cargo.

  1. The discovery of 9/8-ribbons, β/γ-peptides with curved shapes governed by a combined configuration-conformation code.

    Science.gov (United States)

    Grison, Claire M; Robin, Sylvie; Aitken, David J

    2015-11-21

    The de novo design of a β/γ-peptidic foldamer motif has led to the discovery of an unprecedented 9/8-ribbon featuring an uninterrupted alternating C9/C8 hydrogen-bonding network. The ribbons adopt partially curved topologies determined synchronistically by the β-residue configuration and the γ-residue conformation sets.

  2. Designer interface peptide grafts target estrogen receptor alpha dimerization

    International Nuclear Information System (INIS)

    Chakraborty, S.; Asare, B.K.; Biswas, P.K.; Rajnarayanan, R.V.

    2016-01-01

    The nuclear transcription factor estrogen receptor alpha (ERα), triggered by its cognate ligand estrogen, regulates a variety of cellular signaling events. ERα is expressed in 70% of breast cancers and is a widely validated target for anti-breast cancer drug discovery. Administration of anti-estrogen to block estrogen receptor activation is still a viable anti-breast cancer treatment option but anti-estrogen resistance has been a significant bottle-neck. Dimerization of estrogen receptor is required for ER activation. Blocking ERα dimerization is therefore a complementary and alternative strategy to combat anti-estrogen resistance. Dimer interface peptide “I-box” derived from ER residues 503–518 specifically blocks ER dimerization. Recently using a comprehensive molecular simulation we studied the interaction dynamics of ERα LBDs in a homo-dimer. Based on this study, we identified three interface recognition peptide motifs LDKITDT (ERα residues 479–485), LQQQHQRLAQ (residues 497–506), and LSHIRHMSNK (residues 511–520) and reported the suitability of using LQQQHQRLAQ (ER 497–506) as a template to design inhibitors of ERα dimerization. Stability and self-aggregation of peptide based therapeutics poses a significant bottle-neck to proceed further. In this study utilizing peptide grafted to preserve their pharmacophoric recognition motif and assessed their stability and potential to block ERα mediated activity in silico and in vitro. The Grafted peptides blocked ERα mediated cell proliferation and viability of breast cancer cells but did not alter their apoptotic fate. We believe the structural clues identified in this study can be used to identify novel peptidometics and small molecules that specifically target ER dimer interface generating a new breed of anti-cancer agents. - Highlights: • Designer peptide grafts retain core molecular recognition motif during MD simulations. • Designer peptide grafts with Poly-ALA helix form stable

  3. Designer interface peptide grafts target estrogen receptor alpha dimerization

    Energy Technology Data Exchange (ETDEWEB)

    Chakraborty, S. [Laboratory of Computational Biophysics & Bioengineering, Department of Physics, Tougaloo College, Tougaloo, MS 39174 (United States); Asare, B.K. [Department of Pharmacology and Toxicology, University of Buffalo, Buffalo, NY 14214 (United States); Biswas, P.K., E-mail: pbiswas@tougaloo.edu [Laboratory of Computational Biophysics & Bioengineering, Department of Physics, Tougaloo College, Tougaloo, MS 39174 (United States); Rajnarayanan, R.V., E-mail: rajendra@buffalo.edu [Department of Pharmacology and Toxicology, University of Buffalo, Buffalo, NY 14214 (United States)

    2016-09-09

    The nuclear transcription factor estrogen receptor alpha (ERα), triggered by its cognate ligand estrogen, regulates a variety of cellular signaling events. ERα is expressed in 70% of breast cancers and is a widely validated target for anti-breast cancer drug discovery. Administration of anti-estrogen to block estrogen receptor activation is still a viable anti-breast cancer treatment option but anti-estrogen resistance has been a significant bottle-neck. Dimerization of estrogen receptor is required for ER activation. Blocking ERα dimerization is therefore a complementary and alternative strategy to combat anti-estrogen resistance. Dimer interface peptide “I-box” derived from ER residues 503–518 specifically blocks ER dimerization. Recently using a comprehensive molecular simulation we studied the interaction dynamics of ERα LBDs in a homo-dimer. Based on this study, we identified three interface recognition peptide motifs LDKITDT (ERα residues 479–485), LQQQHQRLAQ (residues 497–506), and LSHIRHMSNK (residues 511–520) and reported the suitability of using LQQQHQRLAQ (ER 497–506) as a template to design inhibitors of ERα dimerization. Stability and self-aggregation of peptide based therapeutics poses a significant bottle-neck to proceed further. In this study utilizing peptide grafted to preserve their pharmacophoric recognition motif and assessed their stability and potential to block ERα mediated activity in silico and in vitro. The Grafted peptides blocked ERα mediated cell proliferation and viability of breast cancer cells but did not alter their apoptotic fate. We believe the structural clues identified in this study can be used to identify novel peptidometics and small molecules that specifically target ER dimer interface generating a new breed of anti-cancer agents. - Highlights: • Designer peptide grafts retain core molecular recognition motif during MD simulations. • Designer peptide grafts with Poly-ALA helix form stable

  4. Design and Engineering Strategies for Synthetic Antimicrobial Peptides

    Science.gov (United States)

    Tossi, Alessandro

    Thousands of antimicrobial peptides (AMPs) of prokaryotic, fungal, plant, or animal origin have been identified, and their potential as lead compounds for the design of novel therapeutic agents in the treatment of infection, for stimulating the immune system, or in countering septic shock has been widely recognized. Added to this is their possible use in prophylaxis of infectious diseases for animal or plant protection, for disinfection of surgical instruments or industrial surfaces, and for food preservation among other commercially important applications. Since the early eighties, AMPs have been subject to a vast number of studies aimed at understanding what determines their potency and spectrum of activities against bacterial or fungal pathogens, and at maximizing these while limiting cytotoxic activities toward host cells. Much research has also been directed toward understanding specific mechanisms of action underlying the antimicrobial activity and selectivity, to be able to redesign the peptides for optimal performance. A central theme in the mode of action of many AMPs is their dynamic interaction with biological membranes, which involves various properties of these peptides such as, among others, surface hydrophobicity and polarity, charge, structure, and induced conformational variations. These features are often intimately interconnected so that engineering peptides to independently adjust any one property in particular is not an easy task. However, solid-phase peptide synthesis allows the use of a large repertoire of nonproteinogenic amino acids that can be used in the rational design of peptides to finely tune structural and physicochemical properties and precisely probe structure-function relationships.

  5. De Novo Design of Bioactive Small Molecules by Artificial Intelligence.

    Science.gov (United States)

    Merk, Daniel; Friedrich, Lukas; Grisoni, Francesca; Schneider, Gisbert

    2018-01-01

    Generative artificial intelligence offers a fresh view on molecular design. We present the first-time prospective application of a deep learning model for designing new druglike compounds with desired activities. For this purpose, we trained a recurrent neural network to capture the constitution of a large set of known bioactive compounds represented as SMILES strings. By transfer learning, this general model was fine-tuned on recognizing retinoid X and peroxisome proliferator-activated receptor agonists. We synthesized five top-ranking compounds designed by the generative model. Four of the compounds revealed nanomolar to low-micromolar receptor modulatory activity in cell-based assays. Apparently, the computational model intrinsically captured relevant chemical and biological knowledge without the need for explicit rules. The results of this study advocate generative artificial intelligence for prospective de novo molecular design, and demonstrate the potential of these methods for future medicinal chemistry. © 2018 The Authors. Published by Wiley-VCH Verlag GmbH & Co. KGaA.

  6. Concept of combinatorial de novo design of drug-like molecules by particle swarm optimization.

    Science.gov (United States)

    Hartenfeller, Markus; Proschak, Ewgenij; Schüller, Andreas; Schneider, Gisbert

    2008-07-01

    We present a fast stochastic optimization algorithm for fragment-based molecular de novo design (COLIBREE, Combinatorial Library Breeding). The search strategy is based on a discrete version of particle swarm optimization. Molecules are represented by a scaffold, which remains constant during optimization, and variable linkers and side chains. Different linkers represent virtual chemical reactions. Side-chain building blocks were obtained from pseudo-retrosynthetic dissection of large compound databases. Here, ligand-based design was performed using chemically advanced template search (CATS) topological pharmacophore similarity to reference ligands as fitness function. A weighting scheme was included for particle swarm optimization-based molecular design, which permits the use of many reference ligands and allows for positive and negative design to be performed simultaneously. In a case study, the approach was applied to the de novo design of potential peroxisome proliferator-activated receptor subtype-selective agonists. The results demonstrate the ability of the technique to cope with large combinatorial chemistry spaces and its applicability to focused library design. The technique was able to perform exploitation of a known scheme and at the same time explorative search for novel ligands within the framework of a given molecular core structure. It thereby represents a practical solution for compound screening in the early hit and lead finding phase of a drug discovery project.

  7. Design, synthesis, and actions of a novel chimeric natriuretic peptide: CD-NP.

    Science.gov (United States)

    Lisy, Ondrej; Huntley, Brenda K; McCormick, Daniel J; Kurlansky, Paul A; Burnett, John C

    2008-07-01

    Our aim was to design, synthesize and test in vivo and in vitro a new chimeric peptide that would combine the beneficial properties of 2 distinct natriuretic peptides with a biological profile that goes beyond native peptides. Studies have established the beneficial vascular and antiproliferative properties of C-type natriuretic peptide (CNP). While lacking renal actions, CNP is less hypotensive than the cardiac peptides atrial natriuretic peptide and B-type natriuretic peptide but unloads the heart due to venodilation. Dendroaspis natriuretic peptide is a potent natriuretic and diuretic peptide that is markedly hypotensive and functions via a separate guanylyl cyclase receptor compared with CNP. Here we engineered a novel chimeric peptide CD-NP that represents the fusion of the 22-amino acid peptide CNP together with the 15-amino acid linear C-terminus of Dendroaspis natriuretic peptide. We also determined in vitro in cardiac fibroblasts cyclic guanosine monophosphate-activating and antiproliferative properties of CD-NP. Our studies demonstrate in vivo that CD-NP is natriuretic and diuretic, glomerular filtration rate enhancing, cardiac unloading, and renin inhibiting. CD-NP also demonstrates less hypotensive properties when compared with B-type natriuretic peptide. In addition, CD-NP in vitro activates cyclic guanosine monophosphate and inhibits cardiac fibroblast proliferation. The current findings advance an innovative design strategy in natriuretic peptide drug discovery and development to create therapeutic peptides with favorable properties that may be preferable to those associated with native natriuretic peptides.

  8. PLGA nanofibers blended with designer self-assembling peptides for peripheral neural regeneration

    Energy Technology Data Exchange (ETDEWEB)

    Nune, Manasa; Krishnan, Uma Maheswari; Sethuraman, Swaminathan, E-mail: swami@sastra.edu

    2016-05-01

    Electrospun nanofibers are attractive candidates for neural regeneration due to similarity to the extracellular matrix. Several synthetic polymers have been used but they lack in providing the essential biorecognition motifs on their surfaces. Self-assembling peptide nanofiber scaffolds (SAPNFs) like RADA16 and recently, designer SAPs with functional motifs RADA16-I-BMHP1 areexamples, which showed successful spinal cord regeneration. But these peptide nanofiber scaffolds have poor mechanical properties and faster degradation rates that limit their use for larger nerve defects. Hence, we have developed a novel hybrid nanofiber scaffold of polymer poly(L-lactide-co-glycolide) (PLGA) and RADA16-I-BMHP1. The scaffolds were characterized for the presence of peptides both qualitatively and quantitatively using several techniques like SEM, EDX, FTIR, CHN analysis, Circular Dichroism analysis, Confocal and thermal analysis. Peptide self-assembly was retained post-electrospinning and formed rod-like nanostructures on PLGA nanofibers. In vitro cell compatibility was studied using rat Schwann cells and their adhesion, proliferation and gene expression levels on the designed scaffolds were evaluated. Our results have revealed the significant effects of the peptide blended scaffolds on promoting Schwann cell adhesion, extension and phenotypic expression. Neural development markers (SEM3F, NRP2 & PLX1) gene expression levels were significantly upregulated in peptide blended scaffolds compared to the PLGA scaffolds. Thus the hybrid blended novel designer scaffolds seem to be promising candidates for successful and functional regeneration of the peripheral nerve. - Highlights: • A novel blended scaffold of polymer PLGA and designer self-assembling peptide RADA16-I-BMPH1 was designed • The peptide retained the self-assembling features and formed rod like nanostructures on top of PLGA nanofibers • PLGA-peptide scaffolds have promoted the Schwann cell bipolar extension and

  9. C-peptide comparative radioimmunoassays: a study of three commercial kits

    International Nuclear Information System (INIS)

    Villaume, C.; Beck, B.

    1983-01-01

    Plasma C-peptide immunoreactivity (CPR) was measured in 18 fasting subjects with three different commercial kits (RIA-mat C-peptide-, Byk-Mallinckrodt; RIA-gnost-hC-peptide, Hoechst-Behring; human C-peptide radioimmunoassay kit, Novo) The subjects were chosen as to cover a wide range of CPR concentrations (five healthy subjects, six obese subjects, three insulin-dependent diabetics, four normal subjects whose plasmas had been kept at - 20 0 C for periods of 16 or 36 months). CPR was measured with the Novo kit in eight other plasmas which were kept over a period of 36 months, with or without aprotinin. Good correlations have been established among the values found with the three kits. However, absolute concentration values for each subject as well as the dispersion of all plasma C-peptide values varied as a function of the kit used because of antibody specificity differences and because of the various separation methods. The normal range proposed changes with each kit and the blood CPR of a subject can be a normal, reduced or increased one, depending on the kit used. After several months of storage, plasma CPR degradation is observed with the three kits. A protease-inhibitor is necessary in order to avoid this C-peptide degradation due to the apparent existence of a plasma proteolytic enzyme

  10. Interaction of amyloid inhibitor proteins with amyloid beta peptides: insight from molecular dynamics simulations.

    Directory of Open Access Journals (Sweden)

    Payel Das

    Full Text Available Knowledge of the detailed mechanism by which proteins such as human αB- crystallin and human lysozyme inhibit amyloid beta (Aβ peptide aggregation is crucial for designing treatment for Alzheimer's disease. Thus, unconstrained, atomistic molecular dynamics simulations in explicit solvent have been performed to characterize the Aβ17-42 assembly in presence of the αB-crystallin core domain and of lysozyme. Simulations reveal that both inhibitor proteins compete with inter-peptide interaction by binding to the peptides during the early stage of aggregation, which is consistent with their inhibitory action reported in experiments. However, the Aβ binding dynamics appear different for each inhibitor. The binding between crystallin and the peptide monomer, dominated by electrostatics, is relatively weak and transient due to the heterogeneous amino acid distribution of the inhibitor surface. The crystallin-bound Aβ oligomers are relatively long-lived, as they form more extensive contact surface with the inhibitor protein. In contrast, a high local density of arginines from lysozyme allows strong binding with Aβ peptide monomers, resulting in stable complexes. Our findings not only illustrate, in atomic detail, how the amyloid inhibitory mechanism of human αB-crystallin, a natural chaperone, is different from that of human lysozyme, but also may aid de novo design of amyloid inhibitors.

  11. De novo design of peptide immunogens that mimic the coiled coil region of human T-cell leukemia virus type-1 glycoprotein 21 transmembrane subunit for induction of native protein reactive neutralizing antibodies.

    Science.gov (United States)

    Sundaram, Roshni; Lynch, Marcus P; Rawale, Sharad V; Sun, Yiping; Kazanji, Mirdad; Kaumaya, Pravin T P

    2004-06-04

    Peptide vaccines able to induce high affinity and protective neutralizing antibodies must rely in part on the design of antigenic epitopes that mimic the three-dimensional structure of the corresponding region in the native protein. We describe the design, structural characterization, immunogenicity, and neutralizing potential of antibodies elicited by conformational peptides derived from the human T-cell leukemia virus type 1 (HTLV-1) gp21 envelope glycoprotein spanning residues 347-374. We used a novel template design and a unique synthetic approach to construct two peptides (WCCR2T and CCR2T) that would each assemble into a triple helical coiled coil conformation mimicking the gp21 crystal structure. The peptide B-cell epitopes were grafted onto the epsilon side chains of three lysyl residues on a template backbone construct consisting of the sequence acetyl-XGKGKGKGCONH2 (where X represents the tetanus toxoid promiscuous T cell epitope (TT) sequence 580-599). Leucine substitutions were introduced at the a and d positions of the CCR2T sequence to maximize helical character and stability as shown by circular dichroism and guanidinium hydrochloride studies. Serum from an HTLV-1-infected patient was able to recognize the selected epitopes by enzyme-linked immunosorbent assay (ELISA). Mice immunized with the wild-type sequence (WCCR2T) and the mutant sequence (CCR2T) elicited high antibody titers that were capable of recognizing the native protein as shown by flow cytometry and whole virus ELISA. Sera and purified antibodies from immunized mice were able to reduce the formation of syncytia induced by the envelope glycoprotein of HTLV-1, suggesting that antibodies directed against the coiled coil region of gp21 are capable of disrupting cell-cell fusion. Our results indicate that these peptides represent potential candidates for use in a peptide vaccine against HTLV-1.

  12. Antimicrobial peptides in innate immune responses

    DEFF Research Database (Denmark)

    Sorensen, O.E.; Borregaard, N.; Cole, A.M.

    2008-01-01

    Antimicrobial peptides (AMPs) are ancient effector molecules in the innate immune response of eukaryotes. These peptides are important for the antimicrobial efficacy of phagocytes and for the innate immune response mounted by epithelia of humans and other mammals. AMPs are generated either by de...... novo synthesis or by proteolytic cleavage from antimicrobially inactive proproteins. Studies of human diseases and animal studies have given important clues to the in vivo role of AMPs. It is now evident that dysregulation of the generation of AMPs in innate immune responses plays a role in certain...

  13. Biological Applications of Designed Hairpin Peptides: As Antimicrobials and as Inhibitors of Amyloidogenesis

    Science.gov (United States)

    Sivanesam, Kalkena

    More than 40 diseases have been associated with the misfolding of peptides (or proteins) that form fibrils with a very specific morphology. These peptides classified as amyloidogenic peptides have been implicated in the development of Alzheimer's Disease, Parkinson's Disease, Type II Diabetes, Hungtinton's Disease etc. To date, these diseases have no cure, only therapies that can ameliorate the symptoms to a degree. Inhibition of the amyloidogenesis of these peptides has been proposed as a possible treatment option. While small molecules have been heavily tested as inhibitors of amyloidogenesis, peptides have emerged as potential inhibitors. In this work, the ability of a set of designed hairpin peptides to inhibit the amyloidogenesis of two different systems, alpha-synuclein (implicated in Parkinson's Disease) and human amylin (implicated in Type II Diabetes) is tested. Using circular dichroism and thioflavin T fluorescence, the ability of these peptides to inhibit amyloidogenesis is tested. The binding loci of these inhibitors to alpha-synuclein are also explored. The use of peptides as antimicrobials on the other hand is not a novel concept. However, most antimicrobial peptides, both natural and designed, rely heavily on covalent stabilizations in order to maintain secondary structure. In this study, non-covalent stabilizations are applied to a couple of natural as well as designed antimicrobials in order to study the effects of secondary structure stabilization on biological activity.

  14. Designing of peptides with desired half-life in intestine-like environment

    KAUST Repository

    Sharma, Arun; Singla, Deepak; Rashid, Mamoon; Raghava, Gajendra Pal Singh

    2014-01-01

    hindered mainly because of their high susceptibility towards proteases degradation. We have developed an in silico method to predict the half-life of peptides in intestine-like environment and to design better peptides having optimized physicochemical

  15. Lumican Peptides: Rational Design Targeting ALK5/TGFBRI

    Science.gov (United States)

    Gesteira, Tarsis Ferreira; Coulson-Thomas, Vivien J.; Yuan, Yong; Zhang, Jianhua; Nader, Helena B.; Kao, Winston W.-Y.

    2017-02-01

    Lumican, a small leucine rich proteoglycan (SLRP), is a component of extracellular matrix which also functions as a matrikine regulating multiple cell activities. In the cornea, lumican maintains corneal transparency by regulating collagen fibrillogenesis, promoting corneal epithelial wound healing, regulating gene expression and maintaining corneal homeostasis. We have recently shown that a peptide designed from the 13 C-terminal amino acids of lumican (LumC13) binds to ALK5/TGFBR1 (type1 receptor of TGFβ) to promote wound healing. Herein we evaluate the mechanism by which this synthetic C-terminal amphiphilic peptide (LumC13), binds to ALK5. These studies clearly reveal that LumC13-ALK5 form a stable complex. In order to determine the minimal amino acids required for the formation of a stable lumican/ALK5 complex derivatives of LumC13 were designed and their binding to ALK5 investigated in silico. These LumC13 derivatives were tested both in vitro and in vivo to evaluate their ability to promote corneal epithelial cell migration and corneal wound healing, respectively. These validations add to the therapeutic value of LumC13 (Lumikine) and aid its clinical relevance of promoting the healing of corneal epithelium debridement. Moreover, our data validates the efficacy of our computational approach to design active peptides based on interactions of receptor and chemokine/ligand.

  16. The Folding of de Novo Designed Protein DS119 via Molecular Dynamics Simulations

    Directory of Open Access Journals (Sweden)

    Moye Wang

    2016-04-01

    Full Text Available As they are not subjected to natural selection process, de novo designed proteins usually fold in a manner different from natural proteins. Recently, a de novo designed mini-protein DS119, with a βαβ motif and 36 amino acids, has folded unusually slowly in experiments, and transient dimers have been detected in the folding process. Here, by means of all-atom replica exchange molecular dynamics (REMD simulations, several comparably stable intermediate states were observed on the folding free-energy landscape of DS119. Conventional molecular dynamics (CMD simulations showed that when two unfolded DS119 proteins bound together, most binding sites of dimeric aggregates were located at the N-terminal segment, especially residues 5–10, which were supposed to form β-sheet with its own C-terminal segment. Furthermore, a large percentage of individual proteins in the dimeric aggregates adopted conformations similar to those in the intermediate states observed in REMD simulations. These results indicate that, during the folding process, DS119 can easily become trapped in intermediate states. Then, with diffusion, a transient dimer would be formed and stabilized with the binding interface located at N-terminals. This means that it could not quickly fold to the native structure. The complicated folding manner of DS119 implies the important influence of natural selection on protein-folding kinetics, and more improvement should be achieved in rational protein design.

  17. Design and expression of a short peptide as an HIV detection probe

    Energy Technology Data Exchange (ETDEWEB)

    Lines, Jamie A.; Yu, Zhiqiang; Dedkova, Larisa M.; Chen, Shengxi, E-mail: shengxi.chen.1@asu.edu

    2014-01-03

    Highlights: •We designed a short fusion peptide (FP-50) for in vivo expression. •This peptide is a very promising component for detection of gp120 protein. •The detectable level is about 20–200 times lower than previously published methods. •It is a novel probe to detect HIV-1 gp120 during early stages of HIV infection. -- Abstract: To explore a low-cost novel probe for HIV detection, we designed and prepared a 50-amino acid-length short fusion peptide (FP-50) via Escherichia coli in vivo expression. It was employed as a novel probe to detect HIV-1 gp120 protein. The detectable level of gp120 protein using the FP-50 peptide was approximately 20–200 times lower than previously published methods that used a pair of monoclonal antibodies. Thus, this short peptide is a very promising component for detection of gp120 protein during early stages of HIV infection.

  18. Programming peptidomimetic syntheses by translating genetic codes designed de novo.

    Science.gov (United States)

    Forster, Anthony C; Tan, Zhongping; Nalam, Madhavi N L; Lin, Hening; Qu, Hui; Cornish, Virginia W; Blacklow, Stephen C

    2003-05-27

    Although the universal genetic code exhibits only minor variations in nature, Francis Crick proposed in 1955 that "the adaptor hypothesis allows one to construct, in theory, codes of bewildering variety." The existing code has been expanded to enable incorporation of a variety of unnatural amino acids at one or two nonadjacent sites within a protein by using nonsense or frameshift suppressor aminoacyl-tRNAs (aa-tRNAs) as adaptors. However, the suppressor strategy is inherently limited by compatibility with only a small subset of codons, by the ways such codons can be combined, and by variation in the efficiency of incorporation. Here, by preventing competing reactions with aa-tRNA synthetases, aa-tRNAs, and release factors during translation and by using nonsuppressor aa-tRNA substrates, we realize a potentially generalizable approach for template-encoded polymer synthesis that unmasks the substantially broader versatility of the core translation apparatus as a catalyst. We show that several adjacent, arbitrarily chosen sense codons can be completely reassigned to various unnatural amino acids according to de novo genetic codes by translating mRNAs into specific peptide analog polymers (peptidomimetics). Unnatural aa-tRNA substrates do not uniformly function as well as natural substrates, revealing important recognition elements for the translation apparatus. Genetic programming of peptidomimetic synthesis should facilitate mechanistic studies of translation and may ultimately enable the directed evolution of small molecules with desirable catalytic or pharmacological properties.

  19. A de novo designed monomeric, compact three helix bundle protein on a carbohydrate template

    DEFF Research Database (Denmark)

    Malik, Leila; Nygård, Jesper; Christensen, Niels Johan

    2015-01-01

    De novo design and chemical synthesis of proteins and of other artificial structures, which mimic them, is a central strategy for understanding protein folding and for accessing proteins with novel functions. We have previously described carbohydrates as templates for the assembly of artificial...... the template could facilitate protein folding. Here we report the design and synthesis of 3-helix bundle carboproteins on deoxy-hexopyranosides. The carboproteins were analyzed by CD, AUC, SAXS, and NMR, which revealed the formation of the first compact, and folded monomeric carboprotein distinctly different...

  20. Therapeutic peptides for cancer therapy. Part II - cell cycle inhibitory peptides and apoptosis-inducing peptides.

    Science.gov (United States)

    Raucher, Drazen; Moktan, Shama; Massodi, Iqbal; Bidwell, Gene L

    2009-10-01

    Therapeutic peptides have great potential as anticancer agents owing to their ease of rational design and target specificity. However, their utility in vivo is limited by low stability and poor tumor penetration. The authors review the development of peptide inhibitors with potential for cancer therapy. Peptides that arrest the cell cycle by mimicking CDK inhibitors or induce apoptosis directly are discussed. The authors searched Medline for articles concerning the development of therapeutic peptides and their delivery. Inhibition of cancer cell proliferation directly using peptides that arrest the cell cycle or induce apoptosis is a promising strategy. Peptides can be designed that interact very specifically with cyclins and/or cyclin-dependent kinases and with members of apoptotic cascades. Use of these peptides is not limited by their design, as a rational approach to peptide design is much less challenging than the design of small molecule inhibitors of specific protein-protein interactions. However, the limitations of peptide therapy lie in the poor pharmacokinetic properties of these large, often charged molecules. Therefore, overcoming the drug delivery hurdles could open the door for effective peptide therapy, thus making an entirely new class of molecules useful as anticancer drugs.

  1. SwePep, a database designed for endogenous peptides and mass spectrometry.

    Science.gov (United States)

    Fälth, Maria; Sköld, Karl; Norrman, Mathias; Svensson, Marcus; Fenyö, David; Andren, Per E

    2006-06-01

    A new database, SwePep, specifically designed for endogenous peptides, has been constructed to significantly speed up the identification process from complex tissue samples utilizing mass spectrometry. In the identification process the experimental peptide masses are compared with the peptide masses stored in the database both with and without possible post-translational modifications. This intermediate identification step is fast and singles out peptides that are potential endogenous peptides and can later be confirmed with tandem mass spectrometry data. Successful applications of this methodology are presented. The SwePep database is a relational database developed using MySql and Java. The database contains 4180 annotated endogenous peptides from different tissues originating from 394 different species as well as 50 novel peptides from brain tissue identified in our laboratory. Information about the peptides, including mass, isoelectric point, sequence, and precursor protein, is also stored in the database. This new approach holds great potential for removing the bottleneck that occurs during the identification process in the field of peptidomics. The SwePep database is available to the public.

  2. Application of Generative Autoencoder in De Novo Molecular Design.

    Science.gov (United States)

    Blaschke, Thomas; Olivecrona, Marcus; Engkvist, Ola; Bajorath, Jürgen; Chen, Hongming

    2018-01-01

    A major challenge in computational chemistry is the generation of novel molecular structures with desirable pharmacological and physiochemical properties. In this work, we investigate the potential use of autoencoder, a deep learning methodology, for de novo molecular design. Various generative autoencoders were used to map molecule structures into a continuous latent space and vice versa and their performance as structure generator was assessed. Our results show that the latent space preserves chemical similarity principle and thus can be used for the generation of analogue structures. Furthermore, the latent space created by autoencoders were searched systematically to generate novel compounds with predicted activity against dopamine receptor type 2 and compounds similar to known active compounds not included in the trainings set were identified. © 2018 The Authors. Published by Wiley-VCH Verlag GmbH & Co. KGaA.

  3. Designer Self-Assembling Peptide Nanofiber Scaffolds Containing Link Protein N-Terminal Peptide Induce Chondrogenesis of Rabbit Bone Marrow Stem Cells

    Directory of Open Access Journals (Sweden)

    Baichuan Wang

    2014-01-01

    Full Text Available Designer self-assembling peptide nanofiber hydrogel scaffolds have been considered as promising biomaterials for tissue engineering because of their excellent biocompatibility and biofunctionality. Our previous studies have shown that a novel designer functionalized self-assembling peptide nanofiber hydrogel scaffold (RLN/RADA16, LN-NS containing N-terminal peptide sequence of link protein (link N can promote nucleus pulposus cells (NPCs adhesion and three-dimensional (3D migration and stimulate biosynthesis of type II collagen and aggrecan by NPCs in vitro. The present study has extended these investigations to determine the effects of this functionalized LN-NS on bone marrow stem cells (BMSCs, a potential cell source for NP regeneration. Although the functionalized LN-NS cannot promote BMSCs proliferation, it significantly promotes BMSCs adhesion compared with that of the pure RADA16 hydrogel scaffold. Moreover, the functionalized LN-NS remarkably stimulates biosynthesis and deposition of type II collagen and aggrecan. These data demonstrate that the functionalized peptide nanofiber hydrogel scaffold containing link N peptide as a potential matrix substrate will be very useful in the NP tissue regeneration.

  4. Molecular Design, Structures, and Activity of Antimicrobial Peptide-Mimetic Polymers

    Science.gov (United States)

    Takahashi, Haruko; Palermo, Edmund F.; Yasuhara, Kazuma; Caputo, Gregory A.

    2014-01-01

    There is an urgent need for new antibiotics which are effective against drug-resistant bacteria without contributing to resistance development. We have designed and developed antimicrobial copolymers with cationic amphiphilic structures based on the mimicry of naturally occurring antimicrobial peptides. These copolymers exhibit potent antimicrobial activity against a broad spectrum of bacteria including methicillin-resistant Staphylococcus aureus with no adverse hemolytic activity. Notably, these polymers also did not result in any measurable resistance development in E. coli. The peptide-mimetic design principle offers significant flexibility and diversity in the creation of new antimicrobial materials and their potential biomedical applications. PMID:23832766

  5. Computer-aided designing of immunosuppressive peptides based on IL-10 inducing potential

    Science.gov (United States)

    Nagpal, Gandharva; Usmani, Salman Sadullah; Dhanda, Sandeep Kumar; Kaur, Harpreet; Singh, Sandeep; Sharma, Meenu; Raghava, Gajendra P. S.

    2017-01-01

    In the past, numerous methods have been developed to predict MHC class II binders or T-helper epitopes for designing the epitope-based vaccines against pathogens. In contrast, limited attempts have been made to develop methods for predicting T-helper epitopes/peptides that can induce a specific type of cytokine. This paper describes a method, developed for predicting interleukin-10 (IL-10) inducing peptides, a cytokine responsible for suppressing the immune system. All models were trained and tested on experimentally validated 394 IL-10 inducing and 848 non-inducing peptides. It was observed that certain types of residues and motifs are more frequent in IL-10 inducing peptides than in non-inducing peptides. Based on this analysis, we developed composition-based models using various machine-learning techniques. Random Forest-based model achieved the maximum Matthews’s Correlation Coefficient (MCC) value of 0.59 with an accuracy of 81.24% developed using dipeptide composition. In order to facilitate the community, we developed a web server “IL-10pred”, standalone packages and a mobile app for designing IL-10 inducing peptides (http://crdd.osdd.net/raghava/IL-10pred/). PMID:28211521

  6. Novel anti-HIV peptides containing multiple copies of artificially designed heptad repeat motifs

    International Nuclear Information System (INIS)

    Shi Weiguo; Qi Zhi; Pan Chungen; Xue Na; Debnath, Asim K.; Qie Jiankun; Jiang Shibo; Liu Keliang

    2008-01-01

    The peptidic anti-HIV drug T20 (Fuzeon) and its analog C34 share a common heptad repeat (HR) sequence, but they have different functional domains, i.e., pocket- and lipid-binding domains (PBD and LBD, respectively). We hypothesize that novel anti-HIV peptides may be designed by using artificial sequences containing multiple copies of HR motifs plus zero, one or two functional domains. Surprisingly, we found that the peptides containing only the non-natural HR sequences could significantly inhibit HIV-1 infection, while addition of PBD and/or LBD to the peptides resulted in significant improvement of anti-HIV-1 activity. These results suggest that these artificial HR sequences, which may serve as structural domains, could be used as templates for the design of novel antiviral peptides against HIV and other viruses with class I fusion proteins

  7. De novo molecular design

    CERN Document Server

    Schneider, Gisbert

    2013-01-01

    Systematically examining current methods and strategies, this ready reference covers a wide range of molecular structures, from organic-chemical drugs to peptides, Proteins and nucleic acids, in line with emerging new drug classes derived from biomacromolecules. A leader in the field and one of the pioneers of this young discipline has assembled here the most prominent experts from across the world to provide first-hand knowledge. While most of their methods and examples come from the area of pharmaceutical discovery and development, the approaches are equally applicable for chemical probes an

  8. iScreen: world's first cloud-computing web server for virtual screening and de novo drug design based on TCM database@Taiwan.

    Science.gov (United States)

    Tsai, Tsung-Ying; Chang, Kai-Wei; Chen, Calvin Yu-Chian

    2011-06-01

    The rapidly advancing researches on traditional Chinese medicine (TCM) have greatly intrigued pharmaceutical industries worldwide. To take initiative in the next generation of drug development, we constructed a cloud-computing system for TCM intelligent screening system (iScreen) based on TCM Database@Taiwan. iScreen is compacted web server for TCM docking and followed by customized de novo drug design. We further implemented a protein preparation tool that both extract protein of interest from a raw input file and estimate the size of ligand bind site. In addition, iScreen is designed in user-friendly graphic interface for users who have less experience with the command line systems. For customized docking, multiple docking services, including standard, in-water, pH environment, and flexible docking modes are implemented. Users can download first 200 TCM compounds of best docking results. For TCM de novo drug design, iScreen provides multiple molecular descriptors for a user's interest. iScreen is the world's first web server that employs world's largest TCM database for virtual screening and de novo drug design. We believe our web server can lead TCM research to a new era of drug development. The TCM docking and screening server is available at http://iScreen.cmu.edu.tw/.

  9. iScreen: world's first cloud-computing web server for virtual screening and de novo drug design based on TCM database@Taiwan

    Science.gov (United States)

    Tsai, Tsung-Ying; Chang, Kai-Wei; Chen, Calvin Yu-Chian

    2011-06-01

    The rapidly advancing researches on traditional Chinese medicine (TCM) have greatly intrigued pharmaceutical industries worldwide. To take initiative in the next generation of drug development, we constructed a cloud-computing system for TCM intelligent screening system (iScreen) based on TCM Database@Taiwan. iScreen is compacted web server for TCM docking and followed by customized de novo drug design. We further implemented a protein preparation tool that both extract protein of interest from a raw input file and estimate the size of ligand bind site. In addition, iScreen is designed in user-friendly graphic interface for users who have less experience with the command line systems. For customized docking, multiple docking services, including standard, in-water, pH environment, and flexible docking modes are implemented. Users can download first 200 TCM compounds of best docking results. For TCM de novo drug design, iScreen provides multiple molecular descriptors for a user's interest. iScreen is the world's first web server that employs world's largest TCM database for virtual screening and de novo drug design. We believe our web server can lead TCM research to a new era of drug development. The TCM docking and screening server is available at http://iScreen.cmu.edu.tw/.

  10. Rational design and application of responsive α-helical peptide hydrogels

    Science.gov (United States)

    Banwell, Eleanor F.; Abelardo, Edgardo S.; Adams, Dave J.; Birchall, Martin A.; Corrigan, Adam; Donald, Athene M.; Kirkland, Mark; Serpell, Louise C.; Butler, Michael F.; Woolfson, Derek N.

    2009-01-01

    Biocompatible hydrogels have a wide variety of potential applications in biotechnology and medicine, such as the controlled delivery and release of cells, cosmetics and drugs; and as supports for cell growth and tissue engineering1. Rational peptide design and engineering are emerging as promising new routes to such functional biomaterials2-4. Here we present the first examples of rationally designed and fully characterized self-assembling hydrogels based on standard linear peptides with purely α-helical structures, which we call hydrogelating self-assembling fibres (hSAFs). These form spanning networks of α-helical fibrils that interact to give self-supporting physical hydrogels of >99% water content. The peptide sequences can be engineered to alter the underlying mechanism of gelation and, consequently, the hydrogel properties. Interestingly, for example, those with hydrogen-bonded networks melt upon heating, whereas those formed via hydrophobic interactions strengthen when warmed. The hSAFs are dual-peptide systems that only gel on mixing, which gives tight control over assembly5. These properties raise possibilities for using the hSAFs as substrates in cell culture. We have tested this in comparison with the widely used Matrigel substrate, and demonstrate that, like Matrigel, hSAFs support both growth and differentiation of rat adrenal pheochromocytoma cells for sustained periods in culture. PMID:19543314

  11. Design, synthesis, and validation of an in vitro platform peptide-whole cell screening assay using MTT reagent

    Directory of Open Access Journals (Sweden)

    Sahar Ahmed

    2017-05-01

    Full Text Available An in vitro platform to perform peptide screening against different cancer cell lines was designed. The strategy for this screening relied on the design and detection of high-affinity cancer-targeting peptides based on the sequences of NGR and P160. Evaluation of the best binding peptides was performed via incubation of the peptide array-bounded cells with MTT reagent, which is reduced to purple formazan in living cells and further quantified using an Elispot and Kodak imager. For proof of concept, a peptide library (132 spots, and 66 different peptides was designed, synthesized, and screened against different cancer cell lines. The current strategy assists in the identification of positive and negative peptides as well as the relative binding between positive ones. Better binding peptide sequences of the NGR motif were demonstrated to show up to a 2.6-fold increase in CD13+ cell lines with insignificant binding to CD13− ones. Comparable results were observed for P160 peptide sequences, to which different peptides had increased binding, with an up to 3-fold increase relative to the native P160 peptide. Based on our results, new peptide sequences for cancer targeting were identified, and the developed strategy was applied to two different peptide libraries.

  12. Rational design and synthesis of an orally bioavailable peptide guided by NMR amide temperature coefficients

    Science.gov (United States)

    Wang, Conan K.; Northfield, Susan E.; Colless, Barbara; Chaousis, Stephanie; Hamernig, Ingrid; Lohman, Rink-Jan; Nielsen, Daniel S.; Schroeder, Christina I.; Liras, Spiros; Price, David A.; Fairlie, David P.; Craik, David J.

    2014-01-01

    Enhancing the oral bioavailability of peptide drug leads is a major challenge in drug design. As such, methods to address this challenge are highly sought after by the pharmaceutical industry. Here, we propose a strategy to identify appropriate amides for N-methylation using temperature coefficients measured by NMR to identify exposed amides in cyclic peptides. N-methylation effectively caps these amides, modifying the overall solvation properties of the peptides and making them more membrane permeable. The approach for identifying sites for N-methylation is a rapid alternative to the elucidation of 3D structures of peptide drug leads, which has been a commonly used structure-guided approach in the past. Five leucine-rich peptide scaffolds are reported with selectively designed N-methylated derivatives. In vitro membrane permeability was assessed by parallel artificial membrane permeability assay and Caco-2 assay. The most promising N-methylated peptide was then tested in vivo. Here we report a novel peptide (15), which displayed an oral bioavailability of 33% in a rat model, thus validating the design approach. We show that this approach can also be used to explain the notable increase in oral bioavailability of a somatostatin analog. PMID:25416591

  13. G-CSF receptor-binding cyclic peptides designed with artificial amino-acid linkers

    International Nuclear Information System (INIS)

    Shibata, Kenji; Maruyama-Takahashi, Kumiko; Yamasaki, Motoo; Hirayama, Noriaki

    2006-01-01

    Designing small molecules that mimic the receptor-binding local surface structure of large proteins such as cytokines or growth factors is fascinating and challenging. In this study, we designed cyclic peptides that reproduce the receptor-binding loop structures of G-CSF. We found it is important to select a suitable linker to join two or more discontinuous sequences and both termini of the peptide corresponding to the receptor-binding loop. Structural simulations based on the crystallographic structure of KW-2228, a stable and potent analog of human G-CSF, led us to choose 4-aminobenzoic acid (Abz) as a part of the linker. A combination of 4-Abz with β-alanine or glycine, and disulfide bridges between cysteins or homocysteins, gave a structure suitable for receptor binding. In this structure, the side-chains of several amino acids important for the interactions with the receptor are protruding from one side of the peptide ring. This artificial peptide showed G-CSF antagonistic activity in a cell proliferation assay

  14. In silico design of cyclic peptides as influenza virus, a subtype H1N1 ...

    African Journals Online (AJOL)

    Cyclic peptides can be used to make new antiviral drug design especially to inhibit neuraminidase activity by using 'structure-based design' method. Based on molecular docking, new antiviral drug designs have been found. They are DNY, NNY, DDY, DYY, RRR, RPR, RRP and LRL. These cyclic peptides showed better ...

  15. Analysis of skin derived peptides from the Cuyaba Dwarf Frog Physalaemus nattereri by off-line LC MALDI MS/MS

    DEFF Research Database (Denmark)

    Castro, Mariana S; Pires Júnior, Osmindo Rodrigues; Fontes, Wagner

    2017-01-01

    We have investigated the potential for analysis of the complex peptide mixtures secreted from frog skin using off-line LC-MALDI MS/MS. Since only limited information about the sequence of such peptides is available, de novo sequencing followed by Blast search was needed. An automated workflow has...... for future studies of peptides secreted from frogs....

  16. β-Boomerang Antimicrobial and Antiendotoxic Peptides: Lipidation and Disulfide Bond Effects on Activity and Structure.

    Science.gov (United States)

    Mohanram, Harini; Bhattacharjya, Surajit

    2014-04-21

    Drug-resistant Gram-negative bacterial pathogens and endotoxin- or lipopolysaccharide (LPS)-mediated inflammations are among some of the most  prominent health issues globally. Antimicrobial peptides (AMPs) are eminent molecules that can kill drug-resistant strains and neutralize LPS toxicity. LPS, the outer layer of the outer membrane of Gram-negative bacteria safeguards cell integrity against hydrophobic compounds, including antibiotics and AMPs. Apart from maintaining structural integrity, LPS, when released into the blood stream, also induces inflammatory pathways leading to septic shock. In previous works, we have reported the de novo design of a set of 12-amino acid long cationic/hydrophobic peptides for LPS binding and activity. These peptides adopt β-boomerang like conformations in complex with LPS. Structure-activity studies demonstrated some critical features of the β-boomerang scaffold that may be utilized for the further development of potent analogs. In this work, β-boomerang lipopeptides were designed and structure-activity correlation studies were carried out. These lipopeptides were homo-dimerized through a disulfide bridge to stabilize conformations and for improved activity. The designed peptides exhibited potent antibacterial activity and efficiently neutralized LPS toxicity under in vitro assays. NMR structure of C4YI13C in aqueous solution demonstrated the conserved folding of the lipopeptide with a boomerang aromatic lock stabilized with disulfide bond at the C-terminus and acylation at the N-terminus. These lipo-peptides displaying bacterial sterilization and low hemolytic activity may be useful for future applications as antimicrobial and antiendotoxin molecules.

  17. β-Boomerang Antimicrobial and Antiendotoxic Peptides: Lipidation and Disulfide Bond Effects on Activity and Structure

    Directory of Open Access Journals (Sweden)

    Harini Mohanram

    2014-04-01

    Full Text Available Drug-resistant Gram-negative bacterial pathogens and endotoxin- or lipopolysaccharide (LPS-mediated inflammations are among some of the most  prominent health issues globally. Antimicrobial peptides (AMPs are eminent molecules that can kill drug-resistant strains and neutralize LPS toxicity. LPS, the outer layer of the outer membrane of Gram-negative bacteria safeguards cell integrity against hydrophobic compounds, including antibiotics and AMPs. Apart from maintaining structural integrity, LPS, when released into the blood stream, also induces inflammatory pathways leading to septic shock. In previous works, we have reported the de novo design of a set of 12-amino acid long cationic/hydrophobic peptides for LPS binding and activity. These peptides adopt β-boomerang like conformations in complex with LPS. Structure-activity studies demonstrated some critical features of the β-boomerang scaffold that may be utilized for the further development of potent analogs. In this work, β-boomerang lipopeptides were designed and structure-activity correlation studies were carried out. These lipopeptides were homo-dimerized through a disulfide bridge to stabilize conformations and for improved activity. The designed peptides exhibited potent antibacterial activity and efficiently neutralized LPS toxicity under in vitro assays. NMR structure of C4YI13C in aqueous solution demonstrated the conserved folding of the lipopeptide with a boomerang aromatic lock stabilized with disulfide bond at the C-terminus and acylation at the N-terminus. These lipo-peptides displaying bacterial sterilization and low hemolytic activity may be useful for future applications as antimicrobial and antiendotoxin molecules.

  18. Interpretation of Tandem Mass Spectrometry (MSMS) Spectra for Peptide Analysis

    DEFF Research Database (Denmark)

    Hjernø, Karin; Højrup, Peter

    2015-01-01

    The aim of this chapter is to give a short introduction to peptide analysis by mass spectrometry (MS) and interpretation of fragment mass spectra. Through examples and guidelines we demonstrate how to understand and validate search results and how to perform de novo sequencing based on the often...... very complex fragmentation pattern obtained by tandem mass spectrometry (also referred to as MSMS). The focus is on simple rules for interpretation of MSMS spectra of tryptic as well as non-tryptic peptides....

  19. Evaluation of peptide designing strategy against subunit reassociation in mucin 1: A steered molecular dynamics approach.

    Directory of Open Access Journals (Sweden)

    J Lesitha Jeeva Kumari

    Full Text Available Subunit reassociation in mucin 1, a breast cancer tumor marker, is reported as one of the critical factors for its cytoplasmic activation. Inhibition of its heterodimeric association would therefore result in loss of its function and alter disease progression. The present study aimed at evaluating peptide inhibitor designing strategies that may serve as antagonist against this receptor-ligand alliance. Several peptides and their derivatives were designed based on native residues, subunit interface, hydrogen bonding and secondary structure. Docking studies with the peptides were carried on the receptor subunit and their binding affinities were evaluated using steered molecular dynamics simulation and umbrella sampling. Our results showed that among all the different classes of peptides evaluated, the receptor based peptide showed the highest binding affinity. This result was concurrent with the experimental observation that the receptor-ligand alliance in mucin 1 is highly specific. Our results also show that peptide ligand against this subunit association is only stabilized through native residue inter-protein interaction irrespective of the peptide structure, peptide length and number of hydrogen bonds. Consistency in binding affinity, pull force and free energy barrier was observed with only the receptor derived peptides which resulted in favorable interprotein interactions at the interface. Several observations were made and discussed which will eventually lead to designing efficient peptide inhibitors against mucin 1 heterodimeric subunit reassociation.

  20. Designed beta-boomerang antiendotoxic and antimicrobial peptides: structures and activities in lipopolysaccharide.

    Science.gov (United States)

    Bhunia, Anirban; Mohanram, Harini; Domadia, Prerna N; Torres, Jaume; Bhattacharjya, Surajit

    2009-08-14

    Lipopolysaccharide (LPS), an integral part of the outer membrane of Gram-negative bacteria, is involved in a variety of biological processes including inflammation, septic shock, and resistance to host-defense molecules. LPS also provides an environment for folding of outer membrane proteins. In this work, we describe the structure-activity correlation of a series of 12-residue peptides in LPS. NMR structures of the peptides derived in complex with LPS reveal boomerang-like beta-strand conformations that are stabilized by intimate packing between the two aromatic residues located at the 4 and 9 positions. This structural feature renders these peptides with a high ability to neutralize endotoxicity, >80% at 10 nM concentration, of LPS. Replacements of these aromatic residues either with Ala or with Leu destabilizes the boomerang structure with the concomitant loss of antiendotoxic and antimicrobial activities. Furthermore, the aromatic packing stabilizing the beta-boomerang structure in LPS is found to be maintained even in a truncated octapeptide, defining a structured LPS binding motif. The mode of action of the active designed peptides correlates well with their ability to perturb LPS micelle structures. Fourier transform infrared spectroscopy studies of the peptides delineate beta-type conformations and immobilization of phosphate head groups of LPS. Trp fluorescence studies demonstrated selective interactions with LPS and the depth of insertion into the LPS bilayer. Our results demonstrate the requirement of LPS-specific structures of peptides for endotoxin neutralizations. In addition, we propose that structures of these peptides may be employed to design proteins for the outer membrane.

  1. Peptide-based proteasome inhibitors in anticancer drug design.

    Science.gov (United States)

    Micale, Nicola; Scarbaci, Kety; Troiano, Valeria; Ettari, Roberta; Grasso, Silvana; Zappalà, Maria

    2014-09-01

    The identification of the key role of the eukaryotic 26S proteasome in regulated intracellular proteolysis and its importance as a target in many pathological conditions wherein the proteasomal activity is defective (e.g., malignancies, autoimmune diseases, neurodegenerative diseases, etc.) prompted several research groups to the development of specific inhibitors of this multicatalytic complex with the aim of obtaining valid drug candidates. In regard to the anticancer therapy, the peptide boronate bortezomib (Velcade®) represents the first molecule approved by FDA for the treatment of multiple myeloma in 2003 and mantle cell lymphoma in 2006. Since then, a plethora of molecules targeting the proteasome have been identified as potential anticancer agents and a few of them reached clinical trials or are already in the market (i.e., carfilzomib; Kyprolis®). In most cases, the design of new proteasome inhibitors (PIs) takes into account a proven peptide or pseudopeptide motif as a base structure and places other chemical entities throughout the peptide skeleton in such a way to create an efficacious network of interactions within the catalytic sites. The purpose of this review is to provide an in-depth look at the current state of the research in the field of peptide-based PIs, specifically those ones that might find an application as anticancer agents. © 2014 Wiley Periodicals, Inc.

  2. Customizable de novo design strategies for DOCK: Application to HIVgp41 and other therapeutic targets.

    Science.gov (United States)

    Allen, William J; Fochtman, Brian C; Balius, Trent E; Rizzo, Robert C

    2017-11-15

    De novo design can be used to explore vast areas of chemical space in computational lead discovery. As a complement to virtual screening, from-scratch construction of molecules is not limited to compounds in pre-existing vendor catalogs. Here, we present an iterative fragment growth method, integrated into the program DOCK, in which new molecules are built using rules for allowable connections based on known molecules. The method leverages DOCK's advanced scoring and pruning approaches and users can define very specific criteria in terms of properties or features to customize growth toward a particular region of chemical space. The code was validated using three increasingly difficult classes of calculations: (1) Rebuilding known X-ray ligands taken from 663 complexes using only their component parts (focused libraries), (2) construction of new ligands in 57 drug target sites using a library derived from ∼13M drug-like compounds (generic libraries), and (3) application to a challenging protein-protein interface on the viral drug target HIVgp41. The computational testing confirms that the de novo DOCK routines are robust and working as envisioned, and the compelling results highlight the potential utility for designing new molecules against a wide variety of important protein targets. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

  3. Rational design of a cyclin A fluorescent peptide sensor.

    Science.gov (United States)

    Pazos, Elena; Pérez, Miguel; Gutiérrez-de-Terán, Hugo; Orzáez, Mar; Guevara, Tatiana; Mascareñas, José L; Vázquez, M Eugenio

    2011-10-26

    We report the design and development of a fluorescent sensor specifically designed to target cyclin A, a protein that plays a key role in the regulation of the cell cycle. Computational studies provide a molecular picture that explains the observed emission increase, suggesting that the 4-DMAP fluorophore in the peptide is protected from the bulk solvent when inserted into the hydrophobic binding groove of cyclin A.

  4. Biomimetic oligosaccharide and peptide surfactant polymers designed for cardiovascular biomaterials

    Science.gov (United States)

    Ruegsegger, Mark Andrew

    A common problem associated with cardiovascular devices is surface induced thrombosis initiated by the rapid, non-specific adsorption of plasma proteins onto the biomaterial surface. Control of the initial protein adsorption is crucial to achieve the desired longevity of the implanted biomaterial. The cell membrane glycocalyx acts as a non-thrombogenic interface through passive (dense oligosaccharide structures) and active (ligand/receptor interactions) mechanisms. This thesis is designed to investigate biomimicry of the cell glycocalyx to minimize non-specific protein adsorption and promote specific ligand/receptor interactions. Biomimetic macromolecules were designed through the molecular-scale engineering of polymer surfactants, utilizing a poly(vinyl amine) (PVAm) backbone to which hydrophilic (dextran, maltose, peptide) and hydrophobic alkyl (hexanoyl or hexanal) chains are simultaneously attached. The structure was controlled through the molar feed ratio of hydrophobic-to-hydrophilic groups, which also provided control of the solution and surface-active properties. To mimic passive properties, a series of oligomaltose surfactants were synthesized with increasing saccharide length (n = 2, 7, 15 where n is number of glucose units) to investigate the effect of coating height on protein adsorption. The surfactants were characterized by infra red (IR) and nuclear magnetic resonance (NMR) spectroscopies for structural properties and atomic force microscopy (AFM) and contact angle goniometry for surface activity. Protein adsorption under dynamic flow (5 dyn/cm2) was reduced by 85%--95% over the bare hydrophobic substrate; platelet adhesion dropped by ˜80% compared to glass. Peptide ligands were incorporated into the oligosaccharide surfactant to promote functional activity of the passive coating. The surfactants were synthesized to contain 0%, 25%, 50%, 75%, and 100% peptide ligand density and were stable on hydrophobic surfaces. The peptide surface density was

  5. Functional Self-Assembling Peptide Nanofiber Hydrogels Designed for Nerve Degeneration.

    Science.gov (United States)

    Sun, Yuqiao; Li, Wen; Wu, Xiaoli; Zhang, Na; Zhang, Yongnu; Ouyang, Songying; Song, Xiyong; Fang, Xinyu; Seeram, Ramakrishna; Xue, Wei; He, Liumin; Wu, Wutian

    2016-01-27

    Self-assembling peptide (SAP) RADA16-I (Ac-(RADA)4-CONH2) has been suffering from a main drawback associated with low pH, which damages cells and host tissues upon direct exposure. In this study, we presented a strategy to prepare nanofiber hydrogels from two designer SAPs at neutral pH. RADA16-I was appended with functional motifs containing cell adhesion peptide RGD and neurite outgrowth peptide IKVAV. The two SAPs were specially designed to have opposite net charges at neutral pH, the combination of which created a nanofiber hydrogel (-IKVAV/-RGD) characterized by significantly higher G' than G″ in a viscoelasticity examination. Circular dichroism, Fourier transform infrared spectroscopy, and Raman measurements were performed to investigate the secondary structure of the designer SAPs, indicating that both the hydrophobic/hydrophilic properties and electrostatic interactions of the functional motifs play an important role in the self-assembling behavior of the designer SAPs. The neural progenitor cells (NPCs)/stem cells (NSCs) fully embedded in the 3D-IKVAV/-RGD nanofiber hydrogel survived, whereas those embedded within the RADA 16-I hydrogel hardly survived. Moreover, the -IKVAV/-RGD nanofiber hydrogel supported NPC/NSC neuron and astrocyte differentiation in a 3D environment without adding extra growth factors. Studies of three nerve injury models, including sciatic nerve defect, intracerebral hemorrhage, and spinal cord transection, indicated that the designer -IKVAV/-RGD nanofiber hydrogel provided a more permissive environment for nerve regeneration than the RADA 16-I hydrogel. Therefore, we reported a new mechanism that might be beneficial for the synthesis of SAPs for in vitro 3D cell culture and nerve regeneration.

  6. Antimicrobial activity predictors benchmarking analysis using shuffled and designed synthetic peptides.

    Science.gov (United States)

    Porto, William F; Pires, Állan S; Franco, Octavio L

    2017-08-07

    The antimicrobial activity prediction tools aim to help the novel antimicrobial peptides (AMP) sequences discovery, utilizing machine learning methods. Such approaches have gained increasing importance in the generation of novel synthetic peptides by means of rational design techniques. This study focused on predictive ability of such approaches to determine the antimicrobial sequence activities, which were previously characterized at the protein level by in vitro studies. Using four web servers and one standalone software, we evaluated 78 sequences generated by the so-called linguistic model, being 40 designed and 38 shuffled sequences, with ∼60 and ∼25% of identity to AMPs, respectively. The ab initio molecular modelling of such sequences indicated that the structure does not affect the predictions, as both sets present similar structures. Overall, the systems failed on predicting shuffled versions of designed peptides, as they are identical in AMPs composition, which implies in accuracies below 30%. The prediction accuracy is negatively affected by the low specificity of all systems here evaluated, as they, on the other hand, reached 100% of sensitivity. Our results suggest that complementary approaches with high specificity, not necessarily high accuracy, should be developed to be used together with the current systems, overcoming their limitations. Copyright © 2017 Elsevier Ltd. All rights reserved.

  7. Design of Peptide Immunotherapies for MHC Class-II-Associated Autoimmune Disorders

    Directory of Open Access Journals (Sweden)

    Masha Fridkis-Hareli

    2013-01-01

    Full Text Available Autoimmune disorders, that occur when autoreactive immune cells are induced to activate their responses against self-tissues, affect one percent of the world population and represent one of the top 10 leading causes of death. The major histocompatibility complex (MHC is a principal susceptibility locus for many human autoimmune diseases, in which self-tissue antigens providing targets for pathogenic lymphocytes are bound to HLA molecules encoded by disease-associated alleles. In spite of the attempts to design strategies for inhibition of antigen presentation targeting the MHC-peptide/TCR complex via generation of blocking antibodies, altered peptide ligands (APL, or inhibitors of costimulatory molecules, potent therapies with minimal side effects have yet to be developed. Copaxone (glatiramer acetate, GA is a random synthetic amino acid copolymer that reduces the relapse rate by about 30% in relapsing-remitting multiple sclerosis (MS patients. Based on the elucidated binding motifs of Copaxone and of the anchor residues of the immunogenic myelin basic protein (MBP peptide to HLA-DR molecules, novel copolymers have been designed and proved to be more effective in suppressing MS-like disease in mice. In this report, we describe the rationale for design of second-generation synthetic random copolymers as candidate drugs for a number of MHC class-II-associated autoimmune disorders.

  8. Designing peptide inhibitor of insulin receptor to induce diabetes mellitus type 2 in animal model Mus musculus.

    Science.gov (United States)

    Permatasari, Galuh W; Utomo, Didik H; Widodo

    2016-10-01

    A designing peptide as agent for inducing diabetes mellitus type 2 (T2DM) in an animal model is challenging. The computational approach provides a sophisticated tool to design a functional peptide that may block the insulin receptor activity. The peptide that able to inhibit the binding between insulin and insulin receptor is a warrant for inducing T2DM. Therefore, we designed a potential peptide inhibitor of insulin receptor as an agent to generate T2DM animal model by bioinformatics approach. The peptide has been developed based on the structure of insulin receptor binding site of insulin and then modified it to obtain the best properties of half life, hydrophobicity, antigenicity, and stability binding into insulin receptor. The results showed that the modified peptide has characteristics 100h half-life, high-affinity -95.1±20, and high stability 28.17 in complex with the insulin receptor. Moreover, the modified peptide has molecular weight 4420.8g/Mol and has no antigenic regions. Based on the molecular dynamic simulation, the complex of modified peptide-insulin receptor is more stable than the commercial insulin receptor blocker. This study suggested that the modified peptide has the promising performance to block the insulin receptor activity that potentially induce diabetes mellitus type 2 in mice. Copyright © 2016 Elsevier Ltd. All rights reserved.

  9. Preparation of ACE Inhibitory Peptides from Mytilus coruscus Hydrolysate Using Uniform Design

    Directory of Open Access Journals (Sweden)

    Jin-Chao Wu

    2013-01-01

    Full Text Available The angiotensin-I-converting enzyme (ACE inhibitory peptides from mussel, Mytilus coruscus, were investigated and the variable factors, protease concentration, hydrolysis time, pH, and temperature, were optimized using Uniform Design, a new statistical experimental method. The results proved that the hydrolysate of alkali proteases had high ACE-inhibitory activity, especially the alkali protease E1. Optimization by Uniform Design showed that the best hydrolysis conditions for preparation of ACE-inhibitory peptides from Mytilus coruscus were protease concentration of 36.0 U/mL, hydrolysis time of 2.7 hours, pH 8.2, and Temperature at 59.5°C, respectively. The verification experiments under optimum conditions showed that the ACE-inhibitory activity (91.3% were agreed closely with the predicted activity of 90.7%. The amino acid composition analysis of Mytilus coruscus ACE-inhibitory peptides proved that it had high percent of lysine, leucine, glycine, aspartic acid, and glutamic acid.

  10. De novo design of an RNA tile that self-assembles into a homo-octameric nanoprism

    Science.gov (United States)

    Yu, Jinwen; Liu, Zhiyu; Jiang, Wen; Wang, Guansong; Mao, Chengde

    2015-01-01

    Rational, de novo design of RNA nanostructures can potentially integrate a wide array of structural and functional diversities. Such nanostructures have great promises in biomedical applications. Despite impressive progress in this field, all RNA building blocks (or tiles) reported so far are not geometrically well defined. They are generally flexible and can only assemble into a mixture of complexes with different sizes. To achieve defined structures, multiple tiles with different sequences are needed. In this study, we design an RNA tile that can homo-oligomerize into a uniform RNA nanostructure. The designed RNA nanostructure is characterized by gel electrophoresis, atomic force microscopy and cryogenic electron microscopy imaging. We believe that development along this line would help RNA nanotechnology to reach the structural control that is currently associated with DNA nanotechnology.

  11. A folding pathway for betapep-4 peptide 33mer: from unfolded monomers and beta-sheet sandwich dimers to well-structured tetramers.

    OpenAIRE

    Mayo, K. H.; Ilyina, E.

    1998-01-01

    It was recently reported that a de novo designed peptide 33mer, betapep-4, can form well-structured beta-sheet sandwich tetramers (Ilyina E, Roongta V, Mayo KH, 1997b, Biochemistry 36:5245-5250). For insight into the pathway of betapep-4 folding, the present study investigates the concentration dependence of betapep-4 self-association by using 1H-NMR pulsed-field gradient (PFG)-NMR diffusion measurements, and circular dichroism. Downfield chemically shifted alphaH resonances, found to arise o...

  12. Cloning an artificial gene encoding angiostatic anginex: From designed peptide to functional recombinant protein

    International Nuclear Information System (INIS)

    Brandwijk, Ricardo J.M.G.E.; Nesmelova, Irina; Dings, Ruud P.M.; Mayo, Kevin H.; Thijssen, Victor L.J.L.; Griffioen, Arjan W.

    2005-01-01

    Anginex, a designed peptide 33-mer, is a potent angiogenesis inhibitor and anti-tumor agent in vivo. Anginex functions by inhibiting endothelial cell (EC) proliferation and migration leading to detachment and apoptosis of activated EC's. To better understand tumor endothelium targeting properties of anginex and enable its use in gene therapy, we constructed an artificial gene encoding the biologically exogenous peptide and produced the protein recombinantly in Pichia pastoris. Mass spectrometry shows recombinant anginex to be a dimer and circular dichroism shows the recombinant protein folds with β-strand structure like the synthetic peptide. Moreover, like parent anginex, the recombinant protein is active at inhibiting EC growth and migration, as well as inhibiting angiogenesis in vivo in the chorioallantoic membrane of the chick embryo. This study demonstrated that it is possible to produce a functionally active protein version of a rationally designed peptide, using an artificial gene and the recombinant protein approach

  13. NovoPen Echo® insulin delivery device

    Directory of Open Access Journals (Sweden)

    Hyllested-Winge J

    2016-01-01

    Full Text Available Jacob Hyllested-Winge,1 Thomas Sparre,2 Line Kynemund Pedersen2 1Novo Nordisk Pharma Ltd, Tokyo, Japan; 2Novo Nordisk A/S, Søborg, Denmark Abstract: The introduction of insulin pen devices has provided easier, well-tolerated, and more convenient treatment regimens for patients with diabetes mellitus. When compared with vial and syringe regimens, insulin pens offer a greater clinical efficacy, improved quality of life, and increased dosing accuracy, particularly at low doses. The portable and discreet nature of pen devices reduces the burden on the patient, facilitates adherence, and subsequently contributes to the improvement in glycemic control. NovoPen Echo® is one of the latest members of the NovoPen® family that has been specifically designed for the pediatric population and is the first to combine half-unit increment (=0.5 U of insulin dosing with a simple memory function. The half-unit increment dosing amendments and accurate injection of 0.5 U of insulin are particularly beneficial for children (and insulin-sensitive adults/elders, who often require small insulin doses. The memory function can be used to record the time and amount of the last dose, reducing the fear of double dosing or missing a dose. The memory function also provides parents with extra confidence and security that their child is taking insulin at the correct doses and times. NovoPen Echo is a lightweight, durable insulin delivery pen; it is available in two different colors, which may help to distinguish between different types of insulin, providing more confidence for both users and caregivers. Studies have demonstrated a high level of patient satisfaction, with 80% of users preferring NovoPen Echo to other pediatric insulin pens. Keywords: NovoPen Echo®, memory function, half-unit increment dosing, adherence, children, adolescents 

  14. Antimicrobial Peptides Derived from Fusion Peptides of Influenza A Viruses, a Promising Approach to Designing Potent Antimicrobial Agents.

    Science.gov (United States)

    Wang, Jingyu; Zhong, Wenjing; Lin, Dongguo; Xia, Fan; Wu, Wenjiao; Zhang, Heyuan; Lv, Lin; Liu, Shuwen; He, Jian

    2015-10-01

    The emergence and dissemination of antibiotic-resistant bacterial pathogens have spurred the urgent need to develop novel antimicrobial agents with different mode of action. In this respect, we turned several fusogenic peptides (FPs) derived from the hemagglutinin glycoproteins (HAs) of IAV into potent antibacterials by replacing the negatively or neutrally charged residues of FPs with positively charged lysines. Their antibacterial activities were evaluated by testing the MICs against a panel of bacterial strains including S. aureus, S. mutans, P. aeruginosa, and E. coli. The results showed that peptides HA-FP-1, HA-FP-2-1, and HA-FP-3-1 were effective against both Gram-positive and Gram-negative bacteria with MICs ranging from 1.9 to 16.0 μm, while the toxicities toward mammalian cells were low. In addition, the mode of action and the secondary structure of these peptides were also discussed. These data not only provide several potent peptides displaying promising potential in development as broad antimicrobial agents, but also present a useful strategy in designing new antimicrobial agents. © 2015 John Wiley & Sons A/S.

  15. Rational Design of Cyclic Antimicrobial Peptides Based on BPC194 and BPC198

    Directory of Open Access Journals (Sweden)

    Anna D. Cirac

    2017-06-01

    Full Text Available A strategy for the design of antimicrobial cyclic peptides derived from the lead compounds c(KKLKKFKKLQ (BPC194 and c(KLKKKFKKLQ (BPC198 is reported. First, the secondary β-structure of BPC194 and BPC198 was analyzed by carrying out molecular dynamics (MD simulations. Then, based on the sequence pattern and the β-structure of BPC194 or BPC198, fifteen analogues were designed and synthesized on solid-phase. The best peptides (BPC490, BPC918, and BPC924 showed minimum inhibitory concentration (MIC values <6.2 μM against Pseudomonas syringae pv. syringae and Xanthomonas axonopodis pv. vesicatoria, and an MIC value of 12.5 to 25 μM against Erwinia amylovora, being as active as BPC194 and BPC198. Interestingly, these three analogues followed the structural pattern defined from the MD simulations of the parent peptides. Thus, BPC490 maintained the parallel alignment of the hydrophilic pairs K1–K8, K2–K7, and K4–K5, whereas BPC918 and BPC924 included the two hydrophilic interactions K3–Q10 and K5–K8. In short, MD simulations have proved to be very useful for ascertaining the structural features of cyclic peptides that are crucial for their biological activity. Such approaches could be further employed for the development of new antibacterial cyclic peptides.

  16. Towards the Development of Synthetic Antibiotics: Designs Inspired by Natural Antimicrobial Peptides.

    Science.gov (United States)

    Azmi, Fazren; Skwarczynski, Mariusz; Toth, Istvan

    2016-01-01

    Virtually every living organism produces gene-encoded antimicrobial peptides (AMPs) that provide an immediate defence against pathogen invasion. Many AMPs have been isolated and used as antibiotics that are effective against multidrug-resistant bacteria. Although encouraging, AMPs have such poor drug-like properties that their application for clinical use is restricted. In turn, this has diverted research to the development of synthetic molecules that retain the therapeutic efficacy of AMPs but are endowed with greater biological stability and safety profiles. Most of the synthetic molecules, either based on a peptidic or non-peptidic scaffold, have been designed to mimic the amphiphilic properties of native AMPs, which are widely believed to be the key determinant of their antibacterial activity. In this review, the structural, chemical and biophysical features that govern the biological activities of various synthetic designs are discussed extensively. Recent innovative approaches from the literature that exhibit novel concepts towards the development of new synthetic antibacterial agents, including the engineered delivery platform incorporated with AMP mimetics, are also emphasised.

  17. Computational design of selective peptides to discriminate between similar PDZ domains in an oncogenic pathway.

    Science.gov (United States)

    Zheng, Fan; Jewell, Heather; Fitzpatrick, Jeremy; Zhang, Jian; Mierke, Dale F; Grigoryan, Gevorg

    2015-01-30

    Reagents that target protein-protein interactions to rewire signaling are of great relevance in biological research. Computational protein design may offer a means of creating such reagents on demand, but methods for encoding targeting selectivity are sorely needed. This is especially challenging when targeting interactions with ubiquitous recognition modules--for example, PDZ domains, which bind C-terminal sequences of partner proteins. Here we consider the problem of designing selective PDZ inhibitor peptides in the context of an oncogenic signaling pathway, in which two PDZ domains (NHERF-2 PDZ2-N2P2 and MAGI-3 PDZ6-M3P6) compete for a receptor C-terminus to differentially modulate oncogenic activities. Because N2P2 has been shown to increase tumorigenicity and M3P6 to decreases it, we sought to design peptides that inhibit N2P2 without affecting M3P6. We developed a structure-based computational design framework that models peptide flexibility in binding yet is efficient enough to rapidly analyze tradeoffs between affinity and selectivity. Designed peptides showed low-micromolar inhibition constants for N2P2 and no detectable M3P6 binding. Peptides designed for reverse discrimination bound M3P6 tighter than N2P2, further testing our technology. Experimental and computational analysis of selectivity determinants revealed significant indirect energetic coupling in the binding site. Successful discrimination between N2P2 and M3P6, despite their overlapping binding preferences, is highly encouraging for computational approaches to selective PDZ targeting, especially because design relied on a homology model of M3P6. Still, we demonstrate specific deficiencies of structural modeling that must be addressed to enable truly robust design. The presented framework is general and can be applied in many scenarios to engineer selective targeting. Copyright © 2014 Elsevier Ltd. All rights reserved.

  18. Extreme-Scale De Novo Genome Assembly

    Energy Technology Data Exchange (ETDEWEB)

    Georganas, Evangelos [Intel Corporation, Santa Clara, CA (United States); Hofmeyr, Steven [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Joint Genome Inst.; Egan, Rob [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Computational Research Division; Buluc, Aydin [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Joint Genome Inst.; Oliker, Leonid [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Joint Genome Inst.; Rokhsar, Daniel [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Computational Research Division; Yelick, Katherine [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Joint Genome Inst.

    2017-09-26

    De novo whole genome assembly reconstructs genomic sequence from short, overlapping, and potentially erroneous DNA segments and is one of the most important computations in modern genomics. This work presents HipMER, a high-quality end-to-end de novo assembler designed for extreme scale analysis, via efficient parallelization of the Meraculous code. Genome assembly software has many components, each of which stresses different components of a computer system. This chapter explains the computational challenges involved in each step of the HipMer pipeline, the key distributed data structures, and communication costs in detail. We present performance results of assembling the human genome and the large hexaploid wheat genome on large supercomputers up to tens of thousands of cores.

  19. On-target sample preparation of 4-sulfophenyl isothiocyanate-derivatized peptides using AnchorChip Targets

    DEFF Research Database (Denmark)

    Zhang, Xumin; Rogowska-Wrzesinska, Adelina; Roepstorff, Peter

    2008-01-01

    De novo sequencing of tryptic peptides by post source decay (PSD) or collision induced dissociation (CID) analysis using MALDI TOF-TOF instruments is due to the easy interpretation facilitated by the introduction of N-terminal sulfonated derivatives. Recently, a stable and cheap reagent, 4...

  20. Machine learning in the rational design of antimicrobial peptides.

    Science.gov (United States)

    Rondón-Villarreal, Paola; Sierra, Daniel A; Torres, Rodrigo

    2014-01-01

    One of the most important public health issues is the microbial and bacterial resistance to conventional antibiotics by pathogen microorganisms. In recent years, many researches have been focused on the development of new antibiotics. Among these, antimicrobial peptides (AMPs) have raised as a promising alternative to combat antibioticresistant microorganisms. For this reason, many theoretical efforts have been done in the development of new computational tools for the rational design of both better and effective AMPs. In this review, we present an overview of the rational design of AMPs using machine learning techniques and new research fields.

  1. Trends in the Design and Development of Specific Aptamers Against Peptides and Proteins.

    Science.gov (United States)

    Tabarzad, Maryam; Jafari, Marzieh

    2016-04-01

    Aptamers are single stranded oligonucleotides, comparable to monoclonal antibodies (mAbs) in selectivity and affinity and have significant strategic properties in design, development and applications more than mAbs. Ease of design and development, simple chemical modification and the attachment of functional groups, easily handling and more adaptability with analytical methods, small size and adaptation with nanostructures are the valuable characteristics of aptamers in comparison to large protein based ligands. Among a broad range of targets that their specific aptamers developed, proteins and peptides have significant position according to the number of related studies performed so far. Since proteins control many of important physiological and pathological incidents in the living organisms, particularly human beings and because of the benefits of aptamers in clinical and analytical applications, aptamer related technologies in the field of proteins and peptides are under progress, exclusively. Currently, there is only one FDA approved therapeutic aptamer in the pharmaceutical market, which is specific to vascular endothelial growth factor and is prescribed for age related macular degenerative disease. Additionally, there are several aptamers in the different phases of clinical trials. Almost all of these aptamers are specific to clinically important peptide or protein targets. In addition, the application of protein specific aptamers in the design and development of targeted drug delivery systems and diagnostic biosensors is another interesting field of aptamer technology. In this review, significant efforts related to development and applications of aptamer technologies in proteins and peptides sciences were considered to emphasis on the importance of aptamers in medicinal and clinical applications.

  2. Modular Engineering Concept at Novo Nordisk Engineering

    DEFF Research Database (Denmark)

    Moelgaard, Gert; Miller, Thomas Dedenroth

    1997-01-01

    This report describes the concept of a new engineering method at Novo Nordisk Engineering: Modular Engineering (ME). Three tools are designed to support project phases with different levels of detailing and abstraction. ME supports a standard, cross-functional breakdown of projects that facilitates...

  3. Peptide/protein-polymer conjugates: synthetic strategies and design concepts.

    Science.gov (United States)

    Gauthier, Marc A; Klok, Harm-Anton

    2008-06-21

    This feature article provides a compilation of tools available for preparing well-defined peptide/protein-polymer conjugates, which are defined as hybrid constructs combining (i) a defined number of peptide/protein segments with uniform chain lengths and defined monomer sequences (primary structure) with (ii) a defined number of synthetic polymer chains. The first section describes methods for post-translational, or direct, introduction of chemoselective handles onto natural or synthetic peptides/proteins. Addressed topics include the residue- and/or site-specific modification of peptides/proteins at Arg, Asp, Cys, Gln, Glu, Gly, His, Lys, Met, Phe, Ser, Thr, Trp, Tyr and Val residues and methods for producing peptides/proteins containing non-canonical amino acids by peptide synthesis and protein engineering. In the second section, methods for introducing chemoselective groups onto the side-chain or chain-end of synthetic polymers produced by radical, anionic, cationic, metathesis and ring-opening polymerization are described. The final section discusses convergent and divergent strategies for covalently assembling polymers and peptides/proteins. An overview of the use of chemoselective reactions such as Heck, Sonogashira and Suzuki coupling, Diels-Alder cycloaddition, Click chemistry, Staudinger ligation, Michael's addition, reductive alkylation and oxime/hydrazone chemistry for the convergent synthesis of peptide/protein-polymer conjugates is given. Divergent approaches for preparing peptide/protein-polymer conjugates which are discussed include peptide synthesis from synthetic polymer supports, polymerization from peptide/protein macroinitiators or chain transfer agents and the polymerization of peptide side-chain monomers.

  4. High-resolution protein design with backbone freedom.

    Science.gov (United States)

    Harbury, P B; Plecs, J J; Tidor, B; Alber, T; Kim, P S

    1998-11-20

    Recent advances in computational techniques have allowed the design of precise side-chain packing in proteins with predetermined, naturally occurring backbone structures. Because these methods do not model protein main-chain flexibility, they lack the breadth to explore novel backbone conformations. Here the de novo design of a family of alpha-helical bundle proteins with a right-handed superhelical twist is described. In the design, the overall protein fold was specified by hydrophobic-polar residue patterning, whereas the bundle oligomerization state, detailed main-chain conformation, and interior side-chain rotamers were engineered by computational enumerations of packing in alternate backbone structures. Main-chain flexibility was incorporated through an algebraic parameterization of the backbone. The designed peptides form alpha-helical dimers, trimers, and tetramers in accord with the design goals. The crystal structure of the tetramer matches the designed structure in atomic detail.

  5. Antiviral activity of α-helical stapled peptides designed from the HIV-1 capsid dimerization domain

    Directory of Open Access Journals (Sweden)

    Cowburn David

    2011-05-01

    Full Text Available Abstract Background The C-terminal domain (CTD of HIV-1 capsid (CA, like full-length CA, forms dimers in solution and CTD dimerization is a major driving force in Gag assembly and maturation. Mutations of the residues at the CTD dimer interface impair virus assembly and render the virus non-infectious. Therefore, the CTD represents a potential target for designing anti-HIV-1 drugs. Results Due to the pivotal role of the dimer interface, we reasoned that peptides from the α-helical region of the dimer interface might be effective as decoys to prevent CTD dimer formation. However, these small peptides do not have any structure in solution and they do not penetrate cells. Therefore, we used the hydrocarbon stapling technique to stabilize the α-helical structure and confirmed by confocal microscopy that this modification also made these peptides cell-penetrating. We also confirmed by using isothermal titration calorimetry (ITC, sedimentation equilibrium and NMR that these peptides indeed disrupt dimer formation. In in vitro assembly assays, the peptides inhibited mature-like virus particle formation and specifically inhibited HIV-1 production in cell-based assays. These peptides also showed potent antiviral activity against a large panel of laboratory-adapted and primary isolates, including viral strains resistant to inhibitors of reverse transcriptase and protease. Conclusions These preliminary data serve as the foundation for designing small, stable, α-helical peptides and small-molecule inhibitors targeted against the CTD dimer interface. The observation that relatively weak CA binders, such as NYAD-201 and NYAD-202, showed specificity and are able to disrupt the CTD dimer is encouraging for further exploration of a much broader class of antiviral compounds targeting CA. We cannot exclude the possibility that the CA-based peptides described here could elicit additional effects on virus replication not directly linked to their ability to bind

  6. Thermodynamic description of polymorphism in Q- and N-rich peptide aggregates revealed by atomistic simulation.

    Science.gov (United States)

    Berryman, Joshua T; Radford, Sheena E; Harris, Sarah A

    2009-07-08

    Amyloid fibrils are long, helically symmetric protein aggregates that can display substantial variation (polymorphism), including alterations in twist and structure at the beta-strand and protofilament levels, even when grown under the same experimental conditions. The structural and thermodynamic origins of this behavior are not yet understood. We performed molecular-dynamics simulations to determine the thermodynamic properties of different polymorphs of the peptide GNNQQNY, modeling fibrils containing different numbers of protofilaments based on the structure of amyloid-like cross-beta crystals of this peptide. We also modeled fibrils with new orientations of the side chains, as well as a de novo designed structure based on antiparallel beta-strands. The simulations show that these polymorphs are approximately isoenergetic under a range of conditions. Structural analysis reveals a dynamic reorganization of electrostatics and hydrogen bonding in the main and side chains of the Gln and Asn residues that characterize this peptide sequence. Q/N-rich stretches are found in several amyloidogenic proteins and peptides, including the yeast prions Sup35-N and Ure2p, as well as in the human poly-Q disease proteins, including the ataxins and huntingtin. Based on our results, we propose that these residues imbue a unique structural plasticity to the amyloid fibrils that they comprise, rationalizing the ability of proteins enriched in these amino acids to form prion strains with heritable and different phenotypic traits.

  7. Design and synthesis of macrocyclic peptidyl hydroxamates as peptide deformylase inhibitors.

    Science.gov (United States)

    Shen, Gang; Zhu, Jinge; Simpson, Anthony M; Pei, Dehua

    2008-05-15

    Macrocyclic peptidyl hydroxamates were designed, synthesized, and evaluated as peptide deformylase (PDF) inhibitors. The most potent compound exhibited tight, slow-binding inhibition of Escherichia coli PDF (K(I)(*)=4.4 nM) and had potent antibacterial activity against Gram-positive bacterium Bacillus subtilis (MIC=2-4 microg/mL).

  8. Self-assembling peptide nanofiber hydrogels in tissue engineering and regenerative medicine: Progress, design guidelines, and applications.

    Science.gov (United States)

    Koutsopoulos, Sotirios

    2016-04-01

    Until the mid-1980s, mainly biologists were conducting peptide research. This changed with discoveries that opened new paths of research involving the use of peptides in bioengineering, biotechnology, biomedicine, nanotechnology, and bioelectronics. Peptide engineering and rational design of novel peptide sequences with unique and tailor-made properties further expanded the field. The discovery of short self-assembling peptides, which upon association form well-defined supramolecular architectures, created new and exciting areas of research. Depending on the amino acid sequence, the pH, and the type of the electrolyte in the medium, peptide self-assembly leads to the formation of nanofibers, which are further organized to form a hydrogel. In this review, the application of ionic complementary peptides which self-assemble to form nanofiber hydrogels for tissue engineering and regenerative medicine will be discussed through a selective presentation of the most important work performed during the last 25 years. © 2016 Wiley Periodicals, Inc.

  9. In silico design and performance of peptide microarrays for breast cancer tumour-auto-antibody testing

    Directory of Open Access Journals (Sweden)

    Andreas Weinhäusel

    2012-06-01

    Full Text Available The simplicity and potential of minimally invasive testing using sera from patients makes auto-antibody based biomarkers a very promising tool for use in cancer diagnostics. Protein microarrays have been used for the identification of such auto-antibody signatures. Because high throughput protein expression and purification is laborious, synthetic peptides might be a good alternative for microarray generation and multiplexed analyses. In this study, we designed 1185 antigenic peptides, deduced from proteins expressed by 642 cDNA expression clones found to be sero-reactive in both breast tumour patients and controls. The sero-reactive proteins and the corresponding peptides were used for the production of protein and peptide microarrays. Serum samples from females with benign and malignant breast tumours and healthy control sera (n=16 per group were then analysed. Correct classification of the serum samples on peptide microarrays were 78% for discrimination of ‘malignant versus healthy controls’, 72% for ‘benign versus malignant’ and 94% for ‘benign versus controls’. On protein arrays, correct classification for these contrasts was 69%, 59% and 59%, respectively. The over-representation analysis of the classifiers derived from class prediction showed enrichment of genes associated with ribosomes, spliceosomes, endocytosis and the pentose phosphate pathway. Sequence analyses of the peptides with the highest sero-reactivity demonstrated enrichment of the zinc-finger domain. Peptides’ sero-reactivities were found negatively correlated with hydrophobicity and positively correlated with positive charge, high inter-residue protein contact energies and a secondary structure propensity bias. This study hints at the possibility of using in silico designed antigenic peptide microarrays as an alternative to protein microarrays for the improvement of tumour auto-antibody based diagnostics.

  10. PRO_LIGAND: an approach to de novo molecular design. 2. Design of novel molecules from molecular field analysis (MFA) models and pharmacophores.

    Science.gov (United States)

    Waszkowycz, B; Clark, D E; Frenkel, D; Li, J; Murray, C W; Robson, B; Westhead, D R

    1994-11-11

    A computational approach for molecular design, PRO_LIGAND, has been developed within the PROMETHEUS molecular design and simulation system in order to provide a unified framework for the de novo generation of diverse molecules which are either similar or complementary to a specified target. In this instance, the target is a pharmacophore derived from a series of active structures either by a novel interpretation of molecular field analysis data or by a pharmacophore-mapping procedure based on clique detection. After a brief introduction to PRO_LIGAND, a detailed description is given of the two pharmacophore generation procedures and their abilities are demonstrated by the elucidation of pharmacophores for steroid binding and ACE inhibition, respectively. As a further indication of its efficacy in aiding the rational drug design process, PRO_LIGAND is then employed to build novel organic molecules to satisfy the physicochemical constraints implied by the pharmacophores.

  11. Optimization and high-throughput screening of antimicrobial peptides.

    Science.gov (United States)

    Blondelle, Sylvie E; Lohner, Karl

    2010-01-01

    While a well-established process for lead compound discovery in for-profit companies, high-throughput screening is becoming more popular in basic and applied research settings in academia. The development of combinatorial libraries combined with easy and less expensive access to new technologies have greatly contributed to the implementation of high-throughput screening in academic laboratories. While such techniques were earlier applied to simple assays involving single targets or based on binding affinity, they have now been extended to more complex systems such as whole cell-based assays. In particular, the urgent need for new antimicrobial compounds that would overcome the rapid rise of drug-resistant microorganisms, where multiple target assays or cell-based assays are often required, has forced scientists to focus onto high-throughput technologies. Based on their existence in natural host defense systems and their different mode of action relative to commercial antibiotics, antimicrobial peptides represent a new hope in discovering novel antibiotics against multi-resistant bacteria. The ease of generating peptide libraries in different formats has allowed a rapid adaptation of high-throughput assays to the search for novel antimicrobial peptides. Similarly, the availability nowadays of high-quantity and high-quality antimicrobial peptide data has permitted the development of predictive algorithms to facilitate the optimization process. This review summarizes the various library formats that lead to de novo antimicrobial peptide sequences as well as the latest structural knowledge and optimization processes aimed at improving the peptides selectivity.

  12. Therapeutic peptides for cancer therapy. Part I - peptide inhibitors of signal transduction cascades.

    Science.gov (United States)

    Bidwell, Gene L; Raucher, Drazen

    2009-10-01

    Therapeutic peptides have great potential as anticancer agents owing to their ease of rational design and target specificity. However, their utility in vivo is limited by low stability and poor tumor penetration. The authors review the development of peptide inhibitors with potential for cancer therapy. Peptides that inhibit signal transduction cascades are discussed. The authors searched Medline for articles concerning the development of therapeutic peptides and their delivery. Given our current knowledge of protein sequences, structures and interaction interfaces, therapeutic peptides that inhibit interactions of interest are easily designed. These peptides are advantageous because they are highly specific for the interaction of interest, and they are much more easily developed than small molecule inhibitors of the same interactions. The main hurdle to application of peptides for cancer therapy is their poor pharmacokinetic and biodistribution parameters. Therefore, successful development of peptide delivery vectors could potentially make possible the use of this new and very promising class of anticancer agents.

  13. Protein design and engineering of a de novo pathway for microbial production of 1,3-propanediol from glucose.

    Science.gov (United States)

    Chen, Zhen; Geng, Feng; Zeng, An-Ping

    2015-02-01

    Protein engineering to expand the substrate spectrum of native enzymes opens new possibilities for bioproduction of valuable chemicals from non-natural pathways. No natural microorganism can directly use sugars to produce 1,3-propanediol (PDO). Here, we present a de novo route for the biosynthesis of PDO from sugar, which may overcome the mentioned limitations by expanding the homoserine synthesis pathway. The accomplishment of pathway from homoserine to PDO is achieved by protein engineering of glutamate dehydrogenase (GDH) and pyruvate decarboxylase to sequentially convert homoserine to 4-hydroxy-2-ketobutyrate and 3-hydroxypropionaldehyde. The latter is finally converted to PDO by using a native alcohol dehydrogenase. In this work, we report on experimental accomplishment of this non-natural pathway, especially by protein engineering of GDH for the key step of converting homoserine to 4-hydroxy-2-ketobutyrate. These results show the feasibility and significance of protein engineering for de novo pathway design and overproduction of desired industrial products. Copyright © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  14. A novel TNFα antagonizing peptide-Fc fusion protein designed based on CDRs of TNFα neutralizing monoclonal antibody

    International Nuclear Information System (INIS)

    Qin Weisong; Feng Jiannan; Zhang Wei; Li Yan; Shen, Beifen

    2004-01-01

    The variable regions of antibody molecules bind antigens with high affinity and specificity. The binding sites are imparted largely to the hypervariable portions (i.e., CDRs) of the variable region. Peptides derived from CDRs can bind antigen with similar specificity acting as mimic of antibody and become drug-designing core, although with markedly lower affinity. In order to increase the affinity and bioactivity, in this study, a novel peptide (PT) designed on CDRs of a TNFα neutralizing monoclonal antibody Z12 was linked with Fc fragment of human IgG1. The interaction mode of PT-linker-Fc (PLF) with TNFα was analyzed with computer-guided molecular modeling method. After expression in Escherichia coli and purification, recombinant PT-linker-Fc could bind directly with the TNFα coated on the ELISA plates. Furthermore, PLF could competitively inhibit the binding of Z12 to TNFα and also inhibit the TNFα-induced cytotoxicity on L929 cells. The TNFα antagonizing activity of PLF was significantly higher than that of the free peptide. This study highlights the potential of human Fc to enhance the potency of peptides designed on the CDRs of antibodies and could be useful in developing new TNFα antagonists

  15. Airline Maintenance Manpower Optimization from the De Novo Perspective

    Science.gov (United States)

    Liou, James J. H.; Tzeng, Gwo-Hshiung

    Human resource management (HRM) is an important issue for today’s competitive airline marketing. In this paper, we discuss a multi-objective model designed from the De Novo perspective to help airlines optimize their maintenance manpower portfolio. The effectiveness of the model and solution algorithm is demonstrated in an empirical study of the optimization of the human resources needed for airline line maintenance. Both De Novo and traditional multiple objective programming (MOP) methods are analyzed. A comparison of the results with those of traditional MOP indicates that the proposed model and solution algorithm does provide better performance and an improved human resource portfolio.

  16. Natriuretic peptides and integrated risk assessment for cardiovascular disease

    DEFF Research Database (Denmark)

    Willeit, Peter; Kaptoge, S; Welsh, P.

    2016-01-01

    samples and collection of data from studies identified through a systematic search of the literature (PubMed, Scientific Citation Index Expanded, and Embase) for articles published up to Sept 4, 2014, using search terms related to natriuretic peptide family members and the primary outcomes......BACKGROUND: Guidelines for primary prevention of cardiovascular diseases focus on prediction of coronary heart disease and stroke. We assessed whether or not measurement of N-terminal-pro-B-type natriuretic peptide (NT-proBNP) concentration could enable a more integrated approach than at present...... by predicting heart failure and enhancing coronary heart disease and stroke risk assessment. METHODS: In this individual-participant-data meta-analysis, we generated and harmonised individual-participant data from relevant prospective studies via both de-novo NT-proBNP concentration measurement of stored...

  17. A Simple Three-Step Method for Design and Affinity Testing of New Antisense Peptides: An Example of Erythropoietin

    Directory of Open Access Journals (Sweden)

    Nikola Štambuk

    2014-05-01

    Full Text Available Antisense peptide technology is a valuable tool for deriving new biologically active molecules and performing peptide–receptor modulation. It is based on the fact that peptides specified by the complementary (antisense nucleotide sequences often bind to each other with a higher specificity and efficacy. We tested the validity of this concept on the example of human erythropoietin, a well-characterized and pharmacologically relevant hematopoietic growth factor. The purpose of the work was to present and test simple and efficient three-step procedure for the design of an antisense peptide targeting receptor-binding site of human erythropoietin. Firstly, we selected the carboxyl-terminal receptor binding region of the molecule (epitope as a template for the antisense peptide modeling; Secondly, we designed an antisense peptide using mRNA transcription of the epitope sequence in the 3'→5' direction and computational screening of potential paratope structures with BLAST; Thirdly, we evaluated sense–antisense (epitope–paratope peptide binding and affinity by means of fluorescence spectroscopy and microscale thermophoresis. Both methods showed similar Kd values of 850 and 816 µM, respectively. The advantages of the methods were: fast screening with a small quantity of the sample needed, and measurements done within the range of physicochemical parameters resembling physiological conditions. Antisense peptides targeting specific erythropoietin region(s could be used for the development of new immunochemical methods. Selected antisense peptides with optimal affinity are potential lead compounds for the development of novel diagnostic substances, biopharmaceuticals and vaccines.

  18. Functional mimicry of a discontinuous antigenic site by a designed synthetic peptide

    NARCIS (Netherlands)

    Villen, J.; Borras, E.; Schaaper, W.M.M.; Meloen, R.H.; Davila, M.; Domingo, E.; Giralt, E.; Andreu, D.

    2002-01-01

    Functional reproduction of the discontinuous antigenic site D of foot-and-mouth disease virus (FMDV) has been achieved by means of synthetic peptide constructions that integrate each of the three protein loops that define the antigenic site into a single molecule. The site D mimics were designed on

  19. Design, Synthesis and Evaluation of Branched RRWQWR-Based Peptides as Antibacterial Agents Against Clinically Relevant Gram-Positive and Gram-Negative Pathogens

    Directory of Open Access Journals (Sweden)

    Sandra C. Vega

    2018-03-01

    Full Text Available Multidrug resistance of pathogenic bacteria has become a public health crisis that requires the urgent design of new antibacterial drugs such as antimicrobial peptides (AMPs. Seeking to obtain new, lactoferricin B (LfcinB-based synthetic peptides as viable early-stage candidates for future development as AMPs against clinically relevant bacteria, we designed, synthesized and screened three new cationic peptides derived from bovine LfcinB. These peptides contain at least one RRWQWR motif and differ by the copy number (monomeric, dimeric or tetrameric and structure (linear or branched of this motif. They comprise a linear palindromic peptide (RWQWRWQWR, a dimeric peptide (RRWQWR2KAhx and a tetrameric peptide (RRWQWR4K2Ahx2C2. They were screened for antibacterial activity against Enterococcus faecalis (ATCC 29212 and ATCC 51575 strains, Pseudomonas aeruginosa (ATCC 10145 and ATCC 27853 strains and clinical isolates of two Gram-positive bacteria (Enterococcus faecium and Staphylococcus aureus and two Gram-negative bacteria (Klebsiella pneumoniae and Pseudomonas aeruginosa. All three peptides exhibited greater activity than did the reference peptide, LfcinB (17–31, which contains a single linear RRWQWR motif. Against the ATCC reference strains, the three new peptides exhibited minimum inhibitory concentration (MIC50 values of 3.1–198.0 μM and minimum bactericidal concentration (MBC values of 25–200 μM, and against the clinical isolates, MIC50 values of 1.6–75.0 μM and MBC values of 12.5–100 μM. However, the tetrameric peptide was also found to be strongly hemolytic (49.1% at 100 μM. Scanning Electron Microscopy (SEM demonstrated that in the dimeric and tetrameric peptides, the RRWQWR motif is exposed to the pathogen surface. Our results may inform the design of new, RRWQWR-based AMPs.

  20. Investigation of Self-assembly Structure and Properties of a Novel Designed Lego-type Peptide with Double Amphiphilic Surfaces

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Liang [Sichuan University, Sichuan (China); Zhao, Xiao Jun [Massachusetts Institute of Technology, Cambridge (United States)

    2010-12-15

    A typically designed 'Peptide Lego' has two distinct surfaces: a hydrophilic side that contains the complete charge distribution and a hydrophobic side. In this article, we describe the fabrication of a unique lego-type peptide with the AEAEYAKAK sequence. The novel peptide with double amphiphilic surfaces is different from typical peptides due to special arrangement of the residues. The results of CD, FT-IR, AFM and DLS demonstrate that the peptide with the random coil characteristic was able to form stable nanostructures that were mediated by non-covalent interactions in an aqueous solution. The data further indicated that despite its different structure, the peptide was able to undergo self-assembly similar to a typical peptide. In addition, the use of hydrophobic pyrene as a model allowed the peptide to provide a new type of potential nanomaterial for drug delivery. These efforts collectively open up a new direction in the fabrication of nanomaterials that are more perfect and versatile.

  1. Investigation of Self-assembly Structure and Properties of a Novel Designed Lego-type Peptide with Double Amphiphilic Surfaces

    International Nuclear Information System (INIS)

    Wang, Liang; Zhao, Xiao Jun

    2010-01-01

    A typically designed 'Peptide Lego' has two distinct surfaces: a hydrophilic side that contains the complete charge distribution and a hydrophobic side. In this article, we describe the fabrication of a unique lego-type peptide with the AEAEYAKAK sequence. The novel peptide with double amphiphilic surfaces is different from typical peptides due to special arrangement of the residues. The results of CD, FT-IR, AFM and DLS demonstrate that the peptide with the random coil characteristic was able to form stable nanostructures that were mediated by non-covalent interactions in an aqueous solution. The data further indicated that despite its different structure, the peptide was able to undergo self-assembly similar to a typical peptide. In addition, the use of hydrophobic pyrene as a model allowed the peptide to provide a new type of potential nanomaterial for drug delivery. These efforts collectively open up a new direction in the fabrication of nanomaterials that are more perfect and versatile

  2. Pesquisa de novos elementos Pesquisa de novos elementos

    Directory of Open Access Journals (Sweden)

    Gil Mário de Macedo Grassi

    1978-11-01

    Full Text Available The present study deals with the discovery of new elements synthesized by man. The introduction discusses in general the theories about nuclear transmutation, which is the method employed in these syntheses. The study shows the importance of the Periodical Table since it is through this table that one can reach a prevision of new elements and its, properties. The discoveries of the transuranic elements, together wich the data of their first preparations are also tabulated The stability of these elements is also discussed, and future speculations are showedNeste trabalho estuda-se, teoricamente, a descoberta de novos elementos sintetizados pelo homem Na introdução apresentamos um apanhado geral sobre as teorias a respeito da transmutação nuclear, que é o método utilizado nestas sínteses. Em seguida, mostramos a importância da Tabela Periódica, pois é através dela que se chega à previsão dos novos elementos e de suas propriedades. As descobertas dos transurânicos, Já realizadas com êxito, juntamente com os dados de suas primeiras preparações são tabelados. A estabilidade destes novos elementos também é discutida, e apresentadas futuras especulações.

  3. PChopper: high throughput peptide prediction for MRM/SRM transition design

    Directory of Open Access Journals (Sweden)

    Huang Jeffrey T-J

    2011-08-01

    Full Text Available Abstract Background The use of selective reaction monitoring (SRM based LC-MS/MS analysis for the quantification of phosphorylation stoichiometry has been rapidly increasing. At the same time, the number of sites that can be monitored in a single LC-MS/MS experiment is also increasing. The manual processes associated with running these experiments have highlighted the need for computational assistance to quickly design MRM/SRM candidates. Results PChopper has been developed to predict peptides that can be produced via enzymatic protein digest; this includes single enzyme digests, and combinations of enzymes. It also allows digests to be simulated in 'batch' mode and can combine information from these simulated digests to suggest the most appropriate enzyme(s to use. PChopper also allows users to define the characteristic of their target peptides, and can automatically identify phosphorylation sites that may be of interest. Two application end points are available for interacting with the system; the first is a web based graphical tool, and the second is an API endpoint based on HTTP REST. Conclusions Service oriented architecture was used to rapidly develop a system that can consume and expose several services. A graphical tool was built to provide an easy to follow workflow that allows scientists to quickly and easily identify the enzymes required to produce multiple peptides in parallel via enzymatic digests in a high throughput manner.

  4. Identification of single amino acid substitutions (SAAS) in neuraminidase from influenza a virus (H1N1) via mass spectrometry analysis coupled with de novo peptide sequencing.

    Science.gov (United States)

    Peng, Qisheng; Wang, Zijian; Wu, Donglin; Li, Xiaoou; Liu, Xiaofeng; Sun, Wanchun; Liu, Ning

    2016-08-01

    Amino acid substitutions in the neuraminidase of the influenza virus are the main cause of the emergence of resistance to zanamivir or oseltamivir during seasonal influenza treatment; they are the result of non-synonymous mutations in the viral genome that can be successfully detected by polymer chain reaction (PCR)-based approaches. There is always an urgent need to detect variation in amino acid sequences directly at the protein level. Mass spectrometry coupled with de novo sequencing has been explored as an alternative and straightforward strategy for detecting amino acid substitutions, as well - this approach is the primary focus of the present study. Influenza virus (A/Puerto Rico/8/1934 H1N1) propagated in embryonated chicken eggs was purified by ultracentrifugation, followed by PNGase F treatment. The deglycosylated virion was lysed and separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The gel band corresponding to neuraminidase was picked up and subjected to liquid chromatography tandem mass spectrometry (LC-MS/MS) analysis. LC-MS/MS analyses, coupled with manual de novo sequencing, allowed the determination of three amino acid substitutions: R346K, S349 N, and S370I/L, in the neuraminidase from the influenza virus (A/Puerto Rico/8/1934 H1N1), which were located in three mutated peptides of the neuraminidase: YGNGVWIGK, TKNHSSR, and PNGWTETDI/LK, respectively. We found that the amino acid substitutions in the proteins of RNA viruses (including influenza A virus) resulting from non-synonymous gene mutations can indeed be directly analyzed via mass spectrometry, and that manual interpretation of the MS/MS data may be beneficial. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

  5. Web Access to Digitised Content of the Exhibition Novo Mesto 1848-1918 at the Dolenjska Museum, Novo Mesto

    Directory of Open Access Journals (Sweden)

    Majda Pungerčar

    2013-09-01

    Full Text Available EXTENDED ABSTRACTFor the first time, the Dolenjska museum Novo mesto provided access to digitised museum resources when they took the decision to enrich the exhibition Novo mesto 1848-1918 by adding digital content. The following goals were identified: the digital content was created at the time of exhibition planning and design, it met the needs of different age groups of visitors, and during the exhibition the content was accessible via touch screen. As such, it also served for educational purposes (content-oriented lectures or problem solving team work. In the course of exhibition digital content was accessible on the museum website http://www.novomesto1848-1918.si. The digital content was divided into the following sections: the web photo gallery, the quiz and the game. The photo gallery was designed in the same way as the exhibition and the print catalogue and extended by the photos of contemporary Novo mesto and accompanied by the music from the orchestron machine. The following themes were outlined: the Austrian Empire, the Krka and Novo mesto, the town and its symbols, images of the town and people, administration and economy, social life and Novo mesto today followed by digitised archive materials and sources from that period such as the Commemorative book of the Uniformed Town Guard, the National Reading Room Guest Book, the Kazina guest book, the album of postcards and the Diploma of Honoured Citizen Josip Gerdešič. The Web application was also a tool for a simple and on line selection of digitised material and the creation of new digital content which proved to be much more convenient for lecturing than Power Point presentations. The quiz consisted of 40 questions relating to the exhibition theme and the catalogue. Each question offered a set of three answers only one of them being correct and illustrated by photography. The application auto selected ten questions and valued the answers immediately. The quiz could be accessed

  6. Effect of a Fusion Peptide by Covalent Conjugation of a Mitochondrial Cell-Penetrating Peptide and a Glutathione Analog Peptide

    Directory of Open Access Journals (Sweden)

    Carmine Pasquale Cerrato

    2017-06-01

    Full Text Available Previously, we designed and synthesized a library of mitochondrial antioxidative cell-penetrating peptides (mtCPPs superior to the parent peptide, SS31, to protect mitochondria from oxidative damage. A library of antioxidative glutathione analogs called glutathione peptides (UPFs, exceptional in hydroxyl radical elimination compared with glutathione, were also designed and synthesized. Here, a follow-up study is described, investigating the effects of the most promising members from both libraries on reactive oxidative species scavenging ability. None of the peptides influenced cell viability at the concentrations used. Fluorescence microscopy studies showed that the fluorescein-mtCPP1-UPF25 (mtgCPP internalized into cells, and spectrofluorometric analysis determined the presence and extent of peptide into different cell compartments. mtgCPP has superior antioxidative activity compared with mtCPP1 and UPF25 against H2O2 insult, preventing ROS formation by 2- and 3-fold, respectively. Moreover, we neither observed effects on mitochondrial membrane potential nor production of ATP. These data indicate that mtgCPP is targeting mitochondria, protecting them from oxidative damage, while also being present in the cytosol. Our hypothesis is based on a synergistic effect resulting from the fused peptide. The mitochondrial peptide segment is targeting mitochondria, whereas the glutathione analog peptide segment is active in the cytosol, resulting in increased scavenging ability.

  7. Peptide-membrane interactions of arginine-tryptophan peptides probed using quartz crystal microbalance with dissipation monitoring.

    KAUST Repository

    Rydberg, Hanna A

    2014-04-18

    Membrane-active peptides include peptides that can cross cellular membranes and deliver macromolecular cargo as well as peptides that inhibit bacterial growth. Some of these peptides can act as both transporters and antibacterial agents. It is desirable to combine the knowledge from these two different fields of membrane-active peptides into design of new peptides with tailored actions, as transporters of cargo or as antibacterial substances, targeting specific membranes. We have previously shown that the position of the amino acid tryptophan in the peptide sequence of three arginine-tryptophan peptides affects their uptake and intracellular localization in live mammalian cells, as well as their ability to inhibit bacterial growth. Here, we use quartz crystal microbalance with dissipation monitoring to assess the induced changes caused by binding of the three peptides to supported model membranes composed of POPC, POPC/POPG, POPC/POPG/cholesterol or POPC/lactosyl PE. Our results indicate that the tryptophan position in the peptide sequence affects the way these peptides interact with the different model membranes and that the presence of cholesterol in particular seems to affect the membrane interaction of the peptide with an even distribution of tryptophans in the peptide sequence. These results give mechanistic insight into the function of these peptides and may aid in the design of membrane-active peptides with specified cellular targets and actions.

  8. Peptide-membrane interactions of arginine-tryptophan peptides probed using quartz crystal microbalance with dissipation monitoring.

    KAUST Repository

    Rydberg, Hanna A; Kunze, Angelika; Carlsson, Nils; Altgä rde, Noomi; Svedhem, Sofia; Nordé n, Bengt

    2014-01-01

    Membrane-active peptides include peptides that can cross cellular membranes and deliver macromolecular cargo as well as peptides that inhibit bacterial growth. Some of these peptides can act as both transporters and antibacterial agents. It is desirable to combine the knowledge from these two different fields of membrane-active peptides into design of new peptides with tailored actions, as transporters of cargo or as antibacterial substances, targeting specific membranes. We have previously shown that the position of the amino acid tryptophan in the peptide sequence of three arginine-tryptophan peptides affects their uptake and intracellular localization in live mammalian cells, as well as their ability to inhibit bacterial growth. Here, we use quartz crystal microbalance with dissipation monitoring to assess the induced changes caused by binding of the three peptides to supported model membranes composed of POPC, POPC/POPG, POPC/POPG/cholesterol or POPC/lactosyl PE. Our results indicate that the tryptophan position in the peptide sequence affects the way these peptides interact with the different model membranes and that the presence of cholesterol in particular seems to affect the membrane interaction of the peptide with an even distribution of tryptophans in the peptide sequence. These results give mechanistic insight into the function of these peptides and may aid in the design of membrane-active peptides with specified cellular targets and actions.

  9. Isolation and identification of a new intracellular antimicrobial peptide produced by Paenibacillus alvei AN5.

    Science.gov (United States)

    Alkotaini, Bassam; Anuar, Nurina; Kadhum, Abdul Amir Hassan; Sani, Asmahani Azira Abdu

    2014-04-01

    A wild-type, Gram-positive, rod-shaped, endospore-forming and motile bacteria has been isolated from palm oil mill sludge in Malaysia. Molecular identification using 16S rRNA gene sequence analysis indicated that the bacteria belonged to genus Paenibacillus. With 97 % similarity to P. alvei (AUG6), the isolate was designated as P. alvei AN5. An antimicrobial compound was extracted from P. alvei AN5-pelleted cells using 95 % methanol and was then lyophilized. Precipitates were re-suspended in phosphate buffered saline (PBS), producing an antimicrobial crude extract (ACE). The ACE showed antimicrobial activity against Salmonella enteritidis ATCC 13076, Escherichia coli ATCC 29522, Bacillus cereus ATCC 14579 and Lactobacillus plantarum ATCC 8014. By using SP-Sepharose cation exchange chromatography, Sephadex G-25 gel filtration and Tricine SDS-PAGE, the ACE was purified, which produced a ~2-kDa active band. SDS-PAGE and infrared (IR) spectroscopy indicated the proteinaceous nature of the antimicrobial compound in the ACE, and liquid chromatography electrospray ionization mass spectroscopy and de novo sequencing using an automatic, Q-TOF premier system detected a peptide with the amino acid sequence F-C-K-S-L-P-L-P-L-S-V-K (1,330.7789 Da). This novel peptide was designated as AN5-2. The antimicrobial peptide exhibited stability from pH 3 to 12 and maintained its activity after being heated to 90 °C. It also remained active after incubation with denaturants (urea, SDS and EDTA).

  10. Sequence protein identification by randomized sequence database and transcriptome mass spectrometry (SPIDER-TMS): from manual to automatic application of a 'de novo sequencing' approach.

    Science.gov (United States)

    Pascale, Raffaella; Grossi, Gerarda; Cruciani, Gabriele; Mecca, Giansalvatore; Santoro, Donatello; Sarli Calace, Renzo; Falabella, Patrizia; Bianco, Giuliana

    Sequence protein identification by a randomized sequence database and transcriptome mass spectrometry software package has been developed at the University of Basilicata in Potenza (Italy) and designed to facilitate the determination of the amino acid sequence of a peptide as well as an unequivocal identification of proteins in a high-throughput manner with enormous advantages of time, economical resource and expertise. The software package is a valid tool for the automation of a de novo sequencing approach, overcoming the main limits and a versatile platform useful in the proteomic field for an unequivocal identification of proteins, starting from tandem mass spectrometry data. The strength of this software is that it is a user-friendly and non-statistical approach, so protein identification can be considered unambiguous.

  11. De novo transcriptome assembly facilitates characterisation of fast-evolving gene families, MHC class I in the bank vole (Myodes glareolus).

    Science.gov (United States)

    Migalska, M; Sebastian, A; Konczal, M; Kotlík, P; Radwan, J

    2017-04-01

    The major histocompatibility complex (MHC) plays a central role in the adaptive immune response and is the most polymorphic gene family in vertebrates. Although high-throughput sequencing has increasingly been used for genotyping families of co-amplifying MHC genes, its potential to facilitate early steps in the characterisation of MHC variation in nonmodel organism has not been fully explored. In this study we evaluated the usefulness of de novo transcriptome assembly in characterisation of MHC sequence diversity. We found that although de novo transcriptome assembly of MHC I genes does not reconstruct sequences of individual alleles, it does allow the identification of conserved regions for PCR primer design. Using the newly designed primers, we characterised MHC I sequences in the bank vole. Phylogenetic analysis of the partial MHC I coding sequence (2-4 exons) of the bank vole revealed a lack of orthology to MHC I of other Cricetidae, consistent with the high gene turnover of this region. The diversity of expressed alleles was characterised using ultra-deep sequencing of the third exon that codes for the peptide-binding region of the MHC molecule. High allelic diversity was demonstrated, with 72 alleles found in 29 individuals. Interindividual variation in the number of expressed loci was found, with the number of alleles per individual ranging from 5 to 14. Strong signatures of positive selection were found for 8 amino acid sites, most of which are inferred to bind antigens in human MHC, indicating conservation of structure despite rapid sequence evolution.

  12. A Two-Piece Derivative of a Group I Intron RNA as a Platform for Designing Self-Assembling RNA Templates to Promote Peptide Ligation

    Directory of Open Access Journals (Sweden)

    Takahiro Tanaka

    2012-01-01

    Full Text Available Multicomponent RNA-peptide complexes are attractive from the viewpoint of artificial design of functional biomacromolecular systems. We have developed self-folding and self-assembling RNAs that serve as templates to assist chemical ligation between two reactive peptides with RNA-binding capabilities. The design principle of previous templates, however, can be applied only to limited classes of RNA-binding peptides. In this study, we employed a two-piece derivative of a group I intron RNA from the Tetrahymena large subunit ribosomal RNA (LSU rRNA as a platform for new template RNAs. In this group I intron-based self-assembling platform, modules for the recognition of substrate peptides can be installed independently from modules holding the platform structure. The new self-assembling platform allows us to expand the repertoire of substrate peptides in template RNA design.

  13. Ligand-regulated peptides: a general approach for modulating protein-peptide interactions with small molecules.

    Science.gov (United States)

    Binkowski, Brock F; Miller, Russell A; Belshaw, Peter J

    2005-07-01

    We engineered a novel ligand-regulated peptide (LiRP) system where the binding activity of intracellular peptides is controlled by a cell-permeable small molecule. In the absence of ligand, peptides expressed as fusions in an FKBP-peptide-FRB-GST LiRP scaffold protein are free to interact with target proteins. In the presence of the ligand rapamycin, or the nonimmunosuppressive rapamycin derivative AP23102, the scaffold protein undergoes a conformational change that prevents the interaction of the peptide with the target protein. The modular design of the scaffold enables the creation of LiRPs through rational design or selection from combinatorial peptide libraries. Using these methods, we identified LiRPs that interact with three independent targets: retinoblastoma protein, c-Src, and the AMP-activated protein kinase. The LiRP system should provide a general method to temporally and spatially regulate protein function in cells and organisms.

  14. Gene design, fusion technology and TEV cleavage conditions influence the purification of oxidized disulphide-rich venom peptides in Escherichia coli.

    Science.gov (United States)

    Sequeira, Ana Filipa; Turchetto, Jeremy; Saez, Natalie J; Peysson, Fanny; Ramond, Laurie; Duhoo, Yoan; Blémont, Marilyne; Fernandes, Vânia O; Gama, Luís T; Ferreira, Luís M A; Guerreiro, Catarina I P I; Gilles, Nicolas; Darbon, Hervé; Fontes, Carlos M G A; Vincentelli, Renaud

    2017-01-17

    Animal venoms are large, complex libraries of bioactive, disulphide-rich peptides. These peptides, and their novel biological activities, are of increasing pharmacological and therapeutic importance. However, recombinant expression of venom peptides in Escherichia coli remains difficult due to the significant number of cysteine residues requiring effective post-translational processing. There is also an urgent need to develop high-throughput recombinant protocols applicable to the production of reticulated peptides to enable efficient screening of their drug potential. Here, a comprehensive study was developed to investigate how synthetic gene design, choice of fusion tag, compartment of expression, tag removal conditions and protease recognition site affect levels of solubility of oxidized venom peptides produced in E. coli. The data revealed that expression of venom peptides imposes significant pressure on cysteine codon selection. DsbC was the best fusion tag for venom peptide expression, in particular when the fusion was directed to the bacterial periplasm. While the redox activity of DsbC was not essential to maximize expression of recombinant fusion proteins, redox activity did lead to higher levels of correctly folded target peptides. With the exception of proline, the canonical TEV protease recognition site tolerated all other residues at its C-terminus, confirming that no non-native residues, which might affect activity, need to be incorporated at the N-terminus of recombinant peptides for tag removal. This study reveals that E. coli is a convenient heterologous host for the expression of soluble and functional venom peptides. Using the optimal construct design, a large and diverse range of animal venom peptides were produced in the µM scale. These results open up new possibilities for the high-throughput production of recombinant disulphide-rich peptides in E. coli.

  15. PH dependent adhesive peptides

    Science.gov (United States)

    Tomich, John; Iwamoto, Takeo; Shen, Xinchun; Sun, Xiuzhi Susan

    2010-06-29

    A novel peptide adhesive motif is described that requires no receptor or cross-links to achieve maximal adhesive strength. Several peptides with different degrees of adhesive strength have been designed and synthesized using solid phase chemistries. All peptides contain a common hydrophobic core sequence flanked by positively or negatively charged amino acids sequences.

  16. Screening and Identification of Peptides Specifically Targeted to Gastric Cancer Cells from a Phage Display Peptide Library

    Science.gov (United States)

    Sahin, Deniz; Taflan, Sevket Onur; Yartas, Gizem; Ashktorab, Hassan; Smoot, Duane T

    2018-04-25

    Background: Gastric cancer is the second most common cancer among the malign cancer types. Inefficiency of traditional techniques both in diagnosis and therapy of the disease makes the development of alternative and novel techniques indispensable. As an alternative to traditional methods, tumor specific targeting small peptides can be used to increase the efficiency of the treatment and reduce the side effects related to traditional techniques. The aim of this study is screening and identification of individual peptides specifically targeted to human gastric cancer cells using a phage-displayed peptide library and designing specific peptide sequences by using experimentally-eluted peptide sequences. Methods: Here, MKN-45 human gastric cancer cells and HFE-145 human normal gastric epithelial cells were used as the target and control cells, respectively. 5 rounds of biopannning with a phage display 12-peptide library were applied following subtraction biopanning with HFE-145 control cells. The selected phage clones were established by enzyme-linked immunosorbent assay and immunofluorescence detection. We first obtain random phage clones after five biopanning rounds, determine the binding levels of each individual clone. Then, we analyze the frequencies of each amino acid in best binding clones to determine positively overexpressed amino acids for designing novel peptide sequences. Results: DE532 (VETSQYFRGTLS) phage clone was screened positive, showing specific binding on MKN-45 gastric cancer cells. DE-Obs (HNDLFPSWYHNY) peptide, which was designed by using amino acid frequencies of experimentally selected peptides in the 5th round of biopanning, showed specific binding in MKN-45 cells. Conclusion: Selection and characterization of individual clones may give us specifically binding peptides, but more importantly, data extracted from eluted phage clones may be used to design theoretical peptides with better binding properties than even experimentally selected ones

  17. The oncolytic peptide LTX-315 induces cell death and DAMP release by mitochondria distortion in human melanoma cells

    Science.gov (United States)

    Eike, Liv-Marie; Yang, Nannan; Rekdal, Øystein; Sveinbjørnsson, Baldur

    2015-01-01

    Host defense peptides (HDPs) are naturally occurring molecules found in most species, in which they play a significant role in the first line defense against intruding pathogens, and several HDPs have been shown to possess anticancer activity. Structure-activity relationship studies on the HDP bovine lactoferricin revealed a de novo design of a nonamer peptide LTX-315, with oncolytic properties. In the present study, we investigated the oncolytic activity of LTX-315 in human melanoma cells (A375). LTX-315 induced a rapid plasma membrane disruption and cell death within 2 hours. At a low concentration, fluorescence-labeled LTX-315 was internalized and accumulated in cytoplasmic vacuoles in close proximity to the mitochondria. The mitochondrial membrane potential was shown to depolarize as a consequence of LTX-315 treatment and at ultrastructural level, the mitochondria morphology was significantly altered. Release of danger signals (DAMPs) such as ATP, Cytochrome C and HMGB1 into the cell supernatant of cultured cells was evident minutes after peptide treatment. The oncolytic effect of LTX-315 involving perturbation of both the cell membrane and the mitochondria with subsequent release of DAMPs may highlight the ability of LTX-315 to induce complete regression and long-term protective immune responses as previously reported in experimental animal models. PMID:26472184

  18. Towards accurate de novo assembly for genomes with repeats

    NARCIS (Netherlands)

    Bucur, Doina

    2017-01-01

    De novo genome assemblers designed for short k-mer length or using short raw reads are unlikely to recover complex features of the underlying genome, such as repeats hundreds of bases long. We implement a stochastic machine-learning method which obtains accurate assemblies with repeats and

  19. Engineering and introduction of de novo disulphide bridges in ...

    Indian Academy of Sciences (India)

    The engineeringof de novo disulphide bridges has been explored as a means to increase the thermal stability of enzymes in the rationalmethod of protein engineering. In this study, Disulphide by Design software, homology modelling and moleculardynamics simulations were used to select appropriate amino acid pairs for ...

  20. De Novo Construction of Redox Active Proteins.

    Science.gov (United States)

    Moser, C C; Sheehan, M M; Ennist, N M; Kodali, G; Bialas, C; Englander, M T; Discher, B M; Dutton, P L

    2016-01-01

    Relatively simple principles can be used to plan and construct de novo proteins that bind redox cofactors and participate in a range of electron-transfer reactions analogous to those seen in natural oxidoreductase proteins. These designed redox proteins are called maquettes. Hydrophobic/hydrophilic binary patterning of heptad repeats of amino acids linked together in a single-chain self-assemble into 4-alpha-helix bundles. These bundles form a robust and adaptable frame for uncovering the default properties of protein embedded cofactors independent of the complexities introduced by generations of natural selection and allow us to better understand what factors can be exploited by man or nature to manipulate the physical chemical properties of these cofactors. Anchoring of redox cofactors such as hemes, light active tetrapyrroles, FeS clusters, and flavins by His and Cys residues allow cofactors to be placed at positions in which electron-tunneling rates between cofactors within or between proteins can be predicted in advance. The modularity of heptad repeat designs facilitates the construction of electron-transfer chains and novel combinations of redox cofactors and new redox cofactor assisted functions. Developing de novo designs that can support cofactor incorporation upon expression in a cell is needed to support a synthetic biology advance that integrates with natural bioenergetic pathways. © 2016 Elsevier Inc. All rights reserved.

  1. Fibrils from designed non-amyloid-related synthetic peptides induce AA-amyloidosis during inflammation in an animal model.

    Directory of Open Access Journals (Sweden)

    Per Westermark

    Full Text Available BACKGROUND: Mouse AA-amyloidosis is a transmissible disease by a prion-like mechanism where amyloid fibrils act by seeding. Synthetic peptides with no amyloid relationship can assemble into amyloid-like fibrils and these may have seeding capacity for amyloid proteins. PRINCIPAL FINDINGS: Several synthetic peptides, designed for nanotechnology, have been examined for their ability to produce fibrils with Congo red affinity and concomitant green birefringence, affinity for thioflavin S and to accelerate AA-amyloidosis in mice. It is shown that some amphiphilic fibril-forming peptides not only produced Congo red birefringence and showed affinity for thioflavin S, but they also shortened the lag phase for systemic AA-amyloidosis in mice when they were given intravenously at the time of inflammatory induction with silver nitride. Peptides, not forming amyloid-like fibrils, did not have such properties. CONCLUSIONS: These observations should caution researchers and those who work with synthetic peptides and their derivatives to be aware of the potential health concerns.

  2. de novo'' aneurysms following endovascular procedures

    International Nuclear Information System (INIS)

    Briganti, F.; Cirillo, S.; Caranci, F.; Esposito, F.; Maiuri, F.

    2002-01-01

    Two personal cases of ''de novo'' aneurysms of the anterior communicating artery (ACoA) occurring 9 and 4 years, respectively, after endovascular carotid occlusion are described. A review of the 30 reported cases (including our own two) of ''de novo'' aneurysms after occlusion of the major cerebral vessels has shown some features, including a rather long time interval after the endovascular procedure of up to 20-25 years (average 9.6 years), a preferential ACoA (36.3%) and internal carotid artery-posterior communicating artery (ICA-PCoA) (33.3%) location of the ''de novo'' aneurysms, and a 10% rate of multiple aneurysms. These data are compared with those of the group of reported spontaneous ''de novo'' aneurysms after SAH or previous aneurysm clipping. We agree that the frequency of ''de novo'' aneurysms after major-vessel occlusion (two among ten procedures in our series, or 20%) is higher than commonly reported (0 to 11%). For this reason, we suggest that patients who have been submitted to endovascular major-vessel occlusion be followed up for up to 20-25 years after the procedure, using non-invasive imaging studies such as MR angiography and high-resolution CT angiography. On the other hand, periodic digital angiography has a questionable risk-benefit ratio; it may be used when a ''de novo'' aneurysm is detected or suspected on non-invasive studies. The progressive enlargement of the ACoA after carotid occlusion, as described in our case 1, must be considered a radiological finding of risk for ''de novo'' aneurysm formation. (orig.)

  3. The effect of peptides and ions interacting with an electrically neutral membrane interface on the structure and stability of lipid membranes in the liquid-crystalline phase and in the liquid-ordered phase

    Science.gov (United States)

    Sano, Ryoko; Masum, Shah Md; Tanaka, Tomoki; Yamashita, Yuko; Levadny, Victor; Yamazaki, Masahito

    2005-08-01

    We investigated the effects of a de novo designed peptide, WLFLLKKK (peptide-1) and La3+, which can bind with the electrically neutral lipid membrane interface, on the stability of the phosphatidylcholine (PC) membrane in the Lα phase and that of the liquid-ordered (lo) phase membranes. The results of spacing of the multilamellar vesicle and shape changes of the giant unilamellar vesicle (GUV) indicate that the peptide-1 can be partitioned into the membrane interface in the Lα phase but not into that in the lo phase. La3+ induced shape changes of GUVs of the lo phase membrane, which are the same as those of GUVs in the Lα phase. This indicates that the binding of La3+ induced an increase in the lateral compression pressure of the membrane, which decreased the surface area of the membrane in the lo phase. The difference of the membrane interface between the Lα phase and the lo phase is discussed.

  4. The effect of peptides and ions interacting with an electrically neutral membrane interface on the structure and stability of lipid membranes in the liquid-crystalline phase and in the liquid-ordered phase

    International Nuclear Information System (INIS)

    Sano, Ryoko; Masum, Shah Md; Tanaka, Tomoki; Yamashita, Yuko; Levadny, Victor; Yamazaki, Masahito

    2005-01-01

    We investigated the effects of a de novo designed peptide, WLFLLKKK (peptide-1) and La 3+ , which can bind with the electrically neutral lipid membrane interface, on the stability of the phosphatidylcholine (PC) membrane in the L α phase and that of the liquid-ordered (lo) phase membranes. The results of spacing of the multilamellar vesicle and shape changes of the giant unilamellar vesicle (GUV) indicate that the peptide-1 can be partitioned into the membrane interface in the L α phase but not into that in the lo phase. La 3+ induced shape changes of GUVs of the lo phase membrane, which are the same as those of GUVs in the L α phase. This indicates that the binding of La 3+ induced an increase in the lateral compression pressure of the membrane, which decreased the surface area of the membrane in the lo phase. The difference of the membrane interface between the L α phase and the lo phase is discussed

  5. Design, synthesis and DNA interactions of a chimera between a platinum complex and an IHF mimicking peptide.

    Science.gov (United States)

    Rao, Harita; Damian, Mariana S; Alshiekh, Alak; Elmroth, Sofi K C; Diederichsen, Ulf

    2015-12-28

    Conjugation of metal complexes with peptide scaffolds possessing high DNA binding affinity has shown to modulate their biological activities and to enhance their interaction with DNA. In this work, a platinum complex/peptide chimera was synthesized based on a model of the Integration Host Factor (IHF), an architectural protein possessing sequence specific DNA binding and bending abilities through its interaction with a minor groove. The model peptide consists of a cyclic unit resembling the minor grove binding subdomain of IHF, a positively charged lysine dendrimer for electrostatic interactions with the DNA phosphate backbone and a flexible glycine linker tethering the two units. A norvaline derived artificial amino acid was designed to contain a dimethylethylenediamine as a bidentate platinum chelating unit, and introduced into the IHF mimicking peptides. The interaction of the chimeric peptides with various DNA sequences was studied by utilizing the following experiments: thermal melting studies, agarose gel electrophoresis for plasmid DNA unwinding experiments, and native and denaturing gel electrophoresis to visualize non-covalent and covalent peptide-DNA adducts, respectively. By incorporation of the platinum metal center within the model peptide mimicking IHF we have attempted to improve its specificity and DNA targeting ability, particularly towards those sequences containing adjacent guanine residues.

  6. Bioavailability and transport of peptides and peptide drugs into the brain.

    Science.gov (United States)

    Egleton, R D; Davis, T P

    1997-01-01

    Rational drug design and the targeting of specific organs has become a reality in modern drug development, with the emergence of molecular biology and receptor chemistry as powerful tools for the pharmacologist. A greater understanding of peptide function as one of the major extracellular message systems has made neuropeptides an important target in neuropharmaceutical drug design. The major obstacle to targeting the brain with therapeutics is the presence of the blood-brain barrier (BBB), which controls the concentration and entry of solutes into the central nervous system. Peptides are generally polar in nature, do not easily cross the blood-brain barrier by diffusion, and except for a small number do not have specific transport systems. Peptides can also undergo metabolic deactivation by peptidases of the blood, brain and the endothelial cells that comprise the BBB. In this review, we discuss a number of the recent strategies which have been used to promote peptide stability and peptide entry into the brain. In addition, we approach the subject of targeting specific transport systems that can be found on the brain endothelial cells, and describe the limitations of the methodologies that are currently used to study brain entry of neuropharmaceuticals.

  7. Brute-Force Approach for Mass Spectrometry-Based Variant Peptide Identification in Proteogenomics without Personalized Genomic Data

    Science.gov (United States)

    Ivanov, Mark V.; Lobas, Anna A.; Levitsky, Lev I.; Moshkovskii, Sergei A.; Gorshkov, Mikhail V.

    2018-02-01

    In a proteogenomic approach based on tandem mass spectrometry analysis of proteolytic peptide mixtures, customized exome or RNA-seq databases are employed for identifying protein sequence variants. However, the problem of variant peptide identification without personalized genomic data is important for a variety of applications. Following the recent proposal by Chick et al. (Nat. Biotechnol. 33, 743-749, 2015) on the feasibility of such variant peptide search, we evaluated two available approaches based on the previously suggested "open" search and the "brute-force" strategy. To improve the efficiency of these approaches, we propose an algorithm for exclusion of false variant identifications from the search results involving analysis of modifications mimicking single amino acid substitutions. Also, we propose a de novo based scoring scheme for assessment of identified point mutations. In the scheme, the search engine analyzes y-type fragment ions in MS/MS spectra to confirm the location of the mutation in the variant peptide sequence.

  8. A Macrocyclic Peptide that Serves as a Cocrystallization Ligand and Inhibits the Function of a MATE Family Transporter

    Directory of Open Access Journals (Sweden)

    Hiroaki Suga

    2013-08-01

    Full Text Available The random non-standard peptide integrated discovery (RaPID system has proven to be a powerful approach to discover de novo natural product-like macrocyclic peptides that inhibit protein functions. We have recently reported three macrocyclic peptides that bind to Pyrococcus furiosus multidrug and toxic compound extrusion (PfMATE transporter and inhibit the transport function. Moreover, these macrocyclic peptides were successfully employed as cocrystallization ligands of selenomethionine-labeled PfMATE. In this report, we disclose the details of the RaPID selection strategy that led to the identification of these three macrocyclic peptides as well as a fourth macrocyclic peptide, MaD8, which is exclusively discussed in this article. MaD8 was found to bind within the cleft of PfMATE’s extracellular side and blocked the path of organic small molecules being extruded. The results of an ethidium bromide efflux assay confirmed the efflux inhibitory activity of MaD8, whose behavior was similar to that of previously reported MaD5.

  9. Design and bioinformatics analysis of novel biomimetic peptides as nanocarriers for gene transfer

    Directory of Open Access Journals (Sweden)

    Asia Majidi

    2015-01-01

    Full Text Available Objective(s: The introduction of nucleic acids into cells for therapeutic objectives is significantly hindered by the size and charge of these molecules and therefore requires efficient vectors that assist cellular uptake. For several years great efforts have been devoted to the study of development of recombinant vectors based on biological domains with potential applications in gene therapy. Such vectors have been synthesized in genetically engineered approach, resulting in biomacromolecules with new properties that are not present in nature. Materials and Methods: In this study, we have designed new peptides using homology modeling with the purpose of overcoming the cell barriers for successful gene delivery through Bioinformatics tools. Three different carriers were designed and one of those with better score through Bioinformatics tools was cloned, expressed and its affinity for pDNA was monitored. Results: The resultszz demonstrated that the vector can effectively condense pDNAinto nanoparticles with the average sizes about 100 nm. Conclusion: We hope these peptides can overcome the biological barriers associated with gene transfer, and mediate efficient gene delivery.

  10. A workflow for peptide-based proteomics in a poorly sequenced plant: A case study on the plasma membrane proteome of banana

    DEFF Research Database (Denmark)

    Vertommen, A.; Laurell Blom Møller, Anders; Cordewener, J. H. G.

    2011-01-01

    for membrane proteomics. However, their application in non-model plants demands special precautions to prevent false positive identification of proteins.In the current paper, a workflow for membrane proteomics in banana, a poorly sequenced plant, is proposed. The main steps of this workflow are (i......) optimization of the peptide separation, (ii) performing de novo sequencing to allow a sequence homology search and (iii) visualization of identified peptide–protein associations using Cytoscape to remove redundancy and wrongly assigned peptides, based on species-specific information. By applying this workflow...

  11. Structure-activity-based design of a synthetic malaria peptide eliciting sporozoite inhibitory antibodies in a virosomal formulation.

    NARCIS (Netherlands)

    Okitsu, S.L.; Kienzl, U.; Moehle, K.; Silvie, O.; Peduzzi, E.; Mueller, M.S.; Sauerwein, R.W.; Matile, H.; Zurbriggen, R.; Mazier, D.; Robinson, J.A.; Pluschke, G.

    2007-01-01

    The circumsporozoite protein (CSP) of Plasmodium falciparum is a leading candidate antigen for inclusion in a malaria subunit vaccine. We describe here the design of a conformationally constrained synthetic peptide, designated UK-39, which has structural and antigenic similarity to the NPNA-repeat

  12. Peptide Vaccines for Leishmaniasis

    Directory of Open Access Journals (Sweden)

    Rory C. F. De Brito

    2018-05-01

    Full Text Available Due to an increase in the incidence of leishmaniases worldwide, the development of new strategies such as prophylactic vaccines to prevent infection and decrease the disease have become a high priority. Classic vaccines against leishmaniases were based on live or attenuated parasites or their subunits. Nevertheless, the use of whole parasite or their subunits for vaccine production has numerous disadvantages. Therefore, the use of Leishmania peptides to design more specific vaccines against leishmaniases seems promising. Moreover, peptides have several benefits in comparison with other kinds of antigens, for instance, good stability, absence of potentially damaging materials, antigen low complexity, and low-cost to scale up. By contrast, peptides are poor immunogenic alone, and they need to be delivered correctly. In this context, several approaches described in this review are useful to solve these drawbacks. Approaches, such as, peptides in combination with potent adjuvants, cellular vaccinations, adenovirus, polyepitopes, or DNA vaccines have been used to develop peptide-based vaccines. Recent advancements in peptide vaccine design, chimeric, or polypeptide vaccines and nanovaccines based on particles attached or formulated with antigenic components or peptides have been increasingly employed to drive a specific immune response. In this review, we briefly summarize the old, current, and future stands on peptide-based vaccines, describing the disadvantages and benefits associated with them. We also propose possible approaches to overcome the related weaknesses of synthetic vaccines and suggest future guidelines for their development.

  13. Peptide Vaccines for Leishmaniasis.

    Science.gov (United States)

    De Brito, Rory C F; Cardoso, Jamille M De O; Reis, Levi E S; Vieira, Joao F; Mathias, Fernando A S; Roatt, Bruno M; Aguiar-Soares, Rodrigo Dian D O; Ruiz, Jeronimo C; Resende, Daniela de M; Reis, Alexandre B

    2018-01-01

    Due to an increase in the incidence of leishmaniases worldwide, the development of new strategies such as prophylactic vaccines to prevent infection and decrease the disease have become a high priority. Classic vaccines against leishmaniases were based on live or attenuated parasites or their subunits. Nevertheless, the use of whole parasite or their subunits for vaccine production has numerous disadvantages. Therefore, the use of Leishmania peptides to design more specific vaccines against leishmaniases seems promising. Moreover, peptides have several benefits in comparison with other kinds of antigens, for instance, good stability, absence of potentially damaging materials, antigen low complexity, and low-cost to scale up. By contrast, peptides are poor immunogenic alone, and they need to be delivered correctly. In this context, several approaches described in this review are useful to solve these drawbacks. Approaches, such as, peptides in combination with potent adjuvants, cellular vaccinations, adenovirus, polyepitopes, or DNA vaccines have been used to develop peptide-based vaccines. Recent advancements in peptide vaccine design, chimeric, or polypeptide vaccines and nanovaccines based on particles attached or formulated with antigenic components or peptides have been increasingly employed to drive a specific immune response. In this review, we briefly summarize the old, current, and future stands on peptide-based vaccines, describing the disadvantages and benefits associated with them. We also propose possible approaches to overcome the related weaknesses of synthetic vaccines and suggest future guidelines for their development.

  14. One-pot synthesis of water soluble iron nanoparticles using rationally-designed peptides and ligand release.

    Science.gov (United States)

    Papst, Stefanie; Cheong, Soshan; Banholzer, Moritz J; Brimble, Margaret A; Williams, David E; Tilley, Richard D

    2013-05-18

    Herein we report the rational design of new phosphopeptides for control of nucleation, growth and aggregation of water-soluble, superparamagnetic iron-iron oxide core-shell nanoparticles. The use of the designed peptides enables a one-pot synthesis that avoids utilizing unstable or toxic iron precursors, organic solvents, and the need for exchange of capping agent after synthesis of the NPs.

  15. Rosetta FlexPepDock ab-initio: simultaneous folding, docking and refinement of peptides onto their receptors.

    Science.gov (United States)

    Raveh, Barak; London, Nir; Zimmerman, Lior; Schueler-Furman, Ora

    2011-04-29

    Flexible peptides that fold upon binding to another protein molecule mediate a large number of regulatory interactions in the living cell and may provide highly specific recognition modules. We present Rosetta FlexPepDock ab-initio, a protocol for simultaneous docking and de-novo folding of peptides, starting from an approximate specification of the peptide binding site. Using the Rosetta fragments library and a coarse-grained structural representation of the peptide and the receptor, FlexPepDock ab-initio samples efficiently and simultaneously the space of possible peptide backbone conformations and rigid-body orientations over the receptor surface of a given binding site. The subsequent all-atom refinement of the coarse-grained models includes full side-chain modeling of both the receptor and the peptide, resulting in high-resolution models in which key side-chain interactions are recapitulated. The protocol was applied to a benchmark in which peptides were modeled over receptors in either their bound backbone conformations or in their free, unbound form. Near-native peptide conformations were identified in 18/26 of the bound cases and 7/14 of the unbound cases. The protocol performs well on peptides from various classes of secondary structures, including coiled peptides with unusual turns and kinks. The results presented here significantly extend the scope of state-of-the-art methods for high-resolution peptide modeling, which can now be applied to a wide variety of peptide-protein interactions where no prior information about the peptide backbone conformation is available, enabling detailed structure-based studies and manipulation of those interactions. © 2011 Raveh et al.

  16. De novo design, synthesis, and in vitro activity of LFA-1 antagonists based on a bicyclic[5.5]hydantoin scaffold.

    Science.gov (United States)

    Potin, Dominique; Launay, Michele; Nicolai, Eric; Fabreguette, Maud; Malabre, Patrice; Caussade, François; Besse, Dominique; Skala, Stacey; Stetsko, Dawn K; Todderud, Gordon; Beno, Brett R; Cheney, Daniel L; Chang, Chiehying J; Sheriff, Steven; Hollenbaugh, Diane L; Barrish, Joel C; Iwanowicz, Edwin J; Suchard, Suzanne J; Dhar, T G Murali

    2005-02-15

    LFA-1 (leukocyte function-associated antigen-1), is a member of the beta(2)-integrin family and is expressed on all leukocytes. The LFA-1/ICAM interaction promotes tight adhesion between activated leukocytes and the endothelium, as well as between T cells and antigen-presenting cells. Evidence from both animal models and clinical trials provides support for LFA-1 as a target in several different inflammatory diseases. This paper describes the de novo design, synthesis and in vitro activity of LFA-1 antagonists based on a bicyclic[5.5]hydantoin scaffold.

  17. Study on Strategic Planning of Road and Bridge Infrastructure Development in City Planning: Taking Porto-novo City of Benin Republic as Example

    Directory of Open Access Journals (Sweden)

    Boko-haya Dossa Didier

    2018-01-01

    Full Text Available Concern about the townlet infrastructure construction in developing country is one of the crucial part of county town planning and development. By taking the overall planning and design in a case study of Porto-novo city at Republic of Benin, this paper analyzes the characteristics and opportunities of Porto-novo city and puts forward corresponding infrastructure construction strategy. In the end, the paper comes up with specific plan of planning and design under the background of Porto-novo's planning of development strategy.

  18. Designer lipid-like peptides: a class of detergents for studying functional olfactory receptors using commercial cell-free systems.

    Science.gov (United States)

    Corin, Karolina; Baaske, Philipp; Ravel, Deepali B; Song, Junyao; Brown, Emily; Wang, Xiaoqiang; Wienken, Christoph J; Jerabek-Willemsen, Moran; Duhr, Stefan; Luo, Yuan; Braun, Dieter; Zhang, Shuguang

    2011-01-01

    A crucial bottleneck in membrane protein studies, particularly G-protein coupled receptors, is the notorious difficulty of finding an optimal detergent that can solubilize them and maintain their stability and function. Here we report rapid production of 12 unique mammalian olfactory receptors using short designer lipid-like peptides as detergents. The peptides were able to solubilize and stabilize each receptor. Circular dichroism showed that the purified olfactory receptors had alpha-helical secondary structures. Microscale thermophoresis suggested that the receptors were functional and bound their odorants. Blot intensity measurements indicated that milligram quantities of each olfactory receptor could be produced with at least one peptide detergent. The peptide detergents' capability was comparable to that of the detergent Brij-35. The ability of 10 peptide detergents to functionally solubilize 12 olfactory receptors demonstrates their usefulness as a new class of detergents for olfactory receptors, and possibly other G-protein coupled receptors and membrane proteins.

  19. Designer lipid-like peptides: a class of detergents for studying functional olfactory receptors using commercial cell-free systems.

    Directory of Open Access Journals (Sweden)

    Karolina Corin

    Full Text Available A crucial bottleneck in membrane protein studies, particularly G-protein coupled receptors, is the notorious difficulty of finding an optimal detergent that can solubilize them and maintain their stability and function. Here we report rapid production of 12 unique mammalian olfactory receptors using short designer lipid-like peptides as detergents. The peptides were able to solubilize and stabilize each receptor. Circular dichroism showed that the purified olfactory receptors had alpha-helical secondary structures. Microscale thermophoresis suggested that the receptors were functional and bound their odorants. Blot intensity measurements indicated that milligram quantities of each olfactory receptor could be produced with at least one peptide detergent. The peptide detergents' capability was comparable to that of the detergent Brij-35. The ability of 10 peptide detergents to functionally solubilize 12 olfactory receptors demonstrates their usefulness as a new class of detergents for olfactory receptors, and possibly other G-protein coupled receptors and membrane proteins.

  20. Rationally Designed Peptides and Peptidomimetics as Inhibitors of Amyloid-β (Aβ) Aggregation: Potential Therapeutics of Alzheimer's Disease.

    Science.gov (United States)

    Goyal, Deepti; Shuaib, Suniba; Mann, Sukhmani; Goyal, Bhupesh

    2017-02-13

    Alzheimer's disease (AD) is a progressive neurodegenerative disease with no clinically accepted treatment to cure or halt its progression. The worldwide effort to develop peptide-based inhibitors of amyloid-β (Aβ) aggregation can be considered an unplanned combinatorial experiment. An understanding of what has been done and achieved may advance our understanding of AD pathology and the discovery of effective therapeutic agents. We review here the history of such peptide-based inhibitors, including those based on the Aβ sequence and those not derived from that sequence, containing both natural and unnatural amino acid building blocks. Peptide-based aggregation inhibitors hold significant promise for future AD therapy owing to their high selectivity, effectiveness, low toxicity, good tolerance, low accumulation in tissues, high chemical and biological diversity, possibility of rational design, and highly developed methods for analyzing their mode of action, proteolytic stability (modified peptides), and blood-brain barrier (BBB) permeability.

  1. ESI-MS/MS Identification of a Bradykinin-Potentiating Peptide from Amazon Bothrops atrox Snake Venom Using a Hybrid Qq-oaTOF Mass Spectrometer

    Science.gov (United States)

    Coutinho-Neto, Antonio; Caldeira, Cleópatra A. S.; Souza, Gustavo H. M. F.; Zaqueo, Kayena D.; Kayano, Anderson M.; Silva, Rodrigo S.; Zuliani, Juliana P.; Soares, Andreimar M.; Stábeli, Rodrigo G.; Calderon, Leonardo A.

    2013-01-01

    A bradykinin-potentiating peptide (BPP) from Amazon Bothrops atrox venom with m/z 1384.7386 was identified and characterized by collision induced dissociation (CID) using an ESI-MS/MS spectra obtained in positive ion mode on a hybrid Qq-oaTOF mass spectrometer, Xevo G2 QTof MS (Waters, Manchester, UK). De novo peptide sequence analysis of the CID fragmentation spectra showed the amino acid sequence ZKWPRPGPEIPP, with a pyroglutamic acid and theoretical monoisotopic m/z 1384.7378, which is similar to experimental data, showing a mass accuracy of 0.6 ppm. The peptide is homologous to other BPP from Bothrops moojeni and was named as BPP-BAX12. PMID:23430539

  2. The rational design of a peptide-based hydrogel responsive to H2S.

    Science.gov (United States)

    Peltier, Raoul; Chen, Ganchao; Lei, Haipeng; Zhang, Mei; Gao, Liqian; Lee, Su Seong; Wang, Zuankai; Sun, Hongyan

    2015-12-18

    The development of hydrogels that are responsive to external stimuli in a well-controlled manner is important for numerous biomedical applications. Herein we reported the first example of a hydrogel responsive to hydrogen sulphide (H2S). H2S is an important gasotransmitter whose deregulation has been associated with a number of pathological conditions. Our hydrogel design is based on the functionalization of an ultrashort hydrogelating peptide sequence with an azidobenzyl moiety, which was reported to react with H2S selectively under physiological conditions. The resulting peptide was able to produce hydrogels at a concentration as low as 0.1 wt%. It could then be fully degraded in the presence of excess H2S. We envision that the novel hydrogel developed in this study may provide useful tools for biomedical research.

  3. Effect of Enamel Matrix Derivative and of Proline-Rich Synthetic Peptides on the Differentiation of Human Mesenchymal Stem Cells Toward the Osteogenic Lineage

    Czech Academy of Sciences Publication Activity Database

    Ramis, J. M.; Rubert, M.; Vondrášek, Jiří; Gaya, A.; Lyngstadaas, S. P.; Monjo, M.

    2012-01-01

    Roč. 18, 11/12 (2012), s. 1253-1263 ISSN 1937-3341 Grant - others:GA ČR(CZ) GAP302/10/0427 Institutional research plan: CEZ:AV0Z40550506 Keywords : bone-marrow-cells * de-novo peptide * in-vitro * structure prediction Subject RIV: EI - Biotechnology ; Bionics

  4. De novo malignancy after pancreas transplantation in Japan.

    Science.gov (United States)

    Tomimaru, Y; Ito, T; Marubashi, S; Kawamoto, K; Tomokuni, A; Asaoka, T; Wada, H; Eguchi, H; Mori, M; Doki, Y; Nagano, H

    2015-04-01

    Long-term immunosuppression is associated with an increased risk of cancer. Especially, the immunosuppression in pancreas transplantation is more intensive than that in other organ transplantation because of its strong immunogenicity. Therefore, it suggests that the risk of post-transplant de novo malignancy might increase in pancreas transplantation. However, there have been few studies of de novo malignancy after pancreas transplantation. The aim of this study was to analyze the incidence of de novo malignancy after pancreas transplantation in Japan. Post-transplant patients with de novo malignancy were surveyed and characterized in Japan. Among 107 cases receiving pancreas transplantation in Japan between 2001 and 2010, de novo malignancy developed in 9 cases (8.4%): post-transplant lymphoproliferative disorders in 6 cases, colon cancer in 1 case, renal cancer in 1 case, and brain tumor in 1 case. We clarified the incidence of de novo malignancy after pancreas transplantation in Japan. Copyright © 2015 Elsevier Inc. All rights reserved.

  5. The influence of pathological mutations and proline substitutions in TDP-43 glycine-rich peptides on its amyloid properties and cellular toxicity.

    Directory of Open Access Journals (Sweden)

    Chia-Sui Sun

    Full Text Available TAR DNA-binding protein (TDP-43 was identified as the major ubiquitinated component deposited in the inclusion bodies in amyotrophic lateral sclerosis (ALS and frontotemporal lobar degeneration with ubiquitin-positive inclusions (FTLD-U in 2006. Later on, numerous ALS-related mutations were found in either the glycine or glutamine/asparagine-rich region on the TDP-43 C-terminus, which hinted on the importance of mutations on the disease pathogenesis. However, how the structural conversion was influenced by the mutations and the biological significance of these peptides remains unclear. In this work, various peptides bearing pathogenic or de novo designed mutations were synthesized and displayed their ability to form twisted amyloid fibers, cause liposome leakage, and mediate cellular toxicity as confirmed by transmission electron microscopy (TEM, circular dichroism (CD, Thioflavin T (ThT assay, Raman spectroscopy, calcein leakage assay, and cell viability assay. We have also shown that replacing glycines with prolines, known to obstruct β-sheet formation, at the different positions in these peptides may influence the amyloidogenesis process and neurotoxicity. In these cases, GGG308PPP mutant was not able to form beta-amyloid, cause liposome leakage, nor jeopardized cell survival, which hinted on the importance of the glycines (308-310 during amyloidogenesis.

  6. Rational Design of Alpha-Helical Antimicrobial Peptides: Do's and Don'ts

    DEFF Research Database (Denmark)

    Uggerhøj, Lars Erik; Poulsen, Tanja Juul; Munk, Jens Kristian

    2015-01-01

    Antimicrobial peptides (AMPs) are promising candidates for battling multiresistant bacteria. Despite extensive research, structure–activity relationships of AMPs are not fully understood, and there is a lack of structural data relating to AMPs in lipids. Here we present the NMR structure of anoplin...... (GLLKRIKTLL-NH2) in a micellar environment. A vast library of substitutions was designed and tested for antimicrobial and hemolytic activity, as well as for changes in structure and lipid interactions. This showed that improvement of antimicrobial activity without concomitant introduction of strong hemolytic...

  7. A novel 13 residue acyclic peptide from the marine snail, Conus monile, targets potassium channels.

    Science.gov (United States)

    Sudarslal, Sadasivannair; Singaravadivelan, Govindaswamy; Ramasamy, Palanisamy; Ananda, Kuppanna; Sarma, Siddhartha P; Sikdar, Sujit K; Krishnan, K S; Balaram, Padmanabhan

    2004-05-07

    A novel 13-residue peptide Mo1659 has been isolated from the venom of a vermivorous cone snail, Conus monile. HPLC fractions of the venom extract yielded an intense UV absorbing fraction with a mass of 1659Da. De novo sequencing using both matrix assisted laser desorption and ionization and electrospray MS/MS methods together with analysis of proteolytic fragments successfully yielded the amino acid sequence, FHGGSWYRFPWGY-NH(2). This was further confirmed by comparison with the chemically synthesized peptide and by conventional Edman sequencing. Mo1659 has an unusual sequence with a preponderance of aromatic residues and the absence of apolar, aliphatic residues like Ala, Val, Leu, and Ile. Mo1659 has no disulfide bridges distinguishing it from the conotoxins and bears no sequence similarity with any of the acyclic peptides isolated thus far from the venom of cone snails. Electrophysiological studies on the effect of Mo1659 on measured currents in dorsal root ganglion neurons suggest that the peptide targets non-inactivating voltage-dependent potassium channels.

  8. Computational design of a PDZ domain peptide inhibitor that rescues CFTR activity.

    Directory of Open Access Journals (Sweden)

    Kyle E Roberts

    Full Text Available The cystic fibrosis transmembrane conductance regulator (CFTR is an epithelial chloride channel mutated in patients with cystic fibrosis (CF. The most prevalent CFTR mutation, ΔF508, blocks folding in the endoplasmic reticulum. Recent work has shown that some ΔF508-CFTR channel activity can be recovered by pharmaceutical modulators ("potentiators" and "correctors", but ΔF508-CFTR can still be rapidly degraded via a lysosomal pathway involving the CFTR-associated ligand (CAL, which binds CFTR via a PDZ interaction domain. We present a study that goes from theory, to new structure-based computational design algorithms, to computational predictions, to biochemical testing and ultimately to epithelial-cell validation of novel, effective CAL PDZ inhibitors (called "stabilizers" that rescue ΔF508-CFTR activity. To design the "stabilizers", we extended our structural ensemble-based computational protein redesign algorithm K* to encompass protein-protein and protein-peptide interactions. The computational predictions achieved high accuracy: all of the top-predicted peptide inhibitors bound well to CAL. Furthermore, when compared to state-of-the-art CAL inhibitors, our design methodology achieved higher affinity and increased binding efficiency. The designed inhibitor with the highest affinity for CAL (kCAL01 binds six-fold more tightly than the previous best hexamer (iCAL35, and 170-fold more tightly than the CFTR C-terminus. We show that kCAL01 has physiological activity and can rescue chloride efflux in CF patient-derived airway epithelial cells. Since stabilizers address a different cellular CF defect from potentiators and correctors, our inhibitors provide an additional therapeutic pathway that can be used in conjunction with current methods.

  9. Peptides in melanoma therapy.

    Science.gov (United States)

    Mocellin, Simone

    2012-01-01

    Peptides derived from tumor associated antigens can be utilized to elicit a therapeutically effective immune response against melanoma in experimental models. However, patient vaccination with peptides - although it is often followed by the induction of melanoma- specific T lymphocytes - is rarely associated with tumor response of clinical relevance. In this review I summarize the principles of peptide design as well as the results so far obtained in the clinical setting while treating cutaneous melanoma by means of this active immunotherapy strategy. I also discuss some immunological and methodological issues that might be helpful for the successful development of peptide-based vaccines.

  10. De novo design and synthesis of ultra-short peptidomimetic antibiotics having dual antimicrobial and anti-inflammatory activities.

    Science.gov (United States)

    Murugan, Ravichandran N; Jacob, Binu; Ahn, Mija; Hwang, Eunha; Sohn, Hoik; Park, Hyo-Nam; Lee, Eunjung; Seo, Ji-Hyung; Cheong, Chaejoon; Nam, Ky-Youb; Hyun, Jae-Kyung; Jeong, Ki-Woong; Kim, Yangmee; Shin, Song Yub; Bang, Jeong Kyu

    2013-01-01

    Much attention has been focused on the design and synthesis of potent, cationic antimicrobial peptides (AMPs) that possess both antimicrobial and anti-inflammatory activities. However, their development into therapeutic agents has been limited mainly due to their large size (12 to 50 residues in length) and poor protease stability. In an attempt to overcome the issues described above, a set of ultra-short, His-derived antimicrobial peptides (HDAMPs) has been developed for the first time. Through systematic tuning of pendant hydrophobic alkyl tails at the N(π)- and N(τ)-positions on His, and the positive charge of Arg, much higher prokaryotic selectivity was achieved, compared to human AMP LL-37. Additionally, the most potent HDAMPs showed promising dual antimicrobial and anti-inflammatory activities, as well as anti-methicillin-resistant Staphylococcus aureus (MRSA) activity and proteolytic resistance. Our results from transmission electron microscopy, membrane depolarization, confocal laser-scanning microscopy, and calcein-dye leakage experiments propose that HDAMP-1 kills microbial cells via dissipation of the membrane potential by forming pore/ion channels on bacterial cell membranes. The combination of the ultra-short size, high-prokaryotic selectivity, potent anti-MRSA activity, anti-inflammatory activity, and proteolytic resistance of the designed HDAMP-1, -3, -5, and -6 makes these molecules promising candidates for future antimicrobial therapeutics.

  11. Epitope-based peptide vaccine design and target site depiction against Ebola viruses: an immunoinformatics study.

    Science.gov (United States)

    Khan, M A; Hossain, M U; Rakib-Uz-Zaman, S M; Morshed, M N

    2015-07-01

    Ebola viruses (EBOVs) have been identified as an emerging threat in recent year as it causes severe haemorrhagic fever in human. Epitope-based vaccine design for EBOVs remains a top priority because a mere progress has been made in this regard. Another reason is the lack of antiviral drug and licensed vaccine although there is a severe outbreak in Central Africa. In this study, we aimed to design an epitope-based vaccine that can trigger a significant immune response as well as to prognosticate inhibitor that can bind with potential drug target sites using various immunoinformatics and docking simulation tools. The capacity to induce both humoral and cell-mediated immunity by T cell and B cell was checked for the selected protein. The peptide region spanning 9 amino acids from 42 to 50 and the sequence TLASIGTAF were found as the most potential B and T cell epitopes, respectively. This peptide could interact with 12 HLAs and showed high population coverage up to 80.99%. Using molecular docking, the epitope was further appraised for binding against HLA molecules to verify the binding cleft interaction. In addition with this, the allergenicity of the epitopes was also evaluated. In the post-therapeutic strategy, docking study of predicted 3D structure identified suitable therapeutic inhibitor against targeted protein. However, this computational epitope-based peptide vaccine designing and target site prediction against EBOVs open up a new horizon which may be the prospective way in Ebola viruses research; the results require validation by in vitro and in vivo experiments. © 2015 John Wiley & Sons Ltd.

  12. Design and evaluation of antimalarial peptides derived from prediction of short linear motifs in proteins related to erythrocyte invasion.

    Directory of Open Access Journals (Sweden)

    Alessandra Bianchin

    Full Text Available The purpose of this study was to investigate the blood stage of the malaria causing parasite, Plasmodium falciparum, to predict potential protein interactions between the parasite merozoite and the host erythrocyte and design peptides that could interrupt these predicted interactions. We screened the P. falciparum and human proteomes for computationally predicted short linear motifs (SLiMs in cytoplasmic portions of transmembrane proteins that could play roles in the invasion of the erythrocyte by the merozoite, an essential step in malarial pathogenesis. We tested thirteen peptides predicted to contain SLiMs, twelve of them palmitoylated to enhance membrane targeting, and found three that blocked parasite growth in culture by inhibiting the initiation of new infections in erythrocytes. Scrambled peptides for two of the most promising peptides suggested that their activity may be reflective of amino acid properties, in particular, positive charge. However, one peptide showed effects which were stronger than those of scrambled peptides. This was derived from human red blood cell glycophorin-B. We concluded that proteome-wide computational screening of the intracellular regions of both host and pathogen adhesion proteins provides potential lead peptides for the development of anti-malarial compounds.

  13. Design and mechanism of action of a novel bacteria-selective antimicrobial peptide from the cell-penetrating peptide Pep-1

    International Nuclear Information System (INIS)

    Zhu, W.L.; Lan Hongliang; Park, Il-Seon; Kim, Jae Il; Jin, H.Z.; Hahm, Kyung-Soo; Shin, S.Y.

    2006-01-01

    Here, we report the successful design of a novel bacteria-selective antimicrobial peptide, Pep-1-K (KKTWWKTWWTKWSQPKKKRKV). Pep-1-K was designed by replacing Glu-2, Glu-6, and Glu-11 in the cell-penetrating peptide Pep-1 with Lys. Pep-1-K showed strong antibacterial activity against reference strains (MIC = 1-2 μM) of Gram-positive and Gram-negative bacteria as well as against clinical isolates (MIC = 1-8 μM) of methicillin-resistant Staphylococcus aureus and multidrug-resistant Pseudomonas aeruginosa. In contrast, Pep-1-K did not cause hemolysis of human erythrocytes even at 200 μM. These results indicate that Pep-1-K may be a good candidate for antimicrobial drug development, especially as a topical agent against antibiotic-resistant microorganisms. Tryptophan fluorescence studies indicated that the lack of hemolytic activity of Pep-1-K correlated with its weak ability to penetrate zwitterionic phosphatidylcholine/cholesterol (10:1, w/w) vesicles, which mimic eukaryotic membranes. Furthermore, Pep-1-K caused little or no dye leakage from negatively charged phosphatidylethanolamine/phosphatidylglycerol (7:3, w/w) vesicles, which mimic bacterial membranes but had a potent ability to cause depolarization of the cytoplasmic membrane potential of intact S. aureus cells. These results suggested that Pep-1-K kills microorganisms by not the membrane-disrupting mode but the formation of small channels that permit transit of ions or protons but not molecules as large as calcein

  14. The design and fabrication of supramolecular semiconductor nanowires formed by benzothienobenzothiophene (BTBT)-conjugated peptides.

    Science.gov (United States)

    Khalily, Mohammad Aref; Usta, Hakan; Ozdemir, Mehmet; Bakan, Gokhan; Dikecoglu, F Begum; Edwards-Gayle, Charlotte; Hutchinson, Jessica A; Hamley, Ian W; Dana, Aykutlu; Guler, Mustafa O

    2018-05-18

    π-Conjugated small molecules based on a [1]benzothieno[3,2-b]benzothiophene (BTBT) unit are of great research interest in the development of solution-processable semiconducting materials owing to their excellent charge-transport characteristics. However, the BTBT π-core has yet to be demonstrated in the form of electro-active one-dimensional (1D) nanowires that are self-assembled in aqueous media for potential use in bioelectronics and tissue engineering. Here we report the design, synthesis, and self-assembly of benzothienobenzothiophene (BTBT)-peptide conjugates, the BTBT-peptide (BTBT-C3-COHN-Ahx-VVAGKK-Am) and the C8-BTBT-peptide (C8-BTBT-C3-COHN-Ahx-VVAGKK-Am), as β-sheet forming amphiphilic molecules, which self-assemble into highly uniform nanofibers in water with diameters of 11-13(±1) nm and micron-size lengths. Spectroscopic characterization studies demonstrate the J-type π-π interactions among the BTBT molecules within the hydrophobic core of the self-assembled nanofibers yielding an electrical conductivity as high as 6.0 × 10-6 S cm-1. The BTBT π-core is demonstrated, for the first time, in the formation of self-assembled peptide 1D nanostructures in aqueous media for potential use in tissue engineering, bioelectronics and (opto)electronics. The conductivity achieved here is one of the highest reported to date in a non-doped state.

  15. Antimicrobial Peptides in 2014

    Directory of Open Access Journals (Sweden)

    Guangshun Wang

    2015-03-01

    Full Text Available This article highlights new members, novel mechanisms of action, new functions, and interesting applications of antimicrobial peptides reported in 2014. As of December 2014, over 100 new peptides were registered into the Antimicrobial Peptide Database, increasing the total number of entries to 2493. Unique antimicrobial peptides have been identified from marine bacteria, fungi, and plants. Environmental conditions clearly influence peptide activity or function. Human α-defensin HD-6 is only antimicrobial under reduced conditions. The pH-dependent oligomerization of human cathelicidin LL-37 is linked to double-stranded RNA delivery to endosomes, where the acidic pH triggers the dissociation of the peptide aggregate to release its cargo. Proline-rich peptides, previously known to bind to heat shock proteins, are shown to inhibit protein synthesis. A model antimicrobial peptide is demonstrated to have multiple hits on bacteria, including surface protein delocalization. While cell surface modification to decrease cationic peptide binding is a recognized resistance mechanism for pathogenic bacteria, it is also used as a survival strategy for commensal bacteria. The year 2014 also witnessed continued efforts in exploiting potential applications of antimicrobial peptides. We highlight 3D structure-based design of peptide antimicrobials and vaccines, surface coating, delivery systems, and microbial detection devices involving antimicrobial peptides. The 2014 results also support that combination therapy is preferred over monotherapy in treating biofilms.

  16. Conformational assembly and biological properties of collagen mimetic peptides and their thermally responsive polymer conjugates

    Science.gov (United States)

    Krishna, Ohm Divyam

    2011-12-01

    Collagens are one of the most abundant proteins found in body tissues and organs, endowing structural integrity, mechanical strength, and multiple biological functions. Destabilized collagen inside human body leads to various degenerative diseases (ex. osteoarthritis) and ageing. This has continued to motivate the design of synthetic peptides and bio-synthetic polypeptides to closely mimic the native collagens in terms of triple helix structure and stability, potential for higher order assembly, and biological properties. However, the widespread application of de novo collagens has been limited in part by the need for hydroxylated proline in the formation of stable triple helical structures. To address this continued need, a hydroxyproline-free, thermally stable collagen-mimetic peptide (CLP-Cys) was rationally designed via the incorporation of electrostatically stabilized amino acid triplets. CLP-Cys was synthesized via solid phase peptide synthesis. The formation and stability of the triple helical structure were indicated via circular dichroism (CD) experiments and confirmed via differential scanning calorimetry (DSC) results. CLP-Cys also self-assembled into nano-rods and micro-fibrils, as evidenced via a combination of dynamic light scattering and transmission electron microscopy. Given the high thermal stability and its propensity for higher-order assembly, CLP-Cys was further functionalized at both the ends with a thermally responsive polymer, poly(diethylene glycol methyl ether methacrylate), (PDEGMEMA) to synthesize a biohybrid triblock copolymer. The CD results indicated that the triple helical form is retained, the thermal unfolding is sustained and helix to coil transition is reversible in the triblock hybrid context. The LCST of PDEGMEMA homopolymer (26 °C) is increased (to 35 °C) upon conjugation to the hydrophilic collagen peptide domain. Further, a combination of static light scattering, Cryo-SEM, TEM and confocal microscopy elucidated that the

  17. Potent peptidic fusion inhibitors of influenza virus

    Energy Technology Data Exchange (ETDEWEB)

    Kadam, Rameshwar U.; Juraszek, Jarek; Brandenburg, Boerries; Buyck, Christophe; Schepens, Wim B. G.; Kesteleyn, Bart; Stoops, Bart; Vreeken, Rob J.; Vermond, Jan; Goutier, Wouter; Tang, Chan; Vogels, Ronald; Friesen, Robert H. E.; Goudsmit, Jaap; van Dongen, Maria J. P.; Wilson, Ian A.

    2017-09-28

    Influenza therapeutics with new targets and mechanisms of action are urgently needed to combat potential pandemics, emerging viruses, and constantly mutating strains in circulation. We report here on the design and structural characterization of potent peptidic inhibitors of influenza hemagglutinin. The peptide design was based on complementarity-determining region loops of human broadly neutralizing antibodies against the hemagglutinin (FI6v3 and CR9114). The optimized peptides exhibit nanomolar affinity and neutralization against influenza A group 1 viruses, including the 2009 H1N1 pandemic and avian H5N1 strains. The peptide inhibitors bind to the highly conserved stem epitope and block the low pH–induced conformational rearrangements associated with membrane fusion. These peptidic compounds and their advantageous biological properties should accelerate the development of new small molecule– and peptide-based therapeutics against influenza virus.

  18. What can machine learning do for antimicrobial peptides, and what can antimicrobial peptides do for machine learning?

    Science.gov (United States)

    Lee, Ernest Y; Lee, Michelle W; Fulan, Benjamin M; Ferguson, Andrew L; Wong, Gerard C L

    2017-12-06

    Antimicrobial peptides (AMPs) are a diverse class of well-studied membrane-permeating peptides with important functions in innate host defense. In this short review, we provide a historical overview of AMPs, summarize previous applications of machine learning to AMPs, and discuss the results of our studies in the context of the latest AMP literature. Much work has been recently done in leveraging computational tools to design new AMP candidates with high therapeutic efficacies for drug-resistant infections. We show that machine learning on AMPs can be used to identify essential physico-chemical determinants of AMP functionality, and identify and design peptide sequences to generate membrane curvature. In a broader scope, we discuss the implications of our findings for the discovery of membrane-active peptides in general, and uncovering membrane activity in new and existing peptide taxonomies.

  19. The effect of peptides and ions interacting with an electrically neutral membrane interface on the structure and stability of lipid membranes in the liquid-crystalline phase and in the liquid-ordered phase

    Energy Technology Data Exchange (ETDEWEB)

    Sano, Ryoko [Department of Physics, Faculty of Science, Shizuoka University, Shizuoka, 422-8529 (Japan); Masum, Shah Md [Material Science, Graduate School of Science and Engineering, Shizuoka University, 422-8529 (Japan); Tanaka, Tomoki [Material Science, Graduate School of Science and Engineering, Shizuoka University, 422-8529 (Japan); Yamashita, Yuko [Department of Physics, Faculty of Science, Shizuoka University, Shizuoka, 422-8529 (Japan); Levadny, Victor [Department of Physics, Faculty of Science, Shizuoka University, Shizuoka, 422-8529 (Japan); Scientific Council for Cybernetics, Russian Academy of Sciences, Vavilov street 34, 333117, Moscow (Russian Federation); Yamazaki, Masahito [Department of Physics, Faculty of Science, Shizuoka University, Shizuoka, 422-8529 (Japan); Material Science, Graduate School of Science and Engineering, Shizuoka University, 422-8529 (Japan)

    2005-08-10

    We investigated the effects of a de novo designed peptide, WLFLLKKK (peptide-1) and La{sup 3+}, which can bind with the electrically neutral lipid membrane interface, on the stability of the phosphatidylcholine (PC) membrane in the L{sub {alpha}} phase and that of the liquid-ordered (lo) phase membranes. The results of spacing of the multilamellar vesicle and shape changes of the giant unilamellar vesicle (GUV) indicate that the peptide-1 can be partitioned into the membrane interface in the L{sub {alpha}} phase but not into that in the lo phase. La{sup 3+} induced shape changes of GUVs of the lo phase membrane, which are the same as those of GUVs in the L{sub {alpha}} phase. This indicates that the binding of La{sup 3+} induced an increase in the lateral compression pressure of the membrane, which decreased the surface area of the membrane in the lo phase. The difference of the membrane interface between the L{sub {alpha}} phase and the lo phase is discussed.

  20. Rationally designed turn promoting mutation in the amyloid-β peptide sequence stabilizes oligomers in solution.

    Directory of Open Access Journals (Sweden)

    Jayakumar Rajadas

    Full Text Available Enhanced production of a 42-residue beta amyloid peptide (Aβ(42 in affected parts of the brain has been suggested to be the main causative factor for the development of Alzheimer's Disease (AD. The severity of the disease depends not only on the amount of the peptide but also its conformational transition leading to the formation of oligomeric amyloid-derived diffusible ligands (ADDLs in the brain of AD patients. Despite being significant to the understanding of AD mechanism, no atomic-resolution structures are available for these species due to the evanescent nature of ADDLs that hinders most structural biophysical investigations. Based on our molecular modeling and computational studies, we have designed Met35Nle and G37p mutations in the Aβ(42 peptide (Aβ(42Nle35p37 that appear to organize Aβ(42 into stable oligomers. 2D NMR on the Aβ(42Nle35p37 peptide revealed the occurrence of two β-turns in the V24-N27 and V36-V39 stretches that could be the possible cause for the oligomer stability. We did not observe corresponding NOEs for the V24-N27 turn in the Aβ(21-43Nle35p37 fragment suggesting the need for the longer length amyloid peptide to form the stable oligomer promoting conformation. Because of the presence of two turns in the mutant peptide which were absent in solid state NMR structures for the fibrils, we propose, fibril formation might be hindered. The biophysical information obtained in this work could aid in the development of structural models for toxic oligomer formation that could facilitate the development of therapeutic approaches to AD.

  1. Peptide Based Radiopharmaceuticals: Specific Construct Approach

    Energy Technology Data Exchange (ETDEWEB)

    Som, P; Rhodes, B A; Sharma, S S

    1997-10-21

    The objective of this project was to develop receptor based peptides for diagnostic imaging and therapy. A series of peptides related to cell adhesion molecules (CAM) and immune regulation were designed for radiolabeling with 99mTc and evaluated in animal models as potential diagnostic imaging agents for various disease conditions such as thrombus (clot), acute kidney failure, and inflection/inflammation imaging. The peptides for this project were designed by the industrial partner, Palatin Technologies, (formerly Rhomed, Inc.) using various peptide design approaches including a newly developed rational computer assisted drug design (CADD) approach termed MIDAS (Metal ion Induced Distinctive Array of Structures). In this approach, the biological function domain and the 99mTc complexing domain are fused together so that structurally these domains are indistinguishable. This approach allows construction of conformationally rigid metallo-peptide molecules (similar to cyclic peptides) that are metabolically stable in-vivo. All the newly designed peptides were screened in various in vitro receptor binding and functional assays to identify a lead compound. The lead compounds were formulated in a one-step 99mTc labeling kit form which were studied by BNL for detailed in-vivo imaging using various animals models of human disease. Two main peptides usingMIDAS approach evolved and were investigated: RGD peptide for acute renal failure and an immunomodulatory peptide derived from tuftsin (RMT-1) for infection/inflammation imaging. Various RGD based metallopeptides were designed, synthesized and assayed for their efficacy in inhibiting ADP-induced human platelet aggregation. Most of these peptides displayed biological activity in the 1-100 µM range. Based on previous work by others, RGD-I and RGD-II were evaluated in animal models of acute renal failure. These earlier studies showed that after acute ischemic injury the renal cortex displays

  2. Multiarm-polyethylene glycol-polyglutamic acid peptide dendrimer: Design, synthesis, and dissolving thrombus.

    Science.gov (United States)

    Zhang, Shao-Fei; Lü, Shaoyu; Gao, Chunmei; Yang, Jiandong; Yan, Xiang; Li, Tao; Wen, Na; Huang, Mengjie; Liu, Mingzhu

    2018-06-01

    Thrombotic events affect many individuals in a number of ways, all of which can cause significant morbidity and mortality. Nattokinase (NK), as a novel thrombolytic drug, has been used for thrombolytic therapy. It not only possesses plasminogen activator activity, but also directly digests fibrin through limited proteolysis. However, it may undergo inactivation and denaturation in the harsh external environment. In this study, a multiarm-polyethylene glycol-polyglutamic acid peptide dendrimer was fabricated and used as a carrier for NK protection and delivery. Different arm numbers of polyethylene glycol-polyglutamic acid peptide dendrimers (x-PEG(G 3 ) x , x = 2, 4, 6, 8) were designed, prepared, and characterized by 1 H NMR and FTIR. Then, x-PEG(G 3 ) x were loaded with NK to form nanocomposites. Their size and morphology were determined by dynamic light scattering and transmission electron microscopy. Enzyme activity was evaluated via UV-Vis absorbance spectra, fluorescence spectra, circular dichroism spectra, and zeta potential measurements. The study reveals that the obtained x-PEG(G 3 ) x /NK nanocomposites possess high enzyme activity. In addition, the nanocomposites show increased viability of rat macrophage cells, and excellent thrombolysis ability in vitro and in vivo. This work establishes a multiarm-polyethylene glycol-polyglutamic acid peptide dendrimer with potential application in NK carrier and thrombolytic therapy. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 106A: 1687-1696, 2018. © 2018 Wiley Periodicals, Inc.

  3. A direct biocombinatorial strategy toward next generation, mussel-glue inspired saltwater adhesives.

    Science.gov (United States)

    Wilke, Patrick; Helfricht, Nicolas; Mark, Andreas; Papastavrou, Georg; Faivre, Damien; Börner, Hans G

    2014-09-10

    Biological materials exhibit remarkable, purpose-adapted properties that provide a source of inspiration for designing new materials to meet the requirements of future applications. For instance, marine mussels are able to attach to a broad spectrum of hard surfaces under hostile conditions. Controlling wet-adhesion of synthetic macromolecules by analogue processes promises to strongly impact materials sciences by offering advanced coatings, adhesives, and glues. The de novo design of macromolecules to mimic complex aspects of mussel adhesion still constitutes a challenge. Phage display allows material scientists to design specifically interacting molecules with tailored affinity to material surfaces. Here, we report on the integration of enzymatic processing steps into phage display biopanning to expand the biocombinatorial procedure and enable the direct selection of enzymatically activable peptide adhesion domains. Adsorption isotherms and single molecule force spectroscopy show that those de novo peptides mimic complex aspects of bioadhesion, such as enzymatic activation (by tyrosinase), the switchability from weak to strong binders, and adsorption under hostile saltwater conditions. Furthermore, peptide-poly(ethylene oxide) conjugates are synthesized to generate protective coatings, which possess anti-fouling properties and suppress irreversible interactions with blood-plasma protein cocktails. The extended phage display procedure provides a generic way to non-natural peptide adhesion domains, which not only mimic nature but also improve biological sequence sections extractable from mussel-glue proteins. The de novo peptides manage to combine several tasks in a minimal 12-mer sequence and thus pave the way to overcome major challenges of technical wet glues.

  4. Characterization of a human peptide deformylase: implications for antibacterial drug design.

    Science.gov (United States)

    Nguyen, Kiet T; Hu, Xubo; Colton, Craig; Chakrabarti, Ratna; Zhu, Michael X; Pei, Dehua

    2003-08-26

    Ribosomal protein synthesis in eubacteria and eukaryotic organelles initiates with an N-formylmethionyl-tRNA(i), resulting in N-terminal formylation of all nascent polypeptides. Peptide deformylase (PDF) catalyzes the subsequent removal of the N-terminal formyl group from the majority of bacterial proteins. Deformylation was for a long time thought to be a feature unique to the prokaryotes, making PDF an attractive target for designing novel antibiotics. However, recent genomic sequencing has revealed PDF-like sequences in many eukaryotes, including man. In this work, the cDNA encoding Homo sapiens PDF (HsPDF) has been cloned and a truncated form that lacks the N-terminal 58-amino-acid targeting sequence was overexpressed in Escherichia coli. The recombinant, Co(2+)-substituted protein is catalytically active in deformylating N-formylated peptides, shares many of the properties of bacterial PDF, and is strongly inhibited by specific PDF inhibitors. Expression of HsPDF fused to the enhanced green fluorescence protein in human embryonic kidney cells revealed its location in the mitochondrion. However, HsPDF is much less active than its bacterial counterpart, providing a possible explanation for the apparent lack of deformylation in the mammalian mitochondria. The lower catalytic activity is at least partially due to mutation of a highly conserved residue (Leu-91 in E. coli PDF) in mammalian PDF. PDF inhibitors had no detectable effect on two different human cell lines. These results suggest that HsPDF is likely an evolutional remnant without any functional role in protein formylation/deformylation and validates PDF as an excellent target for antibacterial drug design.

  5. A novel 13 residue acyclic peptide from the marine snail, Conus monile, targets potassium channels

    OpenAIRE

    Sudarslal, Sadasivannair; Singaravadivelan, Govindaswamy; Ramasamy, Palanisamy; Ananda, Kuppanna; Sarma, Siddhartha P; Sikdar, Sujit K; Krishnan, KS; Balaram, Padmanabhan

    2004-01-01

    A novel 13-residue peptide Mo1659 has been isolated from the venom of a vermivorous cone snail, Conus monile. HPLC fractions of the venom extract yielded an intense UV absorbing fraction with a mass of 1659 Da. De novo sequencing using both matrix assisted laser desorption and ionization and electrospray MS/MS methods together with analysis of proteolytic fragments successfully yielded the amino acid sequence, HGGSWYRFPWGY-NH2. This was further confirmed by comparison with the chemically synt...

  6. Peptide chemistry toolbox - Transforming natural peptides into peptide therapeutics.

    Science.gov (United States)

    Erak, Miloš; Bellmann-Sickert, Kathrin; Els-Heindl, Sylvia; Beck-Sickinger, Annette G

    2018-06-01

    The development of solid phase peptide synthesis has released tremendous opportunities for using synthetic peptides in medicinal applications. In the last decades, peptide therapeutics became an emerging market in pharmaceutical industry. The need for synthetic strategies in order to improve peptidic properties, such as longer half-life, higher bioavailability, increased potency and efficiency is accordingly rising. In this mini-review, we present a toolbox of modifications in peptide chemistry for overcoming the main drawbacks during the transition from natural peptides to peptide therapeutics. Modifications at the level of the peptide backbone, amino acid side chains and higher orders of structures are described. Furthermore, we are discussing the future of peptide therapeutics development and their impact on the pharmaceutical market. Copyright © 2018 Elsevier Ltd. All rights reserved.

  7. Synergistic Integration of Experimental and Simulation Approaches for the de Novo Design of Silk-Based Materials.

    Science.gov (United States)

    Huang, Wenwen; Ebrahimi, Davoud; Dinjaski, Nina; Tarakanova, Anna; Buehler, Markus J; Wong, Joyce Y; Kaplan, David L

    2017-04-18

    Tailored biomaterials with tunable functional properties are crucial for a variety of task-specific applications ranging from healthcare to sustainable, novel bio-nanodevices. To generate polymeric materials with predictive functional outcomes, exploiting designs from nature while morphing them toward non-natural systems offers an important strategy. Silks are Nature's building blocks and are produced by arthropods for a variety of uses that are essential for their survival. Due to the genetic control of encoded protein sequence, mechanical properties, biocompatibility, and biodegradability, silk proteins have been selected as prototype models to emulate for the tunable designs of biomaterial systems. The bottom up strategy of material design opens important opportunities to create predictive functional outcomes, following the exquisite polymeric templates inspired by silks. Recombinant DNA technology provides a systematic approach to recapitulate, vary, and evaluate the core structure peptide motifs in silks and then biosynthesize silk-based polymers by design. Post-biosynthesis processing allows for another dimension of material design by controlled or assisted assembly. Multiscale modeling, from the theoretical prospective, provides strategies to explore interactions at different length scales, leading to selective material properties. Synergy among experimental and modeling approaches can provide new and more rapid insights into the most appropriate structure-function relationships to pursue while also furthering our understanding in terms of the range of silk-based systems that can be generated. This approach utilizes nature as a blueprint for initial polymer designs with useful functions (e.g., silk fibers) but also employs modeling-guided experiments to expand the initial polymer designs into new domains of functional materials that do not exist in nature. The overall path to these new functional outcomes is greatly accelerated via the integration of

  8. Approaches in the design of 99mTc based peptide radiolabelling for tumour targeting

    International Nuclear Information System (INIS)

    Yokoyama, A.; Horiuchi, K.; Arano, Y.

    2001-01-01

    . Neogalactoalbumin (NGA), a specific protein that was incorporated by hepatic parenchymal cell via receptor-mediated endocytosis immediately after administration, was selected as a biological tool. The fate of the radiolabelled NGA after lysosomal proteolysis in hepatocytes was studied in liver homogenates. Subcellular distribution and identification of radiometabolites were performed using multiple analytical methods such as electrophoresis, TLC, and size exclusion and reverse phase HPLC. The data indicated that the chemical bonding between 99m Tc and HYNIC remains stable in the lysosomes but the persistent liver localization was due to a radiometabolite identified as 99m Tc-HYNIC-lysine (tricine)2, demonstrated by the similar HPLC data obtained with the synthesized and radiolabelled metabolite. Better co-ligands or new ligands or new design of the peptides are required in order to reduce the residence time of radioactivity in non-target tissue for future peptide use in radiodiagnosis or radiotherapy. (author)

  9. Peptides as Therapeutic Agents for Dengue Virus.

    Science.gov (United States)

    Chew, Miaw-Fang; Poh, Keat-Seong; Poh, Chit-Laa

    2017-01-01

    Dengue is an important global threat caused by dengue virus (DENV) that records an estimated 390 million infections annually. Despite the availability of CYD-TDV as a commercial vaccine, its long-term efficacy against all four dengue virus serotypes remains unsatisfactory. There is therefore an urgent need for the development of antiviral drugs for the treatment of dengue. Peptide was once a neglected choice of medical treatment but it has lately regained interest from the pharmaceutical industry following pioneering advancements in technology. In this review, the design of peptide drugs, antiviral activities and mechanisms of peptides and peptidomimetics (modified peptides) action against dengue virus are discussed. The development of peptides as inhibitors for viral entry, replication and translation is also described, with a focus on the three main targets, namely, the host cell receptors, viral structural proteins and viral non-structural proteins. The antiviral peptides designed based on these approaches may lead to the discovery of novel anti-DENV therapeutics that can treat dengue patients.

  10. Treating autoimmune disorders with venom-derived peptides.

    Science.gov (United States)

    Shen, Bingzheng; Cao, Zhijian; Li, Wenxin; Sabatier, Jean-Marc; Wu, Yingliang

    2017-09-01

    The effective treatment of autoimmune diseases remains a challenge. Voltage-gated potassium Kv1.3 channels, which are expressed in lymphocytes, are a new therapeutic target for treating autoimmune disease. Consequently, Kv1.3 channel-inhibiting venom-derived peptides are a prospective resource for new drug discovery and clinical application. Area covered: Preclinical and clinical studies have produced a wealth of information on Kv1.3 channel-inhibiting venom-derived peptides, especially from venomous scorpions and sea anemones. This review highlights the advances in screening and design of these peptides with diverse structures and potencies. It focuses on representative strategies for improving peptide selectivity and discusses the preclinical research on those venom-derived peptides as well as their clinical developmental status. Expert opinion: Encouraging results indicate that peptides isolated from the venom of venomous animals are a large resource for discovering immunomodulators that act on Kv1.3 channels. Since the structural diversity of venom-derived peptides determines the variety of their pharmacological activities, the design and optimization of venom-peptides for improved Kv1.3 channel-specificity has been advanced through some representative strategies, such as peptide chemical modification, amino acid residue truncation and binding interface modulation. These advances should further accelerate research, development and the future clinical application of venom-derived peptides selectively targeting Kv1.3 channels.

  11. Automated de novo phasing and model building of coiled-coil proteins.

    Science.gov (United States)

    Rämisch, Sebastian; Lizatović, Robert; André, Ingemar

    2015-03-01

    Models generated by de novo structure prediction can be very useful starting points for molecular replacement for systems where suitable structural homologues cannot be readily identified. Protein-protein complexes and de novo-designed proteins are examples of systems that can be challenging to phase. In this study, the potential of de novo models of protein complexes for use as starting points for molecular replacement is investigated. The approach is demonstrated using homomeric coiled-coil proteins, which are excellent model systems for oligomeric systems. Despite the stereotypical fold of coiled coils, initial phase estimation can be difficult and many structures have to be solved with experimental phasing. A method was developed for automatic structure determination of homomeric coiled coils from X-ray diffraction data. In a benchmark set of 24 coiled coils, ranging from dimers to pentamers with resolutions down to 2.5 Å, 22 systems were automatically solved, 11 of which had previously been solved by experimental phasing. The generated models contained 71-103% of the residues present in the deposited structures, had the correct sequence and had free R values that deviated on average by 0.01 from those of the respective reference structures. The electron-density maps were of sufficient quality that only minor manual editing was necessary to produce final structures. The method, named CCsolve, combines methods for de novo structure prediction, initial phase estimation and automated model building into one pipeline. CCsolve is robust against errors in the initial models and can readily be modified to make use of alternative crystallographic software. The results demonstrate the feasibility of de novo phasing of protein-protein complexes, an approach that could also be employed for other small systems beyond coiled coils.

  12. Are de novo acute heart failure and acutely worsened chronic heart failure two subgroups of the same syndrome?

    Directory of Open Access Journals (Sweden)

    Banović Marko

    2010-01-01

    Full Text Available Introduction. Acute heart failure (AHF is one of the most common diseases in emergency medicine, associated with poor prognosis and high in-hospital and long-term mortality. Objective. To investigate clinical presentation of patients with de novo AHF and acute worsening of chronic heart failure (CHF and to identify differences in blood levels of biomarkers and echocardiography findings. Methods. This prospective study comprised 64 consecutive patients being grouped according to the onset of the disease into patients with the de novo AHF (45.3%, and patients with acute worsening of CHF (54.7%. Results. Acute congestion (60% was the most common manifestation of de novo AHF, whereas pulmonary oedema (43.1% was the most common manifestation of acutely decompensated CHF. Patients with acutely decompensated CHF had significantly higher blood values of creatinine (147.10 vs 113.16 μmol/l; p<0.05, urea (12.63 vs 7.82 mmol/l; p<0.05, BNP (1440.11 vs 712.24 pg/ml; p<001 and NTproBNP (9097.00 vs 2827.70 pg/ml; p<0.01 on admission, and lower values of M-mode left ventricular ejection fraction (LVEF during hospitalization (49.44% vs 42.94%; p<0.05. The follow-up after one year revealed still significantly higher BNP (365.49 vs 164.02 pg/ ml; p<0.05 and lower average values of both LVEF in patients with acutely worsened CHF (46.62% vs 54.41% and 39.52% vs 47.88%; p<0.05. Conclusion. Considering differences in clinical severity on admission, echocardiography and natriuretic peptide values during hospitalization and after one year follow-up, de novo AHF and acutely worsened CHF are two different subgroups of the same syndrome.

  13. Peptide and Peptide-Dependent Motions in MHC Proteins: Immunological Implications and Biophysical Underpinnings

    Directory of Open Access Journals (Sweden)

    Cory M. Ayres

    2017-08-01

    Full Text Available Structural biology of peptides presented by class I and class II MHC proteins has transformed immunology, impacting our understanding of fundamental immune mechanisms and allowing researchers to rationalize immunogenicity and design novel vaccines. However, proteins are not static structures as often inferred from crystallographic structures. Their components move and breathe individually and collectively over a range of timescales. Peptides bound within MHC peptide-binding grooves are no exception and their motions have been shown to impact recognition by T cell and other receptors in ways that influence function. Furthermore, peptides tune the motions of MHC proteins themselves, which impacts recognition of peptide/MHC complexes by other proteins. Here, we review the motional properties of peptides in MHC binding grooves and discuss how peptide properties can influence MHC motions. We briefly review theoretical concepts about protein motion and highlight key data that illustrate immunological consequences. We focus primarily on class I systems due to greater availability of data, but segue into class II systems as the concepts and consequences overlap. We suggest that characterization of the dynamic “energy landscapes” of peptide/MHC complexes and the resulting functional consequences is one of the next frontiers in structural immunology.

  14. De novo origin of human protein-coding genes.

    Directory of Open Access Journals (Sweden)

    Dong-Dong Wu

    2011-11-01

    Full Text Available The de novo origin of a new protein-coding gene from non-coding DNA is considered to be a very rare occurrence in genomes. Here we identify 60 new protein-coding genes that originated de novo on the human lineage since divergence from the chimpanzee. The functionality of these genes is supported by both transcriptional and proteomic evidence. RNA-seq data indicate that these genes have their highest expression levels in the cerebral cortex and testes, which might suggest that these genes contribute to phenotypic traits that are unique to humans, such as improved cognitive ability. Our results are inconsistent with the traditional view that the de novo origin of new genes is very rare, thus there should be greater appreciation of the importance of the de novo origination of genes.

  15. De Novo Origin of Human Protein-Coding Genes

    Science.gov (United States)

    Wu, Dong-Dong; Irwin, David M.; Zhang, Ya-Ping

    2011-01-01

    The de novo origin of a new protein-coding gene from non-coding DNA is considered to be a very rare occurrence in genomes. Here we identify 60 new protein-coding genes that originated de novo on the human lineage since divergence from the chimpanzee. The functionality of these genes is supported by both transcriptional and proteomic evidence. RNA–seq data indicate that these genes have their highest expression levels in the cerebral cortex and testes, which might suggest that these genes contribute to phenotypic traits that are unique to humans, such as improved cognitive ability. Our results are inconsistent with the traditional view that the de novo origin of new genes is very rare, thus there should be greater appreciation of the importance of the de novo origination of genes. PMID:22102831

  16. Combined "de novo" and "ex novo" lipid fermentation in a mix-medium of corncob acid hydrolysate and soybean oil by Trichosporon dermatis.

    Science.gov (United States)

    Huang, Chao; Luo, Mu-Tan; Chen, Xue-Fang; Qi, Gao-Xiang; Xiong, Lian; Lin, Xiao-Qing; Wang, Can; Li, Hai-Long; Chen, Xin-De

    2017-01-01

    Microbial oil is one important bio-product for its important function in energy, chemical, and food industry. Finding suitable substrates is one key issue for its industrial application. Both hydrophilic and hydrophobic substrates can be utilized by oleaginous microorganisms with two different bio-pathways (" de novo " lipid fermentation and " ex novo " lipid fermentation). To date, most of the research on lipid fermentation has focused mainly on only one fermentation pathway and little work was carried out on both " de novo " and " ex novo " lipid fermentation simultaneously; thus, the advantages of both lipid fermentation cannot be fulfilled comprehensively. In this study, corncob acid hydrolysate with soybean oil was used as a mix-medium for combined " de novo " and " ex novo " lipid fermentation by oleaginous yeast Trichosporon dermatis . Both hydrophilic and hydrophobic substrates (sugars and soybean oil) in the medium can be utilized simultaneously and efficiently by T. dermatis . Different fermentation modes were compared and the batch mode was the most suitable for the combined fermentation. The influence of soybean oil concentration, inoculum size, and initial pH on the lipid fermentation was evaluated and 20 g/L soybean oil, 5% inoculum size, and initial pH 6.0 were suitable for this bioprocess. By this technology, the lipid composition of extracellular hydrophobic substrate (soybean oil) can be modified. Although adding emulsifier showed little beneficial effect on lipid production, it can modify the intracellular lipid composition of T. dermatis . The present study proves the potential and possibility of combined " de novo " and " ex novo " lipid fermentation. This technology can use hydrophilic and hydrophobic sustainable bio-resources to generate lipid feedstock for the production of biodiesel or other lipid-based chemical compounds and to treat some special wastes such as oil-containing wastewater.

  17. Rational design of DKK3 structure-based small peptides as antagonists of Wnt signaling pathway and in silico evaluation of their efficiency.

    Directory of Open Access Journals (Sweden)

    Mansour Poorebrahim

    Full Text Available Dysregulated Wnt signaling pathway is highly associated with the pathogenesis of several human cancers. Dickkopf proteins (DKKs are thought to inhibit Wnt signaling pathway through binding to lipoprotein receptor-related protein (LRP 5/6. In this study, based on the 3-dimensional (3D structure of DKK3 Cys-rich domain 2 (CRD2, we have designed and developed several peptide inhibitors of Wnt signaling pathway. Modeller 9.15 package was used to predict 3D structure of CRD2 based on the Homology modeling (HM protocol. After refinement and minimization with GalaxyRefine and NOMAD-REF servers, the quality of selected models was evaluated utilizing VADAR, SAVES and ProSA servers. Molecular docking studies as well as literature-based information revealed two distinct boxes located at CRD2 which are actively involved in the DKK3-LRP5/6 interaction. A peptide library was constructed conducting the backrub sequence tolerance scanning protocol in Rosetta3.5 according to the DKK3-LRP5/6 binding sites. Seven tolerated peptides were chosen and their binding affinity and stability were improved by some logical amino acid substitutions. Molecular dynamics (MD simulations of peptide-LRP5/6 complexes were carried out using GROMACS package. After evaluation of binding free energies, stability, electrostatic potential and some physicochemical properties utilizing computational approaches, three peptides (PEP-I1, PEP-I3 and PEP-II2 demonstrated desirable features. However, all seven improved peptides could sufficiently block the Wnt-binding site of LRP6 in silico. In conclusion, we have designed and improved several small peptides based on the LRP6-binding site of CRD2 of DKK3. These peptides are highly capable of binding to LRP6 in silico, and may prevent the formation of active Wnt-LRP6-Fz complex.

  18. Design of Cyclic Peptide Based Glucose Receptors and Their Application in Glucose Sensing.

    Science.gov (United States)

    Li, Chao; Chen, Xin; Zhang, Fuyuan; He, Xingxing; Fang, Guozhen; Liu, Jifeng; Wang, Shuo

    2017-10-03

    Glucose assay is of great scientific significance in clinical diagnostics and bioprocess monitoring, and to design a new glucose receptor is necessary for the development of more sensitive, selective, and robust glucose detection techniques. Herein, a series of cyclic peptide (CP) glucose receptors were designed to mimic the binding sites of glucose binding protein (GBP), and CPs' sequence contained amino acid sites Asp, Asn, His, Asp, and Arg, which constituted the first layer interactions of GBP. The properties of these CPs used as a glucose receptor or substitute for the GBP were studied by using a quartz crystal microbalance (QCM) technique. It was found that CPs can form a self-assembled monolayer at the Au quartz electrode surface, and the monolayer's properties were characterized by using cyclic voltammetry, electrochemical impedance spectroscopy, and atomic force microscopy. The CPs' binding affinity to saccharide (i.e., galactose, fructose, lactose, sucrose, and maltose) was investigated, and the CPs' sensitivity and selectivity toward glucose were found to be dependent upon the configuration,i.e., the amino acids sequence of the CPs. The cyclic unit with a cyclo[-CNDNHCRDNDC-] sequence gave the highest selectivity and sensitivity for glucose sensing. This work suggests that a synthetic peptide bearing a particular functional sequence could be applied for developing a new generation of glucose receptors and would find huge application in biological, life science, and clinical diagnostics fields.

  19. Genes from scratch--the evolutionary fate of de novo genes.

    Science.gov (United States)

    Schlötterer, Christian

    2015-04-01

    Although considered an extremely unlikely event, many genes emerge from previously noncoding genomic regions. This review covers the entire life cycle of such de novo genes. Two competing hypotheses about the process of de novo gene birth are discussed as well as the high death rate of de novo genes. Despite the high death rate, some de novo genes are retained and remain functional, even in distantly related species, through their integration into gene networks. Further studies combining gene expression with ribosome profiling in multiple populations across different species will be instrumental for an improved understanding of the evolutionary processes operating on de novo genes. Copyright © 2015 The Author. Published by Elsevier Ltd.. All rights reserved.

  20. Multifunctional hybrid networks based on self assembling peptide sequences

    Science.gov (United States)

    Sathaye, Sameer

    The overall aim of this dissertation is to achieve a comprehensive correlation between the molecular level changes in primary amino acid sequences of amphiphilic beta-hairpin peptides and their consequent solution-assembly properties and bulk network hydrogel behavior. This has been accomplished using two broad approaches. In the first approach, amino acid substitutions were made to peptide sequence MAX1 such that the hydrophobic surfaces of the folded beta-hairpins from the peptides demonstrate shape specificity in hydrophobic interactions with other beta-hairpins during the assembly process, thereby causing changes to the peptide nanostructure and bulk rheological properties of hydrogels formed from the peptides. Steric lock and key complementary hydrophobic interactions were designed to occur between two beta-hairpin molecules of a single molecule, LNK1 during beta-sheet fibrillar assembly of LNK1. Experimental results from circular dichroism, transmission electron microscopy and oscillatory rheology collectively indicate that the molecular design of the LNK1 peptide can be assigned the cause of the drastically different behavior of the networks relative to MAX1. The results indicate elimination or significant reduction of fibrillar branching due to steric complementarity in LNK1 that does not exist in MAX1, thus supporting the original hypothesis. As an extension of the designed steric lock and key complementarity between two beta-hairpin molecules of the same peptide molecule. LNK1, three new pairs of peptide molecules LP1-KP1, LP2-KP2 and LP3-KP3 that resemble complementary 'wedge' and 'trough' shapes when folded into beta-hairpins were designed and studied. All six peptides individually and when blended with their corresponding shape complement formed fibrillar nanostructures with non-uniform thickness values. Loose packing in the assembled structures was observed in all the new peptides as compared to the uniform tight packing in MAX1 by SANS analysis. This

  1. Stabilized helical peptides: overview of the technologies and its impact on drug discovery.

    Science.gov (United States)

    Klein, Mark

    2017-11-01

    Protein-protein interactions are predominant in the workings of all cells. Until now, there have been a few successes in targeting protein-protein interactions with small molecules. Peptides may overcome some of the challenges of small molecules in disrupting protein-protein interactions. However, peptides present a new set of challenges in drug discovery. Thus, the study of the stabilization of helical peptides has been extensive. Areas covered: Several technological approaches to helical peptide stabilization have been studied. In this review, stapled peptides, foldamers, and hydrogen bond surrogates are discussed. Issues regarding design principles are also discussed. Furthermore, this review introduces select computational techniques used to aid peptide design and discusses clinical trials of peptides in a more advanced stage of development. Expert opinion: Stabilized helical peptides hold great promise in a wide array of diseases. However, the field is still relatively new and new design principles are emerging. The possibilities of peptide modification are quite extensive and expanding, so the design of stabilized peptides requires great attention to detail in order to avoid a large number of failed lead peptides. The start of clinical trials with stapled peptides is a promising sign for the future.

  2. Computationally assisted screening and design of cell-interactive peptides by a cell-based assay using peptide arrays and a fuzzy neural network algorithm.

    Science.gov (United States)

    Kaga, Chiaki; Okochi, Mina; Tomita, Yasuyuki; Kato, Ryuji; Honda, Hiroyuki

    2008-03-01

    We developed a method of effective peptide screening that combines experiments and computational analysis. The method is based on the concept that screening efficiency can be enhanced from even limited data by use of a model derived from computational analysis that serves as a guide to screening and combining the model with subsequent repeated experiments. Here we focus on cell-adhesion peptides as a model application of this peptide-screening strategy. Cell-adhesion peptides were screened by use of a cell-based assay of a peptide array. Starting with the screening data obtained from a limited, random 5-mer library (643 sequences), a rule regarding structural characteristics of cell-adhesion peptides was extracted by fuzzy neural network (FNN) analysis. According to this rule, peptides with unfavored residues in certain positions that led to inefficient binding were eliminated from the random sequences. In the restricted, second random library (273 sequences), the yield of cell-adhesion peptides having an adhesion rate more than 1.5-fold to that of the basal array support was significantly high (31%) compared with the unrestricted random library (20%). In the restricted third library (50 sequences), the yield of cell-adhesion peptides increased to 84%. We conclude that a repeated cycle of experiments screening limited numbers of peptides can be assisted by the rule-extracting feature of FNN.

  3. Peptide ligands for targeting the extracellular domain of EGFR: Comparison between linear and cyclic peptides.

    Science.gov (United States)

    Williams, Tyrslai M; Sable, Rushikesh; Singh, Sitanshu; Vicente, Maria Graca H; Jois, Seetharama D

    2018-02-01

    Colorectal cancer (CRC) is the third most common solid internal malignancy among cancers. Early detection of cancer is key to increasing the survival rate of colorectal cancer patients. Overexpression of the EGFR protein is associated with CRC. We have designed a series of peptides that are highly specific for the extracellular domain of EGFR, based on our earlier studies on linear peptides. The previously reported linear peptide LARLLT, known to bind to EGFR, was modified with the goals of increasing its stability and its specificity toward EGFR. Peptide modifications, including D-amino acid substitution, cyclization, and chain reversal, were investigated. In addition, to facilitate labeling of the peptide with a fluorescent dye, an additional lysine residue was introduced onto the linear (KLARLLT) and cyclic peptides cyclo(KLARLLT) (Cyclo.L1). The lysine residue was also converted into an azide group in both a linear and reversed cyclic peptide sequences cyclo(K(N3)larllt) (Cyclo.L1.1) to allow for subsequent "click" conjugation. The cyclic peptides showed enhanced binding to EGFR by SPR. NMR and molecular modeling studies suggest that the peptides acquire a β-turn structure in solution. In vitro stability studies in human serum show that the cyclic peptide is more stable than the linear peptide. © 2017 John Wiley & Sons A/S.

  4. Osmoprotection of Bacillus subtilis through Import and Proteolysis of Proline-Containing Peptides

    Science.gov (United States)

    Zaprasis, Adrienne; Brill, Jeanette; Thüring, Marietta; Wünsche, Guido; Heun, Magnus; Barzantny, Helena; Hoffmann, Tamara

    2013-01-01

    Bacillus subtilis can attain cellular protection against the detrimental effects of high osmolarity through osmotically induced de novo synthesis and uptake of the compatible solute l-proline. We have now found that B. subtilis can also exploit exogenously provided proline-containing peptides of various lengths and compositions as osmoprotectants. Osmoprotection by these types of peptides is generally dependent on their import via the peptide transport systems (Dpp, Opp, App, and DtpT) operating in B. subtilis and relies on their hydrolysis to liberate proline. The effectiveness with which proline-containing peptides confer osmoprotection varies considerably, and this can be correlated with the amount of the liberated and subsequently accumulated free proline by the osmotically stressed cell. Through gene disruption experiments, growth studies, and the quantification of the intracellular proline pool, we have identified the PapA (YqhT) and PapB (YkvY) peptidases as responsible for the hydrolysis of various types of Xaa-Pro dipeptides and Xaa-Pro-Xaa tripeptides. The PapA and PapB peptidases possess overlapping substrate specificities. In contrast, osmoprotection by peptides of various lengths and compositions with a proline residue positioned at their N terminus was not affected by defects in the PapA and PapB peptidases. Taken together, our data provide new insight into the physiology of the osmotic stress response of B. subtilis. They illustrate the flexibility of this ubiquitously distributed microorganism to effectively exploit environmental resources in its acclimatization to sustained high-osmolarity surroundings through the accumulation of compatible solutes. PMID:23144141

  5. Plasmin substrate binding site cooperativity guides the design of potent peptide aldehyde inhibitors.

    Science.gov (United States)

    Swedberg, Joakim E; Harris, Jonathan M

    2011-10-04

    Perioperative bleeding is a cause of major blood loss and is associated with increased rates of postoperative morbidity and mortality. To combat this, antifibrinolytic inhibitors of the serine protease plasmin are commonly used to reduce bleeding during surgery. The most effective and previously widely used of these is the broad range serine protease inhibitor aprotinin. However, adverse clinical outcomes have led to use of alternative serine lysine analogues to inhibit plasmin. These compounds suffer from low selectivity and binding affinity. Consequently, a concerted effort to discover potent and selective plasmin inhibitors has developed. This study used a noncombinatorial peptide library to define plasmin's extended substrate specificity and guide the design of potent transition state analogue inhibitors. The various substrate binding sites of plasmin were found to exhibit a higher degree of cooperativity than had previously been appreciated. Peptide sequences capitalizing on these features produced high-affinity inhibitors of plasmin. The most potent of these, Lys-Met(sulfone)-Tyr-Arg-H [KM(O(2))YR-H], inhibited plasmin with a K(i) of 3.1 nM while maintaining 25-fold selectivity over plasma kallikrein. Furthermore, 125 nM (0.16 μg/mL) KM(O(2))YR-H attenuated fibrinolysis in vitro with an efficacy similar to that of 15 nM (0.20 μg/mL) aprotinin. To date, this is the most potent peptide inhibitor of plasmin that exhibits selectivity against plasma kallikrein, making this compound an attractive candidate for further therapeutic development.

  6. De novo assembly of the perennial ryegrass transcriptome using an RNA-Seq strategy.

    Directory of Open Access Journals (Sweden)

    Jacqueline D Farrell

    Full Text Available Perennial ryegrass is a highly heterozygous outbreeding grass species used for turf and forage production. Heterozygosity can affect de-Bruijn graph assembly making de novo transcriptome assembly of species such as perennial ryegrass challenging. Creating a reference transcriptome from a homozygous perennial ryegrass genotype can circumvent the challenge of heterozygosity. The goals of this study were to perform RNA-sequencing on multiple tissues from a highly inbred genotype to develop a reference transcriptome. This was complemented with RNA-sequencing of a highly heterozygous genotype for SNP calling.De novo transcriptome assembly of the inbred genotype created 185,833 transcripts with an average length of 830 base pairs. Within the inbred reference transcriptome 78,560 predicted open reading frames were found of which 24,434 were predicted as complete. Functional annotation found 50,890 transcripts with a BLASTp hit from the Swiss-Prot non-redundant database, 58,941 transcripts with a Pfam protein domain and 1,151 transcripts encoding putative secreted peptides. To evaluate the reference transcriptome we targeted the high-affinity K+ transporter gene family and found multiple orthologs. Using the longest unique open reading frames as the reference sequence, 64,242 single nucleotide polymorphisms were found. One thousand sixty one open reading frames from the inbred genotype contained heterozygous sites, confirming the high degree of homozygosity.Our study has developed an annotated, comprehensive transcriptome reference for perennial ryegrass that can aid in determining genetic variation, expression analysis, genome annotation, and gene mapping.

  7. Self-Assembly of Peptides at the Air/Water Interface

    Science.gov (United States)

    Sayar, Mehmet

    2013-03-01

    Peptides are commonly used as building blocks for design and development of novel materials with a variety of application areas ranging from drug design to biotechnology. The precise control of molecular architecture and specific nature of the nonbonded interactions among peptides enable aggregates with well defined structural and functional properties. The interaction of peptides with interfaces leads to dramatic changes in their conformational and aggregation behavior. In this talk, I will discuss our research on the interplay of intermolecular forces and influence of interfaces. In the first part the amphiphilic nature of short peptide oligomers and their behavior at the air/water interface will be discussed. The surface driving force and its decomposition will be analyzed. In the second part aggregation of peptides in bulk water and at an interface will be discussed. Different design features which can be tuned to control aggregation behavior will be analyzed.

  8. The non-peptidic part determines the internalization mechanism and intracellular trafficking of peptide amphiphiles.

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    Dimitris Missirlis

    Full Text Available BACKGROUND: Peptide amphiphiles (PAs are a class of amphiphilic molecules able to self-assemble into nanomaterials that have shown efficient in vivo targeted delivery. Understanding the interactions of PAs with cells and the mechanisms of their internalization and intracellular trafficking is critical in their further development for therapeutic delivery applications. METHODOLOGY/PRINCIPAL FINDINGS: PAs of a novel, cell- and tissue-penetrating peptide were synthesized possessing two different lipophilic tail architectures and their interactions with prostate cancer cells were studied in vitro. Cell uptake of peptides was greatly enhanced post-modification. Internalization occurred via lipid-raft mediated endocytosis and was common for the two analogs studied. On the contrary, we identified the non-peptidic part as the determining factor of differences between intracellular trafficking and retention of PAs. PAs composed of di-stearyl lipid tails linked through poly(ethylene glycol to the peptide exhibited higher exocytosis rates and employed different recycling pathways compared to ones consisting of di-palmitic-coupled peptides. As a result, cell association of the former PAs decreased with time. CONCLUSIONS/SIGNIFICANCE: Control over peptide intracellular localization and retention is possible by appropriate modification with synthetic hydrophobic tails. We propose this as a strategy to design improved peptide-based delivery systems.

  9. Structural Interplay - Tuning Mechanics in Peptide-Polyurea Hybrids

    Science.gov (United States)

    Korley, Lashanda

    Utilizing cues from natural materials, we have been inspired to explore the hierarchical arrangement critical to energy absorption and mechanical enhancement in synthetic systems. Of particular interest is the soft domain ordering proposed as a contributing element to the observed toughness in spider silk. Multiblock copolymers, are ideal and dynamic systems in which to explore this approach via variations in secondary structure of nature's building blocks - peptides. We have designed a new class of polyurea hybrids that incorporate peptidic copolymers as the soft segment. The impact of hierarchical ordering on the thermal, mechanical, and morphological behavior of these bio-inspired polyurethanes with a siloxane-based, peptide soft segment was investigated. These peptide-polyurethane/urea hybrids were microphase segregated, and the beta-sheet secondary structure of the soft segment was preserved during polymerization and film casting. Toughness enhancement at low strains was achieved, but the overall extensibility of the peptide-incorporated systems was reduced due to the unique hard domain organization. To decouple the secondary structure influence in the siloxane-peptide soft segment from mechanics dominated by the hard domain, we also developed non-chain extended peptide-polyurea hybrids in which the secondary structure (beta sheet vs. alpha helix) was tuned via choice of peptide and peptide length. It was shown that this structural approach allowed tailoring of extensibility, toughness, and modulus. The sheet-dominant hybrid materials were typically tougher and more elastic due to intermolecular H-bonding facilitating load distribution, while the helical-prevalent systems generally exhibited higher stiffness. Recently, we have explored the impact of a molecular design strategy that overlays a covalent and physically crosslinked architecture in these peptide-polyurea hybrids, demonstrating that physical constraints in the network hybrids influences peptide

  10. A novel peptide thrombopoietin mimetic designing and optimization using computational approach

    Directory of Open Access Journals (Sweden)

    Vimal kishor Singh

    2016-08-01

    Full Text Available Thrombopoietin receptor (TPOR is a cytokine receptor protein; activation of cell surface TPOR by thrombopoietin (TPO triggers an intracellular cascade of megakaryocytopoiesis for the formation of platelets. TPO is a glycoprotein hormone which stimulates megakaryocytes formation and maturation to platelets in bone marrow. Ex vivo megakaryocytopoiesis is in highlight for its vast role in therapeutics and field of regenerative medicine. For therapeutic uses, various TPO alternatives have been used however they are associated with issues like recombinant TPO administration is associated with the generation of auto antibodies and its production is an expensive process. Moreover, reported thrombopoietin mimetic peptide (TMP has no sequence homology with TPO and low specificity to TPOR. Hence, in this study, a novel peptidic TPO mimetic is designed through computational studies by studying the binding sites of TPO and TMP to TPOR and analogs of known mimetics. Screening of combinatorial library was done through molecular docking using ClusPro and mimetic-9 was found to have better binding score of -938.8 kcal/mol with seven hydrogen bonds and a high number of hydrophobic interactions, than known mimetic TMP with docking score of -798.4 kcal/mol and TMP dimer with docking score of -811.9 kcal/mol for TPOR. Mimetic-9 interaction with TPOR was further assessed by the molecular dynamics simulation and their complex was found to be stable with an RMSD value of 0.091 Aº. Resulting mimetic-9 was found to have overall good physiochemical properties with positive grand average hydropathy (GRAVY score and high instability index score and it was found to be localized in the extracellular region. The designed mimetic-9 might prove to be a useful lead molecule for mimicking the role of TPO and to help for platelet production in vitro with higher efficiency.

  11. Novel Formulations for Antimicrobial Peptides

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    Ana Maria Carmona-Ribeiro

    2014-10-01

    Full Text Available Peptides in general hold much promise as a major ingredient in novel supramolecular assemblies. They may become essential in vaccine design, antimicrobial chemotherapy, cancer immunotherapy, food preservation, organs transplants, design of novel materials for dentistry, formulations against diabetes and other important strategical applications. This review discusses how novel formulations may improve the therapeutic index of antimicrobial peptides by protecting their activity and improving their bioavailability. The diversity of novel formulations using lipids, liposomes, nanoparticles, polymers, micelles, etc., within the limits of nanotechnology may also provide novel applications going beyond antimicrobial chemotherapy.

  12. Novel Formulations for Antimicrobial Peptides

    Science.gov (United States)

    Carmona-Ribeiro, Ana Maria; Carrasco, Letícia Dias de Melo

    2014-01-01

    Peptides in general hold much promise as a major ingredient in novel supramolecular assemblies. They may become essential in vaccine design, antimicrobial chemotherapy, cancer immunotherapy, food preservation, organs transplants, design of novel materials for dentistry, formulations against diabetes and other important strategical applications. This review discusses how novel formulations may improve the therapeutic index of antimicrobial peptides by protecting their activity and improving their bioavailability. The diversity of novel formulations using lipids, liposomes, nanoparticles, polymers, micelles, etc., within the limits of nanotechnology may also provide novel applications going beyond antimicrobial chemotherapy. PMID:25302615

  13. High-resolution mass spectrometry driven discovery of peptidic danger signals in insect immunity.

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    Arton Berisha

    Full Text Available The 'danger model' is an alternative concept for immune response postulating that the immune system reacts to entities that do damage (danger associated molecular patterns, DAMP and not only to entities that are foreign (pathogen-associated molecular patterns, PAMP as proposed by classical immunology concepts. In this study we used Galleria mellonella to validate the danger model in insects. Hemolymph of G. mellonella was digested with thermolysin (as a representative for virulence-associated metalloproteinases produced by humanpathogens followed by chromatographic fractionation. Immune-stimulatory activity was tested by measuring lysozyme activity with the lytic zone assays against Micrococcus luteus cell wall components. Peptides were analyzed by nano-scale liquid chromatography coupled to high-resolution Fourier transform mass spectrometers. Addressing the lack of a genome sequence we complemented the rudimentary NCBI protein database with a recently established transcriptome and de novo sequencing methods for peptide identification. This approach led to identification of 127 peptides, 9 of which were identified in bioactive fractions. Detailed MS/MS experiments in comparison with synthetic analogues confirmed the amino acid sequence of all 9 peptides. To test the potential of these putative danger signals to induce immune responses we injected the synthetic analogues into G. mellonella and monitored the anti-bacterial activity against living Micrococcus luteus. Six out of 9 peptides identified in the bioactive fractions exhibited immune-stimulatory activity when injected. Hence, we provide evidence that small peptides resulting from thermolysin-mediated digestion of hemolymph proteins function as endogenous danger signals which can set the immune system into alarm. Consequently, our study indicates that the danger model also plays a role in insect immunity.

  14. Rational design and synthesis of altered peptide ligands based on human myelin oligodendrocyte glycoprotein 35-55 epitope: inhibition of chronic experimental autoimmune encephalomyelitis in mice.

    Science.gov (United States)

    Tselios, Theodore; Aggelidakis, Mihalis; Tapeinou, Anthi; Tseveleki, Vivian; Kanistras, Ioannis; Gatos, Dimitrios; Matsoukas, John

    2014-11-04

    Experimental autoimmune encephalomyelitis (EAE) is a demyelinating disease of the central nervous system and is an animal model of multiple sclerosis (MS). Although the etiology of MS remains unclear, there is evidence T-cell recognition of immunodominant epitopes of myelin proteins, such as the 35-55 epitope of myelin oligodendrocyte glycoprotein (MOG), plays a pathogenic role in the induction of chronic EAE. Cyclization of peptides is of great interest since the limited stability of linear peptides restricts their potential use as therapeutic agents. Herein, we have designed and synthesized a number of linear and cyclic peptides by mutating crucial T cell receptor (TCR) contact residues of the human MOG35-55 epitope. In particular, we have designed and synthesized cyclic altered peptide ligands (APLs) by mutating Arg41 with Ala or Arg41 and Arg46 with Ala. The peptides were synthesized in solid phase on 2-chlorotrityl chloride resin (CLTR-Cl) using the Fmoc/t-Bu methodology. The purity of final products was verified by RP-HPLC and their identification was achieved by ESI-MS. It was found that the substitutions of Arg at positions 41 and 46 with Ala results in peptide analogues that reduce the severity of MOG-induced EAE clinical symptoms in C57BL/6 mice when co-administered with mouse MOG35-55 peptide at the time of immunization.

  15. Self-assembly of fibronectin mimetic peptide-amphiphile nanofibers

    Science.gov (United States)

    Rexeisen, Emilie Lynn

    Many therapeutic strategies incorporate peptides into their designs to mimic the natural protein ligands found in vivo. A few examples are the short peptide sequences RGD and PHSRN that mimic the primary and synergy-binding domains of the extracellular matrix protein, fibronectin, which is recognized by the cell surface receptor, alpha5beta 1 integrin. Even though scaffold modification with biomimetic peptides remains one of the most promising approaches for tissue engineering, the use of these peptides in therapeutic tissue-engineered products and drug delivery systems available on the commercial market is limited because the peptides are not easily able to mimic the natural protein. The design of a peptide that can effectively target the alpha5beta1 integrin would greatly increase biomimetic scaffold therapeutic potential. A novel peptide containing both the RGD primary binding domain and PHSRN synergy-binding domain for fibronectin joined with the appropriate linker should bind alpha 5beta1 integrin more efficiently and lead to greater cell adhesion over RGD alone. Several fibronectin mimetic peptides were designed and coupled to dialkyl hydrocarbon tails to make peptide-amphiphiles. The peptides contained different linkers connecting the two binding domains and different spacers separating the hydrophobic tails from the hydrophilic headgroups. The peptide-amphiphiles were deposited on mica substrates using the Langmuir-Blodgett technique. Langmuir isotherms indicated that the peptide-amphiphiles that contained higher numbers of serine residues formed a more tightly packed monolayer, but the increased number of serines also made transferring the amphiphiles to the mica substrate more difficult. Atomic force microscopy (AFM) images of the bilayers showed that the headgroups might be bent, forming small divots in the surface. These divots may help expose the PHSRN synergy-binding domain. Parallel studies undertaken by fellow group members showed that human

  16. A de novo NADPH generation pathway for improving lysine production of Corynebacterium glutamicum by rational design of the coenzyme specificity of glyceraldehyde 3-phosphate dehydrogenase.

    Science.gov (United States)

    Bommareddy, Rajesh Reddy; Chen, Zhen; Rappert, Sugima; Zeng, An-Ping

    2014-09-01

    Engineering the cofactor availability is a common strategy of metabolic engineering to improve the production of many industrially important compounds. In this work, a de novo NADPH generation pathway is proposed by altering the coenzyme specificity of a native NAD-dependent glyceraldehyde 3-phosphate dehydrogenase (GAPDH) to NADP, which consequently has the potential to produce additional NADPH in the glycolytic pathway. Specifically, the coenzyme specificity of GAPDH of Corynebacterium glutamicum is systematically manipulated by rational protein design and the effect of the manipulation for cellular metabolism and lysine production is evaluated. By a combinatorial modification of four key residues within the coenzyme binding sites, different GAPDH mutants with varied coenzyme specificity were constructed. While increasing the catalytic efficiency of GAPDH towards NADP enhanced lysine production in all of the tested mutants, the most significant improvement of lysine production (~60%) was achieved with the mutant showing similar preference towards both NAD and NADP. Metabolic flux analysis with (13)C isotope studies confirmed that there was no significant change of flux towards the pentose phosphate pathway and the increased lysine yield was mainly attributed to the NADPH generated by the mutated GAPDH. The present study highlights the importance of protein engineering as a key strategy in de novo pathway design and overproduction of desired products. Copyright © 2014 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

  17. De novo assembly of highly diverse viral populations

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    Yang Xiao

    2012-09-01

    Full Text Available Abstract Background Extensive genetic diversity in viral populations within infected hosts and the divergence of variants from existing reference genomes impede the analysis of deep viral sequencing data. A de novo population consensus assembly is valuable both as a single linear representation of the population and as a backbone on which intra-host variants can be accurately mapped. The availability of consensus assemblies and robustly mapped variants are crucial to the genetic study of viral disease progression, transmission dynamics, and viral evolution. Existing de novo assembly techniques fail to robustly assemble ultra-deep sequence data from genetically heterogeneous populations such as viruses into full-length genomes due to the presence of extensive genetic variability, contaminants, and variable sequence coverage. Results We present VICUNA, a de novo assembly algorithm suitable for generating consensus assemblies from genetically heterogeneous populations. We demonstrate its effectiveness on Dengue, Human Immunodeficiency and West Nile viral populations, representing a range of intra-host diversity. Compared to state-of-the-art assemblers designed for haploid or diploid systems, VICUNA recovers full-length consensus and captures insertion/deletion polymorphisms in diverse samples. Final assemblies maintain a high base calling accuracy. VICUNA program is publicly available at: http://www.broadinstitute.org/scientific-community/science/projects/viral-genomics/ viral-genomics-analysis-software. Conclusions We developed VICUNA, a publicly available software tool, that enables consensus assembly of ultra-deep sequence derived from diverse viral populations. While VICUNA was developed for the analysis of viral populations, its application to other heterogeneous sequence data sets such as metagenomic or tumor cell population samples may prove beneficial in these fields of research.

  18. Voltammetry and In Situ Scanning Tunnelling Microscopy of De Novo Designed Heme Protein Monolayers on Au(111)-Electrode Surfaces

    DEFF Research Database (Denmark)

    Albrecht, Tim; Li, Wu; Haehnel, Wolfgang

    2006-01-01

    to the tunnelling current, apparently due to slow electron transfer kinetics. As a consequence, STM images of heme-containing and heme-free MOP-C did not reveal any notable differences in apparent height or physical extension. The apparent height of heme-containing MOP-C did not show any dependence on the substrate...... potential being varied around the redox potential of the protein. The mere presence of an accessible molecular energy level is not sufficient to result in detectable tunnelling current modulation. (c) 2006 Elsevier B.V. All rights reserved.......In the present work, we report the electrochemical characterization and in situ scanning tunnelling microscopy (STM) studies of monolayers of an artificial de novo designed heme protein MOP-C, covalently immobilized on modified Au(111) surfaces. The protein forms closely packed monolayers, which...

  19. Rational Design of Cancer-Targeted Benzoselenadiazole by RGD Peptide Functionalization for Cancer Theranostics.

    Science.gov (United States)

    Yang, Liye; Li, Wenying; Huang, Yanyu; Zhou, Yangliang; Chen, Tianfeng

    2015-09-01

    A cancer-targeted conjugate of the selenadiazole derivative BSeC (benzo[1,2,5] selenadiazole-5-carboxylic acid) with RGD peptide as targeting molecule and PEI (polyethylenimine) as a linker is rationally designed and synthesized in the present study. The results show that RGD-PEI-BSeC forms nanoparticles in aqueous solution with a core-shell nanostructure and high stability under physiological conditions. This rational design effectively enhances the selective cellular uptake and cellular retention of BSeC in human glioma cells, and increases its selectivity between cancer and normal cells. The nanoparticles enter the cells through receptor-mediated endocytosis via clathrin-mediated and nystatin-dependent lipid raft-mediated pathways. Internalized nanoparticles trigger glioma cell apoptosis by activation of ROS-mediated p53 phosphorylation. Therefore, this study provides a strategy for the rational design of selenium-containing cancer-targeted theranostics. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  20. Treatment of Experimental Brain Tumors with Trombospondin-1 Derived Peptides: an In Vivo Imaging Study

    Directory of Open Access Journals (Sweden)

    A. Bogdanov, Jr.

    1999-11-01

    Full Text Available Antiangiogenic and antiproliferative effects of synthetic D-reverse peptides derived from the type 1 repeats of thrombospondin (TSP1 [1,2] were studied in rodent C6 glioma and 9L gliosarcomas. To directly measure tumor size and vascular parameters, we employed in vivo magnetic resonance (MR imaging and corroborated results by traditional morphometric tissue analysis. Rats bearing either C6 or 9L tumors were treated with TSP1-derived peptide (D-reverse amKRFKQDGGWSHWSPWSSac, n=13 or a control peptide (D-reverse am KRAKQAGGASHASPASSac, n=12 at 10 mg/kg, administered either intravenously or through subcutaneous miniosmotic pumps starting 10 days after tumor implantation. Eleven days later, the effect of peptide treatment was evaluated. TSP1 peptide-treated 9L tumors (50.7±44.2 mm3, n=7 and C6 tumors (41.3±34.2 mm3, n=6 were significantly smaller than tumors treated with control peptide (9L: 215.7±67.8 mm3, n=6; C6:184.2±105.2 mm3, n=6. In contrast, the in vivo vascular volume fraction, the mean vascular area (determined by microscopy, and the microvascular density of tumors were not significantly different in any of the experimental groups. In cell culture, TSP1, and the amKRFKQDGGWSHWSPWSSac peptide showed antiproliferative effects against C6 with an IC of 45 nM for TSP1. These results indicate that TSP1derived peptides retard brain tumor growth presumably as a result of slower de novo blood vessel formation and synergistic direct antiproliferative effects on tumor cells. We also show that in vivo MR imaging can be used to assess treatment efficacy of novel antiangiogenic drugs non-invasively, which has obvious implications for clinical trials.

  1. Recognition of GPCRs by peptide ligands and membrane compartments theory: structural studies of endogenous peptide hormones in membrane environment.

    Science.gov (United States)

    Sankararamakrishnan, Ramasubbu

    2006-04-01

    One of the largest family of cell surface proteins, G-protein coupled receptors (GPCRs) regulate virtually all known physiological processes in mammals. With seven transmembrane segments, they respond to diverse range of extracellular stimuli and represent a major class of drug targets. Peptidergic GPCRs use endogenous peptides as ligands. To understand the mechanism of GPCR activation and rational drug design, knowledge of three-dimensional structure of receptor-ligand complex is important. The endogenous peptide hormones are often short, flexible and completely disordered in aqueous solution. According to "Membrane Compartments Theory", the flexible peptide binds to the membrane in the first step before it recognizes its receptor and the membrane-induced conformation is postulated to bind to the receptor in the second step. Structures of several peptide hormones have been determined in membrane-mimetic medium. In these studies, micelles, reverse micelles and bicelles have been used to mimic the cell membrane environment. Recently, conformations of two peptide hormones have also been studied in receptor-bound form. Membrane environment induces stable secondary structures in flexible peptide ligands and membrane-induced peptide structures have been correlated with their bioactivity. Results of site-directed mutagenesis, spectroscopy and other experimental studies along with the conformations determined in membrane medium have been used to interpret the role of individual residues in the peptide ligand. Structural differences of membrane-bound peptides that belong to the same family but differ in selectivity are likely to explain the mechanism of receptor selectivity and specificity of the ligands. Knowledge of peptide 3D structures in membrane environment has potential applications in rational drug design.

  2. Bicyclic peptide inhibitor of urokinase-type plasminogen activator

    DEFF Research Database (Denmark)

    Roodbeen, Renée; Jensen, Berit Paaske; Jiang, Longguang

    2013-01-01

    The development of protease inhibitors for pharmacological intervention has taken a new turn with the use of peptide-based inhibitors. Here, we report the rational design of bicyclic peptide inhibitors of the serine protease urokinase-type plasminogen activator (uPA), based on the established...... investigated the solution structures of the bicyclic peptide by NMR spectroscopy to map possible conformations. An X-ray structure of the bicyclic-peptide-uPA complex confirmed an interaction similar to that for the previous upain-1/upain-2-uPA complexes. These physical studies of the peptide...

  3. Iterative design of peptide-based hydrogels and the effect of network electrostatics on primary chondrocyte behavior.

    Science.gov (United States)

    Sinthuvanich, Chomdao; Haines-Butterick, Lisa A; Nagy, Katelyn J; Schneider, Joel P

    2012-10-01

    Iterative peptide design was used to generate two peptide-based hydrogels to study the effect of network electrostatics on primary chondrocyte behavior. MAX8 and HLT2 peptides have formal charge states of +7 and +5 per monomer, respectively. These peptides undergo triggered folding and self-assembly to afford hydrogel networks having similar rheological behavior and local network morphologies, yet different electrostatic character. Each gel can be used to directly encapsulate and syringe-deliver cells. The influence of network electrostatics on cell viability after encapsulation and delivery, extracellular matrix deposition, gene expression, and the bulk mechanical properties of the gel-cell constructs as a function of culture time was assessed. The less electropositive HLT2 gel provides a microenvironment more conducive to chondrocyte encapsulation, delivery, and phenotype maintenance. Cell viability was higher for this gel and although a moderate number of cells dedifferentiated to a fibroblast-like phenotype, many retained their chondrocytic behavior. As a result, gel-cell constructs prepared with HLT2, cultured under static in vitro conditions, contained more GAG and type II collagen resulting in mechanically superior constructs. Chondrocytes delivered in the more electropositive MAX8 gel experienced a greater degree of cell death during encapsulation and delivery and the remaining viable cells were less prone to maintain their phenotype. As a result, MAX8 gel-cell constructs had fewer cells, of which a limited number were capable of laying down cartilage-specific ECM. Published by Elsevier Ltd.

  4. 13-Helix folding of a β/γ-peptide manifold designed from a "minimal-constraint" blueprint.

    Science.gov (United States)

    Grison, Claire M; Robin, Sylvie; Aitken, David J

    2016-06-14

    A bottom-up design rationale was adopted to devise β/γ-peptide foldamer manifolds which would adopt preferred 13-helix conformations, relying on minimal steric imposition brought by the constituent amino acid residues. In this way, a well-defined 13-helix conformer was revealed for short oligomers of trans-2-aminocyclobutanecarboxylic acid and γ(4)-amino acids in alternation, which gave good topological superposition upon an α-helix motif.

  5. Modular design of synthetic protein mimics. Characterization of the helical conformation of a 13-residue peptide in crystals

    International Nuclear Information System (INIS)

    Karle, I.L.; Flippen-Anderson, J.L.; Uma, K.; Balaram, P.

    1989-01-01

    The incorporation of α-aminoisobutyryl (Aib) residues into peptide sequences facilitates helical folding. Aib-containing sequences have been chosen for the design of rigid helical segments in a modular approach to the construction of a synthetic protein mimic. The helical conformation of the synthetic peptide Boc-Aib-(Val-Ala-Leu-Aib) 3 -OMe in crystals is established by X-ray diffraction. The 13-residue apolar peptide adopts a helical form in the crystal with seven α-type hydrogen bonds in the middle and 3 10 -type hydrogen bonds at either end. The helices stack in columns, zigzag rather than linear, by means of direct NH hor-ellipsis OC head to tail hydrogen bonds. Leucyl side chains are extended on one side of the helix and valyl side chains on the other side. Water molecules form hydrogen bonds with several backbone carbonyl oxygens that also participate in α-helix hydrogen bonds. There is no apparent distortion of the helix caused by hydration

  6. Genome-wide patterns and properties of de novo mutations in humans

    NARCIS (Netherlands)

    Francioli, Laurent C.; Polak, Paz P.; Koren, Amnon; Menelaou, Androniki; Chun, Sung; Renkens, Ivo; van Duijn, Cornelia M.; Swertz, Morris; Wijmenga, Cisca; van Ommen, Gertjan; Slagboom, P. Eline; Boomsma, Dorret I.; Ye, Kai; Guryev, Victor; Arndt, Peter F.; Kloosterman, Wigard P.; de Bakker, Paul I. W.; Sunyaev, Shamil R.

    Mutations create variation in the population, fuel evolution and cause genetic diseases. Current knowledge about de novo mutations is incomplete and mostly indirect(1-10). Here we analyze 11,020 de novo mutations from the whole genomes of 250 families. We show that de novo mutations in the offspring

  7. Genome-wide patterns and properties of de novo mutations in humans

    NARCIS (Netherlands)

    Francioli, L.C.; Polak, P.P.; Koren, A.; Menelaou, A.; Chun, S.; Renkens, I.; van Duijn, C.M.; Swertz, M.A.; Wijmenga, C.; van Ommen, G.J.; Slagboom, P.E.; Boomsma, D.I.; Ye, K.; Guryev, V.; Arndt, P.F.; Kloosterman, W.P.; Bakker, P.I.W.; Sunyaev, S.R.; Dijk, F.; Neerincx, P.B.T.; Pulit, S.L.; Deelen, P.; Elbers, C.C.; Palamara, P.F.; Pe'er, I.; Abdellaoui, A.; van Oven, M.; Vermaat, M.; Li, M.; Laros, J.F.J.; Stoneking, M.; de Knijff, P.; Kayser, M.; Veldink, J.H.; Van den Berg, L.H.; Byelas, H.; den Dunnen, J.T.; Dijkstra, M.; Amin, N.; van der Velde, K.J.; Hottenga, J.J.; van Setten, J.; van Leeuwen, E.M.; Kanterakis, A.; Kattenberg, V.M.; Karssen, L.C.; van Schaik, B.D.C.; Bot, J.; Nijman, I.J.; van Enckevort, D.; Mei, H.; Koval, V.; Estrada, K.; Medina-Gomez, C.; Lameijer, E.W.; Moed, M.H.; Hehir-Kwa, J.Y.; Handsaker, R.E.; McCarroll, S.A.; Vuzman, D.; Sohail, M.; Hormozdiari, F.; Marschall, T.; Schönhuth, A.; Beekman, M.; de Craen, A.J.; Suchiman, H.E.D.; Hofman, A.; Oostra, B.; Isaacs, A.; Rivadeneira, F.; Uitterlinden, A.G.; Willemsen, G.; Platteel, M.; Pitts, S.J.; Potluri, S.; Sundar, P.; Cox, D.R.; Li, Q.; Li, Y.; Du, Y.; Chen, R.; Cao, H.; Li, N.; Cao, S.; Wang, J.; Bovenberg, J.A.; Brandsma, M.

    2015-01-01

    Mutations create variation in the population, fuel evolution and cause genetic diseases. Current knowledge about de novo mutations is incomplete and mostly indirect. Here we analyze 11,020 de novo mutations from the whole genomes of 250 families. We show that de novo mutations in the offspring of

  8. Peptide design using alpha,beta-dehydro amino acids: from beta-turns to helical hairpins.

    Science.gov (United States)

    Mathur, Puniti; Ramakumar, S; Chauhan, V S

    2004-01-01

    Incorporation of alpha,beta-dehydrophenylalanine (DeltaPhe) residue in peptides induces folded conformations: beta-turns in short peptides and 3(10)-helices in larger ones. A few exceptions-namely, alpha-helix or flat beta-bend ribbon structures-have also been reported in a few cases. The most favorable conformation of DeltaPhe residues are (phi,psi) approximately (-60 degrees, -30 degrees ), (-60 degrees, 150 degrees ), (80 degrees, 0 degrees ) or their enantiomers. DeltaPhe is an achiral and planar residue. These features have been exploited in designing DeltaPhe zippers and helix-turn-helix motifs. DeltaPhe can be incorporated in both right and left-handed helices. In fact, consecutive occurrence of three or more DeltaPhe amino acids induce left-handed screw sense in peptides containing L-amino acids. Weak interactions involving the DeltaPhe residue play an important role in molecular association. The C--H.O==C hydrogen bond between the DeltaPhe side-chain and backbone carboxyl moiety, pi-pi stacking interactions between DeltaPhe side chains belonging to enantiomeric helices have shown to stabilize folding. The unusual capability of a DeltaPhe ring to form the hub of multicentered interactions namely, a donor in aromatic C--H.pi and C--H.O==C and an acceptor in a CH(3).pi interaction suggests its exploitation in introducing long-range interactions in the folding of supersecondary structures. Copyright 2004 Wiley Periodicals, Inc. Biopolymers (Pept Sci), 2004

  9. Clinicopathologic factors associated with de novo metastatic breast cancer.

    Science.gov (United States)

    Shen, Tiansheng; Siegal, Gene P; Wei, Shi

    2016-12-01

    While breast cancers with distant metastasis at presentation (de novo metastasis) harbor significantly inferior clinical outcomes, there have been limited studies analyzing the clinicopathologic characteristics in this subset of patients. In this study, we analyzed 6126 breast cancers diagnosed between 1998 and 2013 to identify factors associated with de novo metastatic breast cancer. When compared to patients without metastasis at presentation, race, histologic grade, estrogen/progesterone receptor (ER/PR) and HER2 statuses were significantly associated with de novo metastasis in the entire cohort, whereas age, histologic grade, PR and HER2 status were the significant parameters in the subset of patients with locally advanced breast cancer (Stage IIB/III). The patients with de novo metastatic breast cancer had a significant older mean age and a lower proportion of HER2-positive tumors when compared to those with metastatic recurrence. Further, the HER2-rich subtype demonstrated a drastically higher incidence of de novo metastasis when compared to the luminal and triple-negative breast cancers in the entire cohort [odds ratio (OR)=5.68 and 2.27, respectively] and in the patients with locally advanced disease (OR=4.02 and 2.12, respectively), whereas no significant difference was seen between de novo metastatic cancers and those with metastatic recurrence. Moreover, the luminal and HER2-rich subtypes showed bone-seeking (OR=1.92) and liver-homing (OR=2.99) characteristics, respectively, for the sites of de novo metastasis, while the latter was not observed in those with metastatic recurrence. Our data suggest that an algorithm incorporating clinicopathologic factors, especially histologic grade and receptor profile, remains of significant benefit during decision making in newly diagnosed breast cancer in the pursuit of precision medicine. Copyright © 2016 Elsevier GmbH. All rights reserved.

  10. Foldability of a Natural De Novo Evolved Protein.

    Science.gov (United States)

    Bungard, Dixie; Copple, Jacob S; Yan, Jing; Chhun, Jimmy J; Kumirov, Vlad K; Foy, Scott G; Masel, Joanna; Wysocki, Vicki H; Cordes, Matthew H J

    2017-11-07

    The de novo evolution of protein-coding genes from noncoding DNA is emerging as a source of molecular innovation in biology. Studies of random sequence libraries, however, suggest that young de novo proteins will not fold into compact, specific structures typical of native globular proteins. Here we show that Bsc4, a functional, natural de novo protein encoded by a gene that evolved recently from noncoding DNA in the yeast S. cerevisiae, folds to a partially specific three-dimensional structure. Bsc4 forms soluble, compact oligomers with high β sheet content and a hydrophobic core, and undergoes cooperative, reversible denaturation. Bsc4 lacks a specific quaternary state, however, existing instead as a continuous distribution of oligomer sizes, and binds dyes indicative of amyloid oligomers or molten globules. The combination of native-like and non-native-like properties suggests a rudimentary fold that could potentially act as a functional intermediate in the emergence of new folded proteins de novo. Copyright © 2017 Elsevier Ltd. All rights reserved.

  11. Particulated articular cartilage: CAIS and DeNovo NT.

    Science.gov (United States)

    Farr, Jack; Cole, Brian J; Sherman, Seth; Karas, Vasili

    2012-03-01

    Cartilage Autograft Implantation System (CAIS; DePuy/Mitek, Raynham, MA) and DeNovo Natural Tissue (NT; ISTO, St. Louis, MO) are novel treatment options for focal articular cartilage defects in the knee. These methods involve the implantation of particulated articular cartilage from either autograft or juvenile allograft donor, respectively. In the laboratory and in animal models, both CAIS and DeNovo NT have demonstrated the ability of the transplanted cartilage cells to "escape" from the extracellular matrix, migrate, multiply, and form a new hyaline-like cartilage tissue matrix that integrates with the surrounding host tissue. In clinical practice, the technique for both CAIS and DeNovo NT is straightforward, requiring only a single surgery to affect cartilage repair. Clinical experience is limited, with short-term studies demonstrating both procedures to be safe, feasible, and effective, with improvements in subjective patient scores, and with magnetic resonance imaging evidence of good defect fill. While these treatment options appear promising, prospective randomized controlled studies are necessary to refine the indications and contraindications for both CAIS and DeNovo NT.

  12. Efficient assembly of de novo human artificial chromosomes from large genomic loci

    Directory of Open Access Journals (Sweden)

    Stromberg Gregory

    2005-07-01

    Full Text Available Abstract Background Human Artificial Chromosomes (HACs are potentially useful vectors for gene transfer studies and for functional annotation of the genome because of their suitability for cloning, manipulating and transferring large segments of the genome. However, development of HACs for the transfer of large genomic loci into mammalian cells has been limited by difficulties in manipulating high-molecular weight DNA, as well as by the low overall frequencies of de novo HAC formation. Indeed, to date, only a small number of large (>100 kb genomic loci have been reported to be successfully packaged into de novo HACs. Results We have developed novel methodologies to enable efficient assembly of HAC vectors containing any genomic locus of interest. We report here the creation of a novel, bimolecular system based on bacterial artificial chromosomes (BACs for the construction of HACs incorporating any defined genomic region. We have utilized this vector system to rapidly design, construct and validate multiple de novo HACs containing large (100–200 kb genomic loci including therapeutically significant genes for human growth hormone (HGH, polycystic kidney disease (PKD1 and ß-globin. We report significant differences in the ability of different genomic loci to support de novo HAC formation, suggesting possible effects of cis-acting genomic elements. Finally, as a proof of principle, we have observed sustained ß-globin gene expression from HACs incorporating the entire 200 kb ß-globin genomic locus for over 90 days in the absence of selection. Conclusion Taken together, these results are significant for the development of HAC vector technology, as they enable high-throughput assembly and functional validation of HACs containing any large genomic locus. We have evaluated the impact of different genomic loci on the frequency of HAC formation and identified segments of genomic DNA that appear to facilitate de novo HAC formation. These genomic loci

  13. Marine Peptides: Bioactivities and Applications

    Directory of Open Access Journals (Sweden)

    Randy Chi Fai Cheung

    2015-06-01

    Full Text Available Peptides are important bioactive natural products which are present in many marine species. These marine peptides have high potential nutraceutical and medicinal values because of their broad spectra of bioactivities. Their antimicrobial, antiviral, antitumor, antioxidative, cardioprotective (antihypertensive, antiatherosclerotic and anticoagulant, immunomodulatory, analgesic, anxiolytic anti-diabetic, appetite suppressing and neuroprotective activities have attracted the attention of the pharmaceutical industry, which attempts to design them for use in the treatment or prevention of various diseases. Some marine peptides or their derivatives have high commercial values and had reached the pharmaceutical and nutraceutical markets. A large number of them are already in different phases of the clinical and preclinical pipeline. This review highlights the recent research in marine peptides and the trends and prospects for the future, with special emphasis on nutraceutical and pharmaceutical development into marketed products.

  14. A highly scalable peptide-based assay system for proteomics.

    Directory of Open Access Journals (Sweden)

    Igor A Kozlov

    Full Text Available We report a scalable and cost-effective technology for generating and screening high-complexity customizable peptide sets. The peptides are made as peptide-cDNA fusions by in vitro transcription/translation from pools of DNA templates generated by microarray-based synthesis. This approach enables large custom sets of peptides to be designed in silico, manufactured cost-effectively in parallel, and assayed efficiently in a multiplexed fashion. The utility of our peptide-cDNA fusion pools was demonstrated in two activity-based assays designed to discover protease and kinase substrates. In the protease assay, cleaved peptide substrates were separated from uncleaved and identified by digital sequencing of their cognate cDNAs. We screened the 3,011 amino acid HCV proteome for susceptibility to cleavage by the HCV NS3/4A protease and identified all 3 known trans cleavage sites with high specificity. In the kinase assay, peptide substrates phosphorylated by tyrosine kinases were captured and identified by sequencing of their cDNAs. We screened a pool of 3,243 peptides against Abl kinase and showed that phosphorylation events detected were specific and consistent with the known substrate preferences of Abl kinase. Our approach is scalable and adaptable to other protein-based assays.

  15. ChemTS: an efficient python library for de novo molecular generation

    Science.gov (United States)

    Yang, Xiufeng; Zhang, Jinzhe; Yoshizoe, Kazuki; Terayama, Kei; Tsuda, Koji

    2017-12-01

    Automatic design of organic materials requires black-box optimization in a vast chemical space. In conventional molecular design algorithms, a molecule is built as a combination of predetermined fragments. Recently, deep neural network models such as variational autoencoders and recurrent neural networks (RNNs) are shown to be effective in de novo design of molecules without any predetermined fragments. This paper presents a novel Python library ChemTS that explores the chemical space by combining Monte Carlo tree search and an RNN. In a benchmarking problem of optimizing the octanol-water partition coefficient and synthesizability, our algorithm showed superior efficiency in finding high-scoring molecules. ChemTS is available at https://github.com/tsudalab/ChemTS.

  16. Vaccinomics Approach for Designing Potential Peptide Vaccine by Targeting Shigella spp. Serine Protease Autotransporter Subfamily Protein SigA

    Directory of Open Access Journals (Sweden)

    Arafat Rahman Oany

    2017-01-01

    Full Text Available Shigellosis, a bacillary dysentery, is closely associated with diarrhoea in human and causes infection of 165 million people worldwide per year. Casein-degrading serine protease autotransporter of enterobacteriaceae (SPATE subfamily protein SigA, an outer membrane protein, exerts both cytopathic and enterotoxic effects especially cytopathic to human epithelial cell type-2 (HEp-2 and is shown to be highly immunogenic. In the present study, we have tried to impose the vaccinomics approach for designing a common peptide vaccine candidate against the immunogenic SigA of Shigella spp. At first, 44 SigA proteins from different variants of S. flexneri, S. dysenteriae, S. boydii, and S. sonnei were assessed to find the most antigenic protein. We retrieved 12 peptides based on the highest score for human leukocyte antigen (HLA supertypes analysed by NetCTL. Initially, these peptides were assessed for the affinity with MHC class I and class II alleles, and four potential core epitopes VTARAGLGY, FHTVTVNTL, HTTWTLTGY, and IELAGTLTL were selected. From these, FHTVTVNTL and IELAGTLTL peptides were shown to have 100% conservancy. Finally, IELAGTLTL was shown to have the highest population coverage (83.86% among the whole world population. In vivo study of the proposed epitope might contribute to the development of functional and unique widespread vaccine, which might be an operative alleyway to thwart dysentery from the world.

  17. Can Natural Proteins Designed with ‘Inverted’ Peptide Sequences Adopt Native-Like Protein Folds?

    Science.gov (United States)

    Sridhar, Settu; Guruprasad, Kunchur

    2014-01-01

    We have carried out a systematic computational analysis on a representative dataset of proteins of known three-dimensional structure, in order to evaluate whether it would possible to ‘swap’ certain short peptide sequences in naturally occurring proteins with their corresponding ‘inverted’ peptides and generate ‘artificial’ proteins that are predicted to retain native-like protein fold. The analysis of 3,967 representative proteins from the Protein Data Bank revealed 102,677 unique identical inverted peptide sequence pairs that vary in sequence length between 5–12 and 18 amino acid residues. Our analysis illustrates with examples that such ‘artificial’ proteins may be generated by identifying peptides with ‘similar structural environment’ and by using comparative protein modeling and validation studies. Our analysis suggests that natural proteins may be tolerant to accommodating such peptides. PMID:25210740

  18. Insect Peptides - Perspectives in Human Diseases Treatment.

    Science.gov (United States)

    Chowanski, Szymon; Adamski, Zbigniew; Lubawy, Jan; Marciniak, Pawel; Pacholska-Bogalska, Joanna; Slocinska, Malgorzata; Spochacz, Marta; Szymczak, Monika; Urbanski, Arkadiusz; Walkowiak-Nowicka, Karolina; Rosinski, Grzegorz

    2017-01-01

    Insects are the largest and the most widely distributed group of animals in the world. Their diversity is a source of incredible variety of different mechanisms of life processes regulation. There are many agents that regulate immunology, reproduction, growth and development or metabolism. Hence, it seems that insects may be a source of numerous substances useful in human diseases treatment. Especially important in the regulation of insect physiology are peptides, like neuropeptides, peptide hormones or antimicrobial peptides. There are two main aspects where they can be helpful, 1) Peptides isolated from insects may become potential drugs in therapy of different diseases, 2) A lot of insect peptide hormones show structural or functional homology to mammalian peptide hormones and the comparative studies may give a new look on human disorders. In our review we focused on three group of insect derived peptides: 1) immune-active peptides, 2) peptide hormones and 3) peptides present in venoms. In our review we try to show the considerable potential of insect peptides in searching for new solutions for mammalian diseases treatment. We summarise the knowledge about properties of insect peptides against different virulent agents, anti-inflammatory or anti-nociceptive properties as well as compare insect and mammalian/vertebrate peptide endocrine system to indicate usefulness of knowledge about insect peptide hormones in drug design. The field of possible using of insect delivered peptide to therapy of various human diseases is still not sufficiently explored. Undoubtedly, more attention should be paid to insects due to searching new drugs. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  19. Peptide drugs to target G protein-coupled receptors.

    Science.gov (United States)

    Bellmann-Sickert, Kathrin; Beck-Sickinger, Annette G

    2010-09-01

    Major indications for use of peptide-based therapeutics include endocrine functions (especially diabetes mellitus and obesity), infectious diseases, and cancer. Whereas some peptide pharmaceuticals are drugs, acting as agonists or antagonists to directly treat cancer, others (including peptide diagnostics and tumour-targeting pharmaceuticals) use peptides to 'shuttle' a chemotherapeutic agent or a tracer to the tumour and allow sensitive imaging or targeted therapy. Significant progress has been made in the last few years to overcome disadvantages in peptide design such as short half-life, fast proteolytic cleavage, and low oral bioavailability. These advances include peptide PEGylation, lipidisation or multimerisation; the introduction of peptidomimetic elements into the sequences; and innovative uptake strategies such as liposomal, capsule or subcutaneous formulations. This review focuses on peptides targeting G protein-coupled receptors that are promising drug candidates or that have recently entered the pharmaceutical market. Copyright 2010 Elsevier Ltd. All rights reserved.

  20. ''de novo'' aneurysms following endovascular procedures

    Energy Technology Data Exchange (ETDEWEB)

    Briganti, F.; Cirillo, S.; Caranci, F. [Department of Neurological Sciences, Services of Neuroradiology, ' ' Federico II' ' University, Naples (Italy); Esposito, F.; Maiuri, F. [Department of Neurological Sciences, Services of Neurosurgery, ' ' Federico II' ' University, Naples (Italy)

    2002-07-01

    Two personal cases of ''de novo'' aneurysms of the anterior communicating artery (ACoA) occurring 9 and 4 years, respectively, after endovascular carotid occlusion are described. A review of the 30 reported cases (including our own two) of ''de novo'' aneurysms after occlusion of the major cerebral vessels has shown some features, including a rather long time interval after the endovascular procedure of up to 20-25 years (average 9.6 years), a preferential ACoA (36.3%) and internal carotid artery-posterior communicating artery (ICA-PCoA) (33.3%) location of the ''de novo'' aneurysms, and a 10% rate of multiple aneurysms. These data are compared with those of the group of reported spontaneous ''de novo'' aneurysms after SAH or previous aneurysm clipping. We agree that the frequency of ''de novo'' aneurysms after major-vessel occlusion (two among ten procedures in our series, or 20%) is higher than commonly reported (0 to 11%). For this reason, we suggest that patients who have been submitted to endovascular major-vessel occlusion be followed up for up to 20-25 years after the procedure, using non-invasive imaging studies such as MR angiography and high-resolution CT angiography. On the other hand, periodic digital angiography has a questionable risk-benefit ratio; it may be used when a ''de novo'' aneurysm is detected or suspected on non-invasive studies. The progressive enlargement of the ACoA after carotid occlusion, as described in our case 1, must be considered a radiological finding of risk for ''de novo'' aneurysm formation. (orig.)

  1. Generation of Hybrid Peptide-Silver Nanoparticles for Antibacterial and Antifouling Applications

    KAUST Repository

    Seferji, Kholoud

    2018-05-01

    An alarming increase of antibiotic-resistant bacterial strains has made the demand for novel antibacterial agents, for example, more effective antibiotics, highly crucial. One of the oldest antimicrobial agents is elementary silver which has been used for thousands of years. Even in our days, elementary silver is used for medical purposes, such as for burns, wounds, and microbial infections. We have taken the effectiveness of elementary silver into consideration to generate novel antibacterial and antifouling agents. Our innovative antibacterial agents are hybrid peptide silver nanoparticles (CH-01-AgNPs) that are created de novo and in situ from a silver nitrate solution (AgNO3) in the presence of ultrashort self-assembling peptides compounds. The nucleation of CH-01-AgNPs is initiated by irradiating the peptide solution mixed with the AgNO3 solution using ultraviolet (UV) light at a wavelength of 254 nm, in the absence of any reducing or capping agents. Obviously, the peptide itself serves as the reducing agent. The ultrashort peptides are four amino acids in length with an innate ability to self-assemble into nanofibrous scaffolds. Using these ultrashort peptides CH-01 we were able to create hybrid peptide silver nanoparticles CH-01-AgNPs with a diameter of 4-6 nm. The synthesized CH-01-AgNPs were further characterized using ultraviolet-visible spectroscopy, transmission electron microscopy, dynamic light scattering, and X-ray photoelectron spectroscopy. The antibacterial and antifouling activity of CH-01-AgNPs were then investigated using either gram-negative bacteria, such as antibiotic-resistant Top10 Escherichia coli and Pseudomonas aeruginosa PDO300, or gram-positive bacteria, such as Staphylococcus aureus CECT 976. The hybrid nanoparticles demonstrated very promising antibacterial and antifouling activity with higher antibacterial and antifouling activity as commercial silver nanoparticles. Quantitative Polymerase Chain Reaction (qPCR) results showed

  2. How Nature Morphs Peptide Scaffolds into Antibiotics

    Science.gov (United States)

    Nolan, Elizabeth M.; Walsh, Christopher T.

    2010-01-01

    The conventional notion that peptides are poor candidates for orally available drugs because of protease-sensitive peptide bonds, intrinsic hydrophilicity, and ionic charges contrasts with the diversity of antibiotic natural products with peptide-based frameworks that are synthesized and utilized by Nature. Several of these antibiotics, including penicillin and vancomycin, are employed to treat bacterial infections in humans and have been best-selling therapeutics for decades. Others might provide new platforms for the design of novel therapeutics to combat emerging antibiotic-resistant bacterial pathogens. PMID:19058272

  3. Language and national identity in Novo Cinema Galego

    Directory of Open Access Journals (Sweden)

    Brais ROMERO SUÁREZ

    2015-12-01

    Full Text Available The talk of town since its inception in 2010, the Cinema Novo Galego has been successful in all competitions and festivals that has been present. From the FIPRESCI prize in Cannes to the Best Emerging Director at Locarno, this new wave of cinema places Galicia in the world film stage. But does Novo Cinema Galego an accurate representation of Galicia? What's the role of Galicia in this movement?

  4. Identification of constrained peptides that bind to and preferentially inhibit the activity of the hepatitis C viral RNA-dependent RNA polymerase

    International Nuclear Information System (INIS)

    Amin, Anthony; Zaccardi, Joe; Mullen, Stanley; Olland, Stephane; Orlowski, Mark; Feld, Boris; Labonte, Patrick; Mak, Paul

    2003-01-01

    A class of disulfide constrained peptides containing a core motif FPWG was identified from a screen of phage displayed library using the HCV RNA-dependent RNA polymerase (NS5B) as a bait. Surface plasmon resonance studies showed that three highly purified synthetic constrained peptides bound to immobilized NS5B with estimated K d values ranging from 30 to 60 μM. In addition, these peptides inhibited the NS5B activity in vitro with IC 50 ranging from 6 to 48 μM, whereas in contrast they had no inhibitory effect on the enzymatic activities of calf thymus polymerase α, human polymerase β, RSV polymerase, and HIV reverse transcriptase in vitro. Two peptides demonstrated conformation-dependent inhibition since their synthetic linear versions were not inhibitory in the NS5B assay. A constrained peptide with the minimum core motif FPWG retained selective inhibition of NS5B activity with an IC 50 of 50 μM. Alanine scan analyses of a representative constrained peptide, FPWGNTW, indicated that residues F1 and W7 were critical for the inhibitory effect of this peptide, although residues P2 and N5 had some measurable inhibitory effect as well. Further analyses of the mechanism of inhibition indicated that these peptides inhibited the formation of preelongation complexes required for the elongation reaction. However, once the preelongation complex was formed, its activity was refractory to peptide inhibition. Furthermore, the constrained peptide FPWGNTW inhibited de novo initiated RNA synthesis by NS5B from a poly(rC) template. These data indicate that the peptides confer selective inhibition of NS5B activity by binding to the enzyme and perturbing an early step preceding the processive elongation step of RNA synthesis

  5. OptMAVEn--a new framework for the de novo design of antibody variable region models targeting specific antigen epitopes.

    Directory of Open Access Journals (Sweden)

    Tong Li

    Full Text Available Antibody-based therapeutics provides novel and efficacious treatments for a number of diseases. Traditional experimental approaches for designing therapeutic antibodies rely on raising antibodies against a target antigen in an immunized animal or directed evolution of antibodies with low affinity for the desired antigen. However, these methods remain time consuming, cannot target a specific epitope and do not lead to broad design principles informing other studies. Computational design methods can overcome some of these limitations by using biophysics models to rationally select antibody parts that maximize affinity for a target antigen epitope. This has been addressed to some extend by OptCDR for the design of complementary determining regions. Here, we extend this earlier contribution by addressing the de novo design of a model of the entire antibody variable region against a given antigen epitope while safeguarding for immunogenicity (Optimal Method for Antibody Variable region Engineering, OptMAVEn. OptMAVEn simulates in silico the in vivo steps of antibody generation and evolution, and is capable of capturing the critical structural features responsible for affinity maturation of antibodies. In addition, a humanization procedure was developed and incorporated into OptMAVEn to minimize the potential immunogenicity of the designed antibody models. As case studies, OptMAVEn was applied to design models of neutralizing antibodies targeting influenza hemagglutinin and HIV gp120. For both HA and gp120, novel computational antibody models with numerous interactions with their target epitopes were generated. The observed rates of mutations and types of amino acid changes during in silico affinity maturation are consistent with what has been observed during in vivo affinity maturation. The results demonstrate that OptMAVEn can efficiently generate diverse computational antibody models with both optimized binding affinity to antigens and reduced

  6. Propensity of a single-walled carbon nanotube-peptide to mimic a KK10 peptide in an HLA-TCR complex

    Science.gov (United States)

    Feng, Mei; Bell, David R.; Zhou, Ruhong

    2017-12-01

    The application of nanotechnology to improve disease diagnosis, treatment, monitoring, and prevention is the goal of nanomedicine. We report here a theoretical study of a functionalized single-walled carbon nanotube (CNT) mimic binding to a human leukocyte antigen-T cell receptor (HLA-TCR) immune complex as a first attempt of a potential nanomedicine for human immunodeficiency virus (HIV) vaccine development. The carbon nanotube was coated with three arginine residues to imitate the HIV type 1 immunodominant viral peptide KK10 (gag 263-272: KRWIILGLNK), named CNT-peptide hereafter. Through molecular dynamics simulations, we explore the CNT-peptide and KK10 binding to an important HLA-TCR complex. Our results suggest that the CNT-peptide and KK10 bind comparably to the HLA-TCR complex, but the CNT-peptide forms stronger interactions with the TCR. Desorption simulations highlight the innate flexibility of KK10 over the CNT-peptide, resulting in a slightly higher desorption energy required for KK10 over the CNT-peptide. Our findings indicate that the designed CNT-peptide mimic has favorable propensity to activate TCR pathways and should be further explored to understand therapeutic potential.

  7. Template-Directed Ligation of Peptides to Oligonucleotides

    Science.gov (United States)

    Bruick, Richard K.; Dawson, Philip E.; Kent, Stephen BH; Usman, Nassim; Joyce, Gerald F.

    1996-01-01

    Synthetic oligonucleotides and peptides have enjoyed a wide range of applications in both biology and chemistry. As a consequence, oligonucleotide-peptide conjugates have received considerable attention, most notably in the development of antisense constructs with improved pharmacological properties. In addition, oligonucleotide-peptide conjugates have been used as molecular tags, in the assembly of supramolecular arrays and in the construction of encoded combinatorial libraries. To make these chimeric molecules more accessible for a broad range of investigations, we sought to develop a facile method for joining fully deprotected oligonucleotides and peptides through a stable amide bond linkage. Furthermore, we wished to make this ligation reaction addressable, enabling one to direct the ligation of specific oligonucleotide and peptide components.To confer specificity and accelerate the rate of the reaction, the ligation process was designed to be dependent on the presence of a complementary oligonucleotide template.

  8. On the Origin of De Novo Genes in Arabidopsis thaliana Populations.

    Science.gov (United States)

    Li, Zi-Wen; Chen, Xi; Wu, Qiong; Hagmann, Jörg; Han, Ting-Shen; Zou, Yu-Pan; Ge, Song; Guo, Ya-Long

    2016-08-03

    De novo genes, which originate from ancestral nongenic sequences, are one of the most important sources of protein-coding genes. This origination process is crucial for the adaptation of organisms. However, how de novo genes arise and become fixed in a population or species remains largely unknown. Here, we identified 782 de novo genes from the model plant Arabidopsis thaliana and divided them into three types based on the availability of translational evidence, transcriptional evidence, and neither transcriptional nor translational evidence for their origin. Importantly, by integrating multiple types of omics data, including data from genomes, epigenomes, transcriptomes, and translatomes, we found that epigenetic modifications (DNA methylation and histone modification) play an important role in the origination process of de novo genes. Intriguingly, using the transcriptomes and methylomes from the same population of 84 accessions, we found that de novo genes that are transcribed in approximately half of the total accessions within the population are highly methylated, with lower levels of transcription than those transcribed at other frequencies within the population. We hypothesized that, during the origin of de novo gene alleles, those neutralized to low expression states via DNA methylation have relatively high probabilities of spreading and becoming fixed in a population. Our results highlight the process underlying the origin of de novo genes at the population level, as well as the importance of DNA methylation in this process. © The Author 2016. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

  9. Mechanistic studies of a cell-permeant peptide designed to enhance myosin light chain phosphorylation in polarized intestinal epithelia.

    Science.gov (United States)

    Almansour, Khaled; Taverner, Alistair; Eggleston, Ian M; Mrsny, Randall J

    2018-06-10

    Tight junction (TJ) structures restrict the movement of solutes between adjacent epithelial cells to maintain homeostatic conditions. A peptide, termed PIP 640, with the capacity to regulate the transient opening of intestinal TJ structures through an endogenous mechanism involving the induction of myosin light chain (MLC) phosphorylation at serine 19 (MLC-pS 19 ) has provided a promising new method to enhance the in vivo oral bioavailability of peptide therapeutics. PIP 640 is a decapeptide composed of all D-amino acids (rrdykvevrr-NH 2 ) that contains a central sequence designed to emulates a specific domain of C-kinase potentiated protein phosphatase-1 inhibitor-17 kDa (CPI-17) surrounded by positively-charged amino acids that provide a cell penetrating peptide (CPP)-like character. Here, we examine compositional requirements of PIP 640 with regard to its actions on MLC phosphorylation, its intracellular localization to TJ structures, and its interactions with MLC phosphatase (MLCP) elements that correlate with enhanced solute uptake. These studies showed that a glutamic acid and tyrosine within this peptide are critical for PIP 640 to retain its ability to increase MLC-pS 19 levels and enhance the permeability of macromolecular solutes of the size range of therapeutic peptides without detectable cytotoxicity. On the other hand, exchange of the aspartic acid for alanine and then arginine resulted in an increasingly greater bias toward protein phosphatase-1 (PP1) relative to MLCP inhibition, an outcome that resulted in increased paracellular permeability for solutes in the size range of therapeutic peptides, but with a significant increase in cytotoxicity. Together, these data further our understanding of the composition requirements of PIP 640 with respect to the desired goal of transiently altering the intestinal epithelial cell paracellular barrier properties through an endogenous mechanism, providing a novel approach to enhance the oral bioavailability of

  10. Neoglycolipids for Prolonging the Effects of Peptides

    DEFF Research Database (Denmark)

    van Witteloostuijn, Søren Blok; Mannerstedt, Karin Margareta Sophia; Wismann, Pernille

    2017-01-01

    Novel principles for optimizing the properties of peptide-based drugs are needed in order to leverage their full pharmacological potential. We present the design, synthesis, and evaluation of a library of neoglycolipidated glucagon-like peptide 1 (GLP-1) analogues, which are valuable drug...... was maintained or even improved compared to native GLP-1. This translated into pronounced in vivo efficacy in terms of both decreased acute food intake and improved glucose homeostasis in mice. Thus, we propose neoglycolipidation as a novel, general method for modulating the properties of therapeutic peptides...

  11. A novel chimeric peptide with antimicrobial activity.

    Science.gov (United States)

    Alaybeyoglu, Begum; Akbulut, Berna Sariyar; Ozkirimli, Elif

    2015-04-01

    Beta-lactamase-mediated bacterial drug resistance exacerbates the prognosis of infectious diseases, which are sometimes treated with co-administration of beta-lactam type antibiotics and beta-lactamase inhibitors. Antimicrobial peptides are promising broad-spectrum alternatives to conventional antibiotics in this era of evolving bacterial resistance. Peptides based on the Ala46-Tyr51 beta-hairpin loop of beta-lactamase inhibitory protein (BLIP) have been previously shown to inhibit beta-lactamase. Here, our goal was to modify this peptide for improved beta-lactamase inhibition and cellular uptake. Motivated by the cell-penetrating pVEC sequence, which includes a hydrophobic stretch at its N-terminus, our approach involved the addition of LLIIL residues to the inhibitory peptide N-terminus to facilitate uptake. Activity measurements of the peptide based on the 45-53 loop of BLIP for enhanced inhibition verified that the peptide was a competitive beta-lactamase inhibitor with a K(i) value of 58 μM. Incubation of beta-lactam-resistant cells with peptide decreased the number of viable cells, while it had no effect on beta-lactamase-free cells, indicating that this peptide had antimicrobial activity via beta-lactamase inhibition. To elucidate the molecular mechanism by which this peptide moves across the membrane, steered molecular dynamics simulations were carried out. We propose that addition of hydrophobic residues to the N-terminus of the peptide affords a promising strategy in the design of novel antimicrobial peptides not only against beta-lactamase but also for other intracellular targets. Copyright © 2015 European Peptide Society and John Wiley & Sons, Ltd.

  12. Self-assembling peptide-based building blocks in medical applications

    Energy Technology Data Exchange (ETDEWEB)

    Acar, Handan; Srivastava, Samanvaya; Chung, Eun Ji; Schnorenberg, Mathew R.; Barrett, John C.; LaBelle, James L.; Tirrell, Matthew

    2017-02-01

    Peptides and peptide-conjugates, comprising natural and synthetic building blocks, are an increasingly popular class of biomaterials. Self-assembled nanostructures based on peptides and peptide-conjugates offer advantages such as precise selectivity and multifunctionality that can address challenges and limitations in the clinic. In this review article, we discuss recent developments in the design and self-assembly of various nanomaterials based on peptides and peptide-conjugates for medical applications, and categorize them into two themes based on the driving forces of molecular self-assembly. First, we present the self-assembled nanostructures driven by the supramolecular interactions between the peptides, with or without the presence of conjugates. The studies where nanoassembly is driven by the interactions between the conjugates of peptide-conjugates are then presented. Particular emphasis is given to in vivo studies focusing on therapeutics, diagnostics, immune modulation and regenerative medicine. Finally, challenges and future perspectives are presented.

  13. Radiopharmaceutical development of radiolabelled peptides

    Energy Technology Data Exchange (ETDEWEB)

    Fani, Melpomeni; Maecke, Helmut R. [University Hospital Freiburg, Department of Nuclear Medicine, Freiburg (Germany)

    2012-02-15

    Receptor targeting with radiolabelled peptides has become very important in nuclear medicine and oncology in the past few years. The overexpression of many peptide receptors in numerous cancers, compared to their relatively low density in physiological organs, represents the molecular basis for in vivo imaging and targeted radionuclide therapy with radiolabelled peptide-based probes. The prototypes are analogs of somatostatin which are routinely used in the clinic. More recent developments include somatostatin analogs with a broader receptor subtype profile or with antagonistic properties. Many other peptide families such as bombesin, cholecystokinin/gastrin, glucagon-like peptide-1 (GLP-1)/exendin, arginine-glycine-aspartic acid (RGD) etc. have been explored during the last few years and quite a number of potential radiolabelled probes have been derived from them. On the other hand, a variety of strategies and optimized protocols for efficient labelling of peptides with clinically relevant radionuclides such as {sup 99m}Tc, M{sup 3+} radiometals ({sup 111}In, {sup 86/90}Y, {sup 177}Lu, {sup 67/68}Ga), {sup 64/67}Cu, {sup 18}F or radioisotopes of iodine have been developed. The labelling approaches include direct labelling, the use of bifunctional chelators or prosthetic groups. The choice of the labelling approach is driven by the nature and the chemical properties of the radionuclide. Additionally, chemical strategies, including modification of the amino acid sequence and introduction of linkers/spacers with different characteristics, have been explored for the improvement of the overall performance of the radiopeptides, e.g. metabolic stability and pharmacokinetics. Herein, we discuss the development of peptides as radiopharmaceuticals starting from the choice of the labelling method and the conditions to the design and optimization of the peptide probe, as well as some recent developments, focusing on a selected list of peptide families, including somatostatin

  14. Nanocellulose-based biosensors: design, preparation, and activity of peptide-linked cotton cellulose nanocrystals having fluorimetric and colorimetric elastase detection sensitivity

    Science.gov (United States)

    Nanocrystalline cellulose is an amphiphilic, high surface area material that can be easily functionalized and is biocom-patible and eco-friendly. It has been used singularly and in combination with other nanomaterials to optimize biosensor design. The attachment of peptides and proteins to nanocryst...

  15. A molecular dynamics and circular dichroism study of a novel synthetic antimicrobial peptide

    International Nuclear Information System (INIS)

    Rodina, N P; Yudenko, A N; Terterov, I N; Eliseev, I E

    2013-01-01

    Antimicrobial peptides are a class of small, usually positively charged amphiphilic peptides that are used by the innate immune system to combat bacterial infection in multicellular eukaryotes. Antimicrobial peptides are known for their broad-spectrum antimicrobial activity and thus can be used as a basis for a development of new antibiotics against multidrug-resistant bacteria. The most challengeous task on the way to a therapeutic use of antimicrobial peptides is a rational design of new peptides with enhanced activity and reduced toxicity. Here we report a molecular dynamics and circular dichroism study of a novel synthetic antimicrobial peptide D51. This peptide was earlier designed by Loose et al. using a linguistic model of natural antimicrobial peptides. Molecular dynamics simulation of the peptide folding in explicit solvent shows fast formation of two antiparallel beta strands connected by a beta-turn that is confirmed by circular dichroism measurements. Obtained from simulation amphipatic conformation of the peptide is analysed and possible mechanism of it's interaction with bacterial membranes together with ways to enhance it's antibacterial activity are suggested

  16. Glucagon infusion increases rate of purine synthesis de novo in rat liver

    International Nuclear Information System (INIS)

    Itakura, Mitsuo; Maeda, Noriaki; Tsuchiya, Masami; Yamashita, Kamejiro

    1987-01-01

    Based on the parallel increases of glucagon, the second peak of hepatic cAMP, and the rate of purine synthesis de novo in the prereplicative period in regenerating rate liver after a 70% hepatectomy, it was hypothesized that glucagon is responsible for the increased rate of purine synthesis de novo. To test this hypothesis, the effect of glucagon or dibutyryl cAMP infusion on the rate of purine synthesis de novo in rat liver was studied. Glucagon infusion but not insulin or glucose infusion increased the rate of purine synthesis de novo, which was assayed by [ 14 C]glycine or [ 14 C]formate incorporation, by 2.7- to 4.3-fold. Glucagon infusion increased cAMP concentrations by 4.9-fold and 5-phosphoribosyl-1-pyrophosphate concentrations by 1.5-fold in liver but did not change the specific activity of amidophosphoribosyltransferase or purine ribonucleotide concentrations. Dibutyryl cAMP infusion also increased the rate of purine synthesis de novo by 2.2- to 4.0-fold. Because glucagon infusion increased the rate of purine synthesis de novo in the presence of unchanged purine ribonucleotide concentrations, it is concluded that glucagon after infusion or in animals after a 70% hepatectomy is playing an anabolic role to increase the rate of purine synthesis de novo by increasing cAMP and 5-phosphoribosyl-1-pyrophosphate concentrations

  17. Targeting EphA2-Sam and Its Interactome: Design and Evaluation of Helical Peptides Enriched in Charged Residues.

    Science.gov (United States)

    Mercurio, Flavia A; Marasco, Daniela; Di Natale, Concetta; Pirone, Luciano; Costantini, Susan; Pedone, Emilia M; Leone, Marilisa

    2016-11-17

    The EphA2 receptor controls diverse physiological and pathological conditions and its levels are often upregulated in cancer. Targeting receptor overexpression, through modulation of endocytosis and consequent degradation, appears to be an appealing strategy for attacking tumor malignancy. In this scenario, the Sam domain of EphA2 plays a pivotal role because it is the site where protein regulators of endocytosis and stability are recruited by means of heterotypic Sam-Sam interactions. Because EphA2-Sam heterotypic complexes are largely based on electrostatic contacts, we have investigated the possibility of attacking these interactions with helical peptides enriched in charged residues. Several peptide sequences with high predicted helical propensities were designed, and detailed conformational analyses were conducted by diverse techniques including NMR, CD, and molecular dynamics (MD) simulations. Interaction studies were also performed by NMR, surface plasmon resonance (SPR), and microscale thermophoresis (MST) and led to the identification of two peptides capable of binding to the first Sam domain of Odin. These molecules represent early candidates for the generation of efficient Sam domain binders and antagonists of Sam-Sam interactions involving EphA2. © 2016 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  18. High frequencies of de novo CNVs in bipolar disorder and schizophrenia.

    LENUS (Irish Health Repository)

    Malhotra, Dheeraj

    2011-12-22

    While it is known that rare copy-number variants (CNVs) contribute to risk for some neuropsychiatric disorders, the role of CNVs in bipolar disorder is unclear. Here, we reasoned that a contribution of CNVs to mood disorders might be most evident for de novo mutations. We performed a genome-wide analysis of de novo CNVs in a cohort of 788 trios. Diagnoses of offspring included bipolar disorder (n = 185), schizophrenia (n = 177), and healthy controls (n = 426). Frequencies of de novo CNVs were significantly higher in bipolar disorder as compared with controls (OR = 4.8 [1.4,16.0], p = 0.009). De novo CNVs were particularly enriched among cases with an age at onset younger than 18 (OR = 6.3 [1.7,22.6], p = 0.006). We also confirmed a significant enrichment of de novo CNVs in schizophrenia (OR = 5.0 [1.5,16.8], p = 0.007). Our results suggest that rare spontaneous mutations are an important contributor to risk for bipolar disorder and other major neuropsychiatric diseases.

  19. Soybean Peptide as Additive on Yellow Feather Broiler Chicks ...

    African Journals Online (AJOL)

    GM soybean and sold on the Chinese market by China CF Duqing (Heze) Biotech Co. Ltd, Shandong Prov-ince, China. For the purpose of this study, the peptide was designated marketed soybean peptide (MSBP). In order to show its efficacy ...

  20. De Novo Human Cardiac Myocytes for Medical Research: Promises and Challenges

    Directory of Open Access Journals (Sweden)

    Veronique Hamel

    2017-01-01

    Full Text Available The advent of cellular reprogramming technology has revolutionized biomedical research. De novo human cardiac myocytes can now be obtained from direct reprogramming of somatic cells (such as fibroblasts, from induced pluripotent stem cells (iPSCs, which are reprogrammed from somatic cells, and from human embryonic stem cells (hESCs. Such de novo human cardiac myocytes hold great promise for in vitro disease modeling and drug screening and in vivo cell therapy of heart disease. Here, we review the technique advancements for generating de novo human cardiac myocytes. We also discuss several challenges for the use of such cells in research and regenerative medicine, such as the immature phenotype and heterogeneity of de novo cardiac myocytes obtained with existing protocols. We focus on the recent advancements in addressing such challenges.

  1. StraPep: a structure database of bioactive peptides

    Science.gov (United States)

    Wang, Jian; Yin, Tailang; Xiao, Xuwen; He, Dan; Xue, Zhidong; Jiang, Xinnong; Wang, Yan

    2018-01-01

    Abstract Bioactive peptides, with a variety of biological activities and wide distribution in nature, have attracted great research interest in biological and medical fields, especially in pharmaceutical industry. The structural information of bioactive peptide is important for the development of peptide-based drugs. Many databases have been developed cataloguing bioactive peptides. However, to our knowledge, database dedicated to collect all the bioactive peptides with known structure is not available yet. Thus, we developed StraPep, a structure database of bioactive peptides. StraPep holds 3791 bioactive peptide structures, which belong to 1312 unique bioactive peptide sequences. About 905 out of 1312 (68%) bioactive peptides in StraPep contain disulfide bonds, which is significantly higher than that (21%) of PDB. Interestingly, 150 out of 616 (24%) bioactive peptides with three or more disulfide bonds form a structural motif known as cystine knot, which confers considerable structural stability on proteins and is an attractive scaffold for drug design. Detailed information of each peptide, including the experimental structure, the location of disulfide bonds, secondary structure, classification, post-translational modification and so on, has been provided. A wide range of user-friendly tools, such as browsing, sequence and structure-based searching and so on, has been incorporated into StraPep. We hope that this database will be helpful for the research community. Database URL: http://isyslab.info/StraPep PMID:29688386

  2. Effect of sequence and stereochemistry reversal on p53 peptide mimicry.

    Directory of Open Access Journals (Sweden)

    Alessio Atzori

    Full Text Available Peptidomimetics effective in modulating protein-protein interactions and resistant to proteolysis have potential in therapeutic applications. An appealing yet underperforming peptidomimetic strategy is to employ D-amino acids and reversed sequences to mimic a lead peptide conformation, either separately or as the combined retro-inverso peptide. In this work, we examine the conformations of inverse, reverse and retro-inverso peptides of p53(15-29 using implicit solvent molecular dynamics simulation and circular dichroism spectroscopy. In order to obtain converged ensembles for the peptides, we find enhanced sampling is required via the replica exchange molecular dynamics method. From these replica exchange simulations, the D-peptide analogues of p53(15-29 result in a predominantly left-handed helical conformation. When the parent sequence is reversed sequence as either the L-peptide and D-peptide, these peptides display a greater helical propensity, feature reflected by NMR and CD studies in TFE/water solvent. The simulations also indicate that, while approximately similar orientations of the side-chains are possible by the peptide analogues, their ability to mimic the parent peptide is severely compromised by backbone orientation (for D-amino acids and side-chain orientation (for reversed sequences. A retro-inverso peptide is disadvantaged as a mimic in both aspects, and further chemical modification is required to enable this concept to be used fruitfully in peptidomimetic design. The replica exchange molecular simulation approach adopted here, with its ability to provide detailed conformational insights into modified peptides, has potential as a tool to guide structure-based design of new improved peptidomimetics.

  3. Peptides and Anti-peptide Antibodies for Small and Medium Scale Peptide and Anti-peptide Affinity Microarrays: Antigenic Peptide Selection, Immobilization, and Processing.

    Science.gov (United States)

    Zhang, Fan; Briones, Andrea; Soloviev, Mikhail

    2016-01-01

    This chapter describes the principles of selection of antigenic peptides for the development of anti-peptide antibodies for use in microarray-based multiplex affinity assays and also with mass-spectrometry detection. The methods described here are mostly applicable to small to medium scale arrays. Although the same principles of peptide selection would be suitable for larger scale arrays (with 100+ features) the actual informatics software and printing methods may well be different. Because of the sheer number of proteins/peptides to be processed and analyzed dedicated software capable of processing all the proteins and an enterprise level array robotics may be necessary for larger scale efforts. This report aims to provide practical advice to those who develop or use arrays with up to ~100 different peptide or protein features.

  4. Melhoramento do cafeeiro: IV - Café Mundo Novo

    Directory of Open Access Journals (Sweden)

    A. Carvalho

    1952-06-01

    Full Text Available Em um conjunto de cafeeiros existentes em Mundo Novo, hoje Urupês, na região Araraquarense do Estado de São Paulo, foram feitas seleções de vários cafeeiros baseando-se no seu aspecto vegetativo, na produção existente na época da seleção e na provável produção do ano seguinte. Estudou-se a origem da plantação inicial desse café, tanto em Urupês como em Jaú, chegando-se à conclusão de que é provavelmente originário desta última localidade. Progênies do café "Mundo Novo", anteriormente conhecido por "Sumatra" e derivado de plantas selecionadas em Urupês e Jaú, acham-se em estudo em seis localidades do Estado : Campinas, Ribeirão Prêto, Pindorama, Mococa, Jaú e Monte Alegre do Sul. No presente trabalho são apenas aproveitados dados referentes à variabilidade morfológica e característicos da produção das progênies dos primeiros cafeeiros selecionados em Urupês e estudados em Campinas, Jaú, Pindorama e Mococa. Em tôdas as localidades, observou-se variação nos caracteres morfológicos das progênies, verificando-se a ocorrência de plantas quase improdutivas. A maioria das progênies, no entanto, se caracteriza por acentuado vigor vegetativo. Foram estudadas as produções totais das progénies e das plantas, no período 1946-1951, notando-se que algumas progénies se salientaram pela elevada produção em tôdas as localidades. Os tipos de sementes "moca", "concha" e "chato" foram determinados em amostras de tôdas as plantas, por um período de três anos, notando-se que a variação ocorrida é da mesma ordem que a encontrada em outros cafeeiros em seleção. Procurou-se eliminar, pela seleção, cafeeiros com elevada produção de frutos sem sementes em uma ou duas lojas, característico êsse que parece ser hereditário. Os resultados obtidos de cruzamento entre os melhores cafeeiros "Mundo Novo" de Campinas e plantas da variedade murta, indicaram que esses cafeeiros são do tipo bourbon. Provavelmente

  5. Conformational Restriction of Peptides Using Dithiol Bis-Alkylation.

    Science.gov (United States)

    Peraro, L; Siegert, T R; Kritzer, J A

    2016-01-01

    Macrocyclic peptides are highly promising as inhibitors of protein-protein interactions. While many bond-forming reactions can be used to make cyclic peptides, most have limitations that make this chemical space challenging to access. Recently, a variety of cysteine alkylation reactions have been used in rational design and library approaches for cyclic peptide discovery and development. We and others have found that this chemistry is versatile and robust enough to produce a large variety of conformationally constrained cyclic peptides. In this chapter, we describe applications, methods, mechanistic insights, and troubleshooting for dithiol bis-alkylation reactions for the production of cyclic peptides. This method for efficient solution-phase macrocyclization is highly useful for the rapid production and screening of loop-based inhibitors of protein-protein interactions. © 2016 Elsevier Inc. All rights reserved.

  6. Structure-Based Design of Peptidic Inhibitors of the Interaction between CC Chemokine Ligand 5 (CCL5) and Human Neutrophil Peptides 1 (HNP1)

    NARCIS (Netherlands)

    Wichapong, Kanin; Alard, Jean-Eric; Ortega-Gomez, Almudena; Weber, Christian; Hackeng, Tilman M.; Soehnlein, Oliver; Nicolaes, Gerry A. F.

    2016-01-01

    Protein-protein interactions (PPIs) are receiving increasing interest, much sparked by the realization that they represent druggable targets. Recently, we successfully developed a peptidic inhibitor, RRYGTSKYQ ("SKY" peptide), that shows high potential in vitro and in vivo to interrupt a PPI between

  7. De Novo Collapsing Glomerulopathy in a Renal Allograft Recipient

    Directory of Open Access Journals (Sweden)

    Kanodia K

    2008-01-01

    Full Text Available Collapsing glomerulopathy (CG, characterized histologically by segmental/global glomerular capillary collapse, podocyte hypertrophy and hypercellularity and tubulo-interstitial injury; is characterized clinically by massive proteinuria and rapid progressive renal failure. CG is known to recur in renal allograft and rarely de novo. We report de novo CG 3 years post-transplant in a patient who received renal allograft from haplo-identical type donor.

  8. Design and characterization of hirulogs: A novel class of bivalent peptide inhibitors of thrombin

    Energy Technology Data Exchange (ETDEWEB)

    Maraganore, J.M.; Bourdon, P.; Jablonski, J.; Ramachandran, K.L. (Biogen, Inc., Cambridge, MA (USA)); Fenton, J.W. II (New York State Department of Health, Albany (USA))

    1990-07-31

    A novel class of synthetic peptides has been designed that inhibit the thrombin catalytic site and exhibit specificity for the anion-binding exosite (ABE) of {alpha}-thrombin. These peptides, called hirulogs, consist of (i) an active-site specificity sequence with a restricted Arg-Pro scissile bond, (ii) a polymeric linker of glycyl residues from 6 to 18 {angstrom} in length, and (iii) an ABE recognition sequence such as that in the hirudin C-terminus. Hirulog-1 ((D-Phe)-Pro-Arg-Pro-(Gly){sub 4}-Asn-Gly-Asp-Phe-Glu-Glu-Ile-Pro-Glu-Tyr-Leu) inhibits the thrombin-catalyzed hydrolysis of a tripeptide p-nitroanilide substrate with K{sub i} = 2.3 nM. In contrast, the synthetic C-terminal hirudin peptide S-Hir{sub 53-64}, which binds to the thrombin ABE, blocked the fibrinogen clotting activity of the enzyme with K{sub i} = 144 nM but failed to inhibit the hydrolysis of p-nitroanilide substrates at concentrations as high as 1 mM. Hirulog-1, but not S-Hir{sub 53-64}, was found to inhibit the incorporation of ({sup 14}C)diisopropyl fluorophosphate in thrombin. Hirulog-1 appears specific for thrombin as it lacks inhibitory activities toward human factor Xa, human plasmin, and bovine trypsin at inhibitor:enzyme concentrations 3 orders of magnitude higher than those required to inhibit thrombin. The optimal inhibitory activity of hirulog-1 depends upon all three components of its structure. Comparison of anticoagulant activities of hirulog-1, hirudin, and S-Hir{sub 53-64} showed that the synthetic hirulog-1 is 2-fold more potent than hirudin and 100-fold more active than S-Hir{sub 53-64} in increasing the activated partial thromboplastin time of normal human plasma.

  9. Purine biosynthesis de novo by lymphocytes in gout

    International Nuclear Information System (INIS)

    Kamoun, P.; Chanard, J.; Brami, M.; Funck-Brentano, J.L.

    1978-01-01

    A method of measurement in vitro of purine biosynthesis de novo in human circulating blood lymphocytes is proposed. The rate of early reactions of purine biosynthesis de novo was determined by the incorporation of [ 14 C]formate into N-formyl glycinamide ribonucleotide when the subsequent reactions of the metabolic pathway were completely inhibited by the antibiotic azaserine. Synthesis of 14 C-labelled N-formyl glycinamide ribonucleotide by lymphocytes was measured in healthy control subjects and patients with primary gout or hyperuricaemia secondary to renal failure, with or without allopurinol therapy. The average synthesis was higher in gouty patients without therapy than in control subjects, but the values contained overlap the normal range. In secondary hyperuricaemia the synthesis was at same value as in control subjects. These results are in agreement with the inconstant acceleration of purine biosynthesis de novo in gouty patients as seen by others with measurement of [ 14 C]glycine incorporation into urinary uric acid. (author)

  10. Studies on lactoferricin-derived Escherichia coli membrane-active peptides reveal differences in the mechanism of N-acylated versus nonacylated peptides.

    Science.gov (United States)

    Zweytick, Dagmar; Deutsch, Günter; Andrä, Jörg; Blondelle, Sylvie E; Vollmer, Ekkehard; Jerala, Roman; Lohner, Karl

    2011-06-17

    To improve the low antimicrobial activity of LF11, an 11-mer peptide derived from human lactoferricin, mutant sequences were designed based on the defined structure of LF11 in the lipidic environment. Thus, deletion of noncharged polar residues and strengthening of the hydrophobic N-terminal part upon adding a bulky hydrophobic amino acid or N-acylation resulted in enhanced antimicrobial activity against Escherichia coli, which correlated with the peptides' degree of perturbation of bacterial membrane mimics. Nonacylated and N-acylated peptides exhibited different effects at a molecular level. Nonacylated peptides induced segregation of peptide-enriched and peptide-poor lipid domains in negatively charged bilayers, although N-acylated peptides formed small heterogeneous domains resulting in a higher degree of packing defects. Additionally, only N-acylated peptides perturbed the lateral packing of neutral lipids and exhibited increased permeability of E. coli lipid vesicles. The latter did not correlate with the extent of improvement of the antimicrobial activity, which could be explained by the fact that elevated binding of N-acylated peptides to lipopolysaccharides of the outer membrane of gram-negative bacteria seems to counteract the elevated membrane permeabilization, reflected in the respective minimal inhibitory concentration for E. coli. The antimicrobial activity of the peptides correlated with an increase of membrane curvature stress and hence bilayer instability. Transmission electron microscopy revealed that only the N-acylated peptides induced tubular protrusions from the outer membrane, whereas all peptides caused detachment of the outer and inner membrane of E. coli bacteria. Viability tests demonstrated that these bacteria were dead before onset of visible cell lysis.

  11. Studies on Lactoferricin-derived Escherichia coli Membrane-active Peptides Reveal Differences in the Mechanism of N-Acylated Versus Nonacylated Peptides*

    Science.gov (United States)

    Zweytick, Dagmar; Deutsch, Günter; Andrä, Jörg; Blondelle, Sylvie E.; Vollmer, Ekkehard; Jerala, Roman; Lohner, Karl

    2011-01-01

    To improve the low antimicrobial activity of LF11, an 11-mer peptide derived from human lactoferricin, mutant sequences were designed based on the defined structure of LF11 in the lipidic environment. Thus, deletion of noncharged polar residues and strengthening of the hydrophobic N-terminal part upon adding a bulky hydrophobic amino acid or N-acylation resulted in enhanced antimicrobial activity against Escherichia coli, which correlated with the peptides' degree of perturbation of bacterial membrane mimics. Nonacylated and N-acylated peptides exhibited different effects at a molecular level. Nonacylated peptides induced segregation of peptide-enriched and peptide-poor lipid domains in negatively charged bilayers, although N-acylated peptides formed small heterogeneous domains resulting in a higher degree of packing defects. Additionally, only N-acylated peptides perturbed the lateral packing of neutral lipids and exhibited increased permeability of E. coli lipid vesicles. The latter did not correlate with the extent of improvement of the antimicrobial activity, which could be explained by the fact that elevated binding of N-acylated peptides to lipopolysaccharides of the outer membrane of Gram-negative bacteria seems to counteract the elevated membrane permeabilization, reflected in the respective minimal inhibitory concentration for E. coli. The antimicrobial activity of the peptides correlated with an increase of membrane curvature stress and hence bilayer instability. Transmission electron microscopy revealed that only the N-acylated peptides induced tubular protrusions from the outer membrane, whereas all peptides caused detachment of the outer and inner membrane of E. coli bacteria. Viability tests demonstrated that these bacteria were dead before onset of visible cell lysis. PMID:21515687

  12. De novo giant A2 aneurysm following anterior communicating artery occlusion.

    Science.gov (United States)

    Ibrahim, Tarik F; Hafez, Ahmad; Andrade-Barazarte, Hugo; Raj, Rahul; Niemela, Mika; Lehto, Hanna; Numminen, Jussi; Jarvelainen, Juha; Hernesniemi, Juha

    2015-01-01

    De novo intracranial aneurysms are reported to occur with varying incidence after intracranial aneurysm treatment. They are purported to be observed, however, with increased incidence after Hunterian ligation; particularly in cases of carotid artery occlusion for giant or complex aneurysms deemed unclippable. We report a case of right-sided de novo giant A2 aneurysm 6 years after an anterior communicating artery (ACoA) aneurysm clipping. We believe this de novo aneurysm developed in part due to patient-specific risk factors but also a significant change in cerebral hemodynamics. The ACoA became occluded after surgery that likely altered the cerebral hemodynamics and contributed to the de novo aneurysm. We believe this to be the first reported case of a giant de novo aneurysm in this location. Following parent vessel occlusion (mostly of the carotid artery), there are no reports of any de novo aneurysms in the pericallosal arteries let alone a giant one. The patient had a dominant right A1 and the sudden increase in A2 blood flow likely resulted in increased wall shear stress, particularly in the medial wall of the A2 where the aneurysm occurred 2 mm distal to the A1-2 junction. ACoA preservation is a key element of aneurysm surgery in this location. Suspected occlusion of this vessel may warrant closer radiographic follow-up in patients with other risk factors for aneurysm development.

  13. Peptides, polypeptides and peptide-polymer hybrids as nucleic acid carriers.

    Science.gov (United States)

    Ahmed, Marya

    2017-10-24

    Cell penetrating peptides (CPPs), and protein transduction domains (PTDs) of viruses and other natural proteins serve as a template for the development of efficient peptide based gene delivery vectors. PTDs are sequences of acidic or basic amphipathic amino acids, with superior membrane trespassing efficacies. Gene delivery vectors derived from these natural, cationic and cationic amphipathic peptides, however, offer little flexibility in tailoring the physicochemical properties of single chain peptide based systems. Owing to significant advances in the field of peptide chemistry, synthetic mimics of natural peptides are often prepared and have been evaluated for their gene expression, as a function of amino acid functionalities, architecture and net cationic content of peptide chains. Moreover, chimeric single polypeptide chains are prepared by a combination of multiple small natural or synthetic peptides, which imparts distinct physiological properties to peptide based gene delivery therapeutics. In order to obtain multivalency and improve the gene delivery efficacies of low molecular weight cationic peptides, bioactive peptides are often incorporated into a polymeric architecture to obtain novel 'polymer-peptide hybrids' with improved gene delivery efficacies. Peptide modified polymers prepared by physical or chemical modifications exhibit enhanced endosomal escape, stimuli responsive degradation and targeting efficacies, as a function of physicochemical and biological activities of peptides attached onto a polymeric scaffold. The focus of this review is to provide comprehensive and step-wise progress in major natural and synthetic peptides, chimeric polypeptides, and peptide-polymer hybrids for nucleic acid delivery applications.

  14. A Proteomic Workflow Using High-Throughput De Novo Sequencing Towards Complementation of Genome Information for Improved Comparative Crop Science.

    Science.gov (United States)

    Turetschek, Reinhard; Lyon, David; Desalegn, Getinet; Kaul, Hans-Peter; Wienkoop, Stefanie

    2016-01-01

    The proteomic study of non-model organisms, such as many crop plants, is challenging due to the lack of comprehensive genome information. Changing environmental conditions require the study and selection of adapted cultivars. Mutations, inherent to cultivars, hamper protein identification and thus considerably complicate the qualitative and quantitative comparison in large-scale systems biology approaches. With this workflow, cultivar-specific mutations are detected from high-throughput comparative MS analyses, by extracting sequence polymorphisms with de novo sequencing. Stringent criteria are suggested to filter for confidential mutations. Subsequently, these polymorphisms complement the initially used database, which is ready to use with any preferred database search algorithm. In our example, we thereby identified 26 specific mutations in two cultivars of Pisum sativum and achieved an increased number (17 %) of peptide spectrum matches.

  15. Serum stabilities of short tryptophan- and arginine-rich antimicrobial peptide analogs.

    Directory of Open Access Journals (Sweden)

    Leonard T Nguyen

    2010-09-01

    Full Text Available Several short antimicrobial peptides that are rich in tryptophan and arginine residues were designed with a series of simple modifications such as end capping and cyclization. The two sets of hexapeptides are based on the Trp- and Arg-rich primary sequences from the "antimicrobial centre" of bovine lactoferricin as well as an antimicrobial sequence obtained through the screening of a hexapeptide combinatorial library.HPLC, mass spectrometry and antimicrobial assays were carried out to explore the consequences of the modifications on the serum stability and microbicidal activity of the peptides. The results show that C-terminal amidation increases the antimicrobial activity but that it makes little difference to its proteolytic degradation in human serum. On the other hand, N-terminal acetylation decreases the peptide activities but significantly increases their protease resistance. Peptide cyclization of the hexameric peptides was found to be highly effective for both serum stability and antimicrobial activity. However the two cyclization strategies employed have different effects, with disulfide cyclization resulting in more active peptides while backbone cyclization results in more proteolytically stable peptides. However, the benefit of backbone cyclization did not extend to longer 11-mer peptides derived from the same region of lactoferricin. Mass spectrometry data support the serum stability assay results and allowed us to determine preferred proteolysis sites in the peptides. Furthermore, isothermal titration calorimetry experiments showed that the peptides all had weak interactions with albumin, the most abundant protein in human serum.Taken together, the results provide insight into the behavior of the peptides in human serum and will therefore aid in advancing antimicrobial peptide design towards systemic applications.

  16. Structural basis for precursor protein-directed ribosomal peptide macrocyclization

    Science.gov (United States)

    Li, Kunhua; Condurso, Heather L.; Li, Gengnan; Ding, Yousong; Bruner, Steven D.

    2016-01-01

    Macrocyclization is a common feature of natural product biosynthetic pathways including the diverse family of ribosomal peptides. Microviridins are architecturally complex cyanobacterial ribosomal peptides whose members target proteases with potent reversible inhibition. The product structure is constructed by three macrocyclizations catalyzed sequentially by two members of the ATP-grasp family, a unique strategy for ribosomal peptide macrocyclization. Here, we describe the detailed structural basis for the enzyme-catalyzed macrocyclizations in the microviridin J pathway of Microcystis aeruginosa. The macrocyclases, MdnC and MdnB, interact with a conserved α-helix of the precursor peptide using a novel precursor peptide recognition mechanism. The results provide insight into the unique protein/protein interactions key to the chemistry, suggest an origin of the natural combinatorial synthesis of microviridin peptides and provide a framework for future engineering efforts to generate designed compounds. PMID:27669417

  17. Structural basis for precursor protein-directed ribosomal peptide macrocyclization.

    Science.gov (United States)

    Li, Kunhua; Condurso, Heather L; Li, Gengnan; Ding, Yousong; Bruner, Steven D

    2016-11-01

    Macrocyclization is a common feature of natural product biosynthetic pathways including the diverse family of ribosomal peptides. Microviridins are architecturally complex cyanobacterial ribosomal peptides that target proteases with potent reversible inhibition. The product structure is constructed via three macrocyclizations catalyzed sequentially by two members of the ATP-grasp family, a unique strategy for ribosomal peptide macrocyclization. Here we describe in detail the structural basis for the enzyme-catalyzed macrocyclizations in the microviridin J pathway of Microcystis aeruginosa. The macrocyclases MdnC and MdnB interact with a conserved α-helix of the precursor peptide using a novel precursor-peptide recognition mechanism. The results provide insight into the unique protein-protein interactions that are key to the chemistry, suggest an origin for the natural combinatorial synthesis of microviridin peptides, and provide a framework for future engineering efforts to generate designed compounds.

  18. Computational Design of Biomimetic Phosphate Scavengers

    DEFF Research Database (Denmark)

    Gruber, Mathias Felix; Wood, Elizabeth Baker; Truelsen, Sigurd Friis

    2015-01-01

    Phosphorus has long been the target of much research, but in recent years the focus has shifted from being limited only to reducing its detrimental environmental impact, to also looking at how it is linked to the global food security. Therefore, the interest in finding novel techniques for phosph......Phosphorus has long been the target of much research, but in recent years the focus has shifted from being limited only to reducing its detrimental environmental impact, to also looking at how it is linked to the global food security. Therefore, the interest in finding novel techniques...... for phosphorus recovery, as well as improving existing techniques, has increased. In this study we apply a hybrid simulation approach of molecular dynamics and quantum mechanics to investigate the binding modes of phosphate anions by a small intrinsically disordered peptide. Our results confirm...... phosphate could be the starting point of new novel technological approaches toward phosphorus recovery, and they represent an excellent model system for investigating the nature and dynamics of functional de novo designed intrinsically disordered proteins....

  19. Direct Visualization of De novo Lipogenesis in Single Living Cells

    Science.gov (United States)

    Li, Junjie; Cheng, Ji-Xin

    2014-10-01

    Increased de novo lipogenesis is being increasingly recognized as a hallmark of cancer. Despite recent advances in fluorescence microscopy, autoradiography and mass spectrometry, direct observation of de novo lipogenesis in living systems remains to be challenging. Here, by coupling stimulated Raman scattering (SRS) microscopy with isotope labeled glucose, we were able to trace the dynamic metabolism of glucose in single living cells with high spatial-temporal resolution. As the first direct visualization, we observed that glucose was largely utilized for lipid synthesis in pancreatic cancer cells, which occurs at a much lower rate in immortalized normal pancreatic epithelial cells. By inhibition of glycolysis and fatty acid synthase (FAS), the key enzyme for fatty acid synthesis, we confirmed the deuterium labeled lipids in cancer cells were from de novo lipid synthesis. Interestingly, we also found that prostate cancer cells exhibit relatively lower level of de novo lipogenesis, but higher fatty acid uptake compared to pancreatic cancer cells. Together, our results demonstrate a valuable tool to study dynamic lipid metabolism in cancer and other disorders.

  20. Integrating valve-inspired design features into poly(ethylene glycol) hydrogel scaffolds for heart valve tissue engineering.

    Science.gov (United States)

    Zhang, Xing; Xu, Bin; Puperi, Daniel S; Yonezawa, Aline L; Wu, Yan; Tseng, Hubert; Cuchiara, Maude L; West, Jennifer L; Grande-Allen, K Jane

    2015-03-01

    The development of advanced scaffolds that recapitulate the anisotropic mechanical behavior and biological functions of the extracellular matrix in leaflets would be transformative for heart valve tissue engineering. In this study, anisotropic mechanical properties were established in poly(ethylene glycol) (PEG) hydrogels by crosslinking stripes of 3.4 kDa PEG diacrylate (PEGDA) within 20 kDa PEGDA base hydrogels using a photolithographic patterning method. Varying the stripe width and spacing resulted in a tensile elastic modulus parallel to the stripes that was 4.1-6.8 times greater than that in the perpendicular direction, comparable to the degree of anisotropy between the circumferential and radial orientations in native valve leaflets. Biomimetic PEG-peptide hydrogels were prepared by tethering the cell-adhesive peptide RGDS and incorporating the collagenase-degradable peptide PQ (GGGPQG↓IWGQGK) into the polymer network. The specific amounts of RGDS and PEG-PQ within the resulting hydrogels influenced the elongation, de novo extracellular matrix deposition and hydrogel degradation behavior of encapsulated valvular interstitial cells (VICs). In addition, the morphology and activation of VICs grown atop PEG hydrogels could be modulated by controlling the concentration or micro-patterning profile of PEG-RGDS. These results are promising for the fabrication of PEG-based hydrogels using anatomically and biologically inspired scaffold design features for heart valve tissue engineering. Copyright © 2014 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

  1. Response monitoring in de novo patients with Parkinson's disease.

    Directory of Open Access Journals (Sweden)

    Rita Willemssen

    Full Text Available BACKGROUND: Parkinson's disease (PD is accompanied by dysfunctions in a variety of cognitive processes. One of these is error processing, which depends upon phasic decreases of medial prefrontal dopaminergic activity. Until now, there is no study evaluating these processes in newly diagnosed, untreated patients with PD ("de novo PD". METHODOLOGY/PRINCIPAL FINDINGS: Here we report large changes in performance monitoring processes using event-related potentials (ERPs in de novo PD-patients. The results suggest that increases in medial frontal dopaminergic activity after an error (Ne are decreased, relative to age-matched controls. In contrast, neurophysiological processes reflecting general motor response monitoring (Nc are enhanced in de novo patients. CONCLUSIONS/SIGNIFICANCE: It may be hypothesized that the Nc-increase is at costs of dopaminergic activity after an error; on a functional level errors may not always be detected and correct responses sometimes be misinterpreted as errors. This pattern differs from studies examining patients with a longer history of PD and may reflect compensatory processes, frequently occurring in pre-manifest stages of PD. From a clinical point of view the clearly attenuated Ne in the de novo PD patients may prove a useful additional tool for the early diagnosis of basal ganglia dysfunction in PD.

  2. Precise detection of de novo single nucleotide variants in human genomes.

    Science.gov (United States)

    Gómez-Romero, Laura; Palacios-Flores, Kim; Reyes, José; García, Delfino; Boege, Margareta; Dávila, Guillermo; Flores, Margarita; Schatz, Michael C; Palacios, Rafael

    2018-05-07

    The precise determination of de novo genetic variants has enormous implications across different fields of biology and medicine, particularly personalized medicine. Currently, de novo variations are identified by mapping sample reads from a parent-offspring trio to a reference genome, allowing for a certain degree of differences. While widely used, this approach often introduces false-positive (FP) results due to misaligned reads and mischaracterized sequencing errors. In a previous study, we developed an alternative approach to accurately identify single nucleotide variants (SNVs) using only perfect matches. However, this approach could be applied only to haploid regions of the genome and was computationally intensive. In this study, we present a unique approach, coverage-based single nucleotide variant identification (COBASI), which allows the exploration of the entire genome using second-generation short sequence reads without extensive computing requirements. COBASI identifies SNVs using changes in coverage of exactly matching unique substrings, and is particularly suited for pinpointing de novo SNVs. Unlike other approaches that require population frequencies across hundreds of samples to filter out any methodological biases, COBASI can be applied to detect de novo SNVs within isolated families. We demonstrate this capability through extensive simulation studies and by studying a parent-offspring trio we sequenced using short reads. Experimental validation of all 58 candidate de novo SNVs and a selection of non-de novo SNVs found in the trio confirmed zero FP calls. COBASI is available as open source at https://github.com/Laura-Gomez/COBASI for any researcher to use. Copyright © 2018 the Author(s). Published by PNAS.

  3. Intelligent "Peptide-Gathering Mechanical Arm" Tames Wild "Trojan-Horse" Peptides for the Controlled Delivery of Cancer Nanotherapeutics.

    Science.gov (United States)

    Shi, Nian-Qiu; Li, Yan; Zhang, Yong; Shen, Nan; Qi, Ling; Wang, Shu-Ran; Qi, Xian-Rong

    2017-12-06

    Cell-penetrating peptides (CPPs), also called "Trojan-Horse" peptides, have been used for facilitating intracellular delivery of numerous diverse cargoes and even nanocarriers. However, the lack of targeting specificity ("wildness" or nonselectivity) of CPP-nanocarriers remains an intractable challenge for many in vivo applications. In this work, we used an intelligent "peptide-gathering mechanical arm" (Int PMA) to curb CPPs' wildness and enhance the selectivity of R 9 -liposome-based cargo delivery for tumor targeting. The peptide NGR, serving as a cell-targeting peptide for anchoring, and peptide PLGLAG, serving as a substrate peptide for deanchoring, were embedded in the Int PMA motif. The Int PMA construct was designed to be sensitive to tumor microenvironmental stimuli, including aminopeptidase N (CD13) and matrix metalloproteinases (MMP-2/9). Moreover, Int PMA could be specifically recognized by tumor tissues via CD13-mediated anchoring and released for cell entry by MMP-2/9-mediated deanchoring. To test the Int PMA design, a series of experiments were conducted in vitro and in vivo. Functional conjugates Int PMA-R 9 -poly(ethylene glycol) (PEG) 2000 -distearoylphosphatidyl-ethanolamine (DSPE) and R 9 -PEG 2000 -DSPE were synthesized by Michael addition reaction and were characterized by thin-layer chromatography and matrix-assisted laser desorption ionization-time-of-flight mass spectrometry. The Int PMA-R 9 -modified doxorubicin-loaded liposomes (Int PMA-R 9 -Lip-DOX) exhibited a proper particle diameter (approximately 155 nm) with in vitro sustained release characteristics. Cleavage assay showed that Int PMA-R 9 peptide molecules could be cleaved by MMP-2/9 for completion of deanchoring. Flow cytometry and confocal microscopy studies indicated that Int PMA-R 9 -Lip-DOX can respond to both endogenous and exogenous stimuli in the presence/absence of excess MMP-2/9 and MMP-2/9 inhibitor (GM6001) and effectively function under competitive receptor

  4. Computational design and elaboration of a de novo heterotetrameric alpha-helical protein that selectively binds an emissive abiological (porphinato)zinc chromophore.

    Science.gov (United States)

    Fry, H Christopher; Lehmann, Andreas; Saven, Jeffery G; DeGrado, William F; Therien, Michael J

    2010-03-24

    The first example of a computationally de novo designed protein that binds an emissive abiological chromophore is presented, in which a sophisticated level of cofactor discrimination is pre-engineered. This heterotetrameric, C(2)-symmetric bundle, A(His):B(Thr), uniquely binds (5,15-di[(4-carboxymethyleneoxy)phenyl]porphinato)zinc [(DPP)Zn] via histidine coordination and complementary noncovalent interactions. The A(2)B(2) heterotetrameric protein reflects ligand-directed elements of both positive and negative design, including hydrogen bonds to second-shell ligands. Experimental support for the appropriate formulation of [(DPP)Zn:A(His):B(Thr)](2) is provided by UV/visible and circular dichroism spectroscopies, size exclusion chromatography, and analytical ultracentrifugation. Time-resolved transient absorption and fluorescence spectroscopic data reveal classic excited-state singlet and triplet PZn photophysics for the A(His):B(Thr):(DPP)Zn protein (k(fluorescence) = 4 x 10(8) s(-1); tau(triplet) = 5 ms). The A(2)B(2) apoprotein has immeasurably low binding affinities for related [porphinato]metal chromophores that include a (DPP)Fe(III) cofactor and the zinc metal ion hemin derivative [(PPIX)Zn], underscoring the exquisite active-site binding discrimination realized in this computationally designed protein. Importantly, elements of design in the A(His):B(Thr) protein ensure that interactions within the tetra-alpha-helical bundle are such that only the heterotetramer is stable in solution; corresponding homomeric bundles present unfavorable ligand-binding environments and thus preclude protein structural rearrangements that could lead to binding of (porphinato)iron cofactors.

  5. The Drosophila melanogaster PeptideAtlas facilitates the use of peptide data for improved fly proteomics and genome annotation

    Directory of Open Access Journals (Sweden)

    King Nichole L

    2009-02-01

    Full Text Available Abstract Background Crucial foundations of any quantitative systems biology experiment are correct genome and proteome annotations. Protein databases compiled from high quality empirical protein identifications that are in turn based on correct gene models increase the correctness, sensitivity, and quantitative accuracy of systems biology genome-scale experiments. Results In this manuscript, we present the Drosophila melanogaster PeptideAtlas, a fly proteomics and genomics resource of unsurpassed depth. Based on peptide mass spectrometry data collected in our laboratory the portal http://www.drosophila-peptideatlas.org allows querying fly protein data observed with respect to gene model confirmation and splice site verification as well as for the identification of proteotypic peptides suited for targeted proteomics studies. Additionally, the database provides consensus mass spectra for observed peptides along with qualitative and quantitative information about the number of observations of a particular peptide and the sample(s in which it was observed. Conclusion PeptideAtlas is an open access database for the Drosophila community that has several features and applications that support (1 reduction of the complexity inherently associated with performing targeted proteomic studies, (2 designing and accelerating shotgun proteomics experiments, (3 confirming or questioning gene models, and (4 adjusting gene models such that they are in line with observed Drosophila peptides. While the database consists of proteomic data it is not required that the user is a proteomics expert.

  6. Design of peptide substrates for nanosecond time-resolved fluorescence assays of proteases: 2,3-diazabicyclo[2.2.2]oct-2-ene as a noninvasive fluorophore.

    Science.gov (United States)

    Hennig, Andreas; Florea, Mara; Roth, Doris; Enderle, Thilo; Nau, Werner M

    2007-01-15

    Fluorescence protease assays were investigated with peptide substrates containing a 2,3-diazabicyclo[2.2.2]oct-2-ene-labeled asparagine (Dbo) as a fluorescent amino acid. The special characteristic of the fluorophore Dbo is its exceedingly long fluorescence lifetime (ca. 300 ns in water under air), which allows the use of nanosecond time-resolved fluorescence (Nano-TRF) detection to efficiently suppress shorter-lived background emission. In addition, the natural amino acids tryptophan and tyrosine can be employed as intramolecular fluorescence quenchers, which facilitates substrate design. Fourteen synthetic peptide substrates (composed of 2-19 amino acids) and five enzymes (trypsin, pepsin, carboxypeptidase A, leucine aminopeptidase, and chymotrypsin) were investigated and, in all 28 examined combinations, enzymatic activity was detected by monitoring the increase in steady state fluorescence with time and determining the reaction rates as kcat/Km values, which ranged from 0.2 to 80x10(6) M-1 min-1. The results suggest an excellent compatibility of the very small and hydrophilic fluorescent probe Dbo with solid-phase peptide synthesis and the investigated proteases. For all 14 peptides the fluorescence lifetimes before and after enzymatic cleavage were measured and Nano-TRF measurements were performed in 384-well microplates. The fluorescence lifetimes of the different peptides provide the basis for the rational design of Dbo-based fluorescent substrates for protease assays. Measurements in Nano-TRF mode revealed, in addition to efficient suppression of background fluorescence, an increased differentiation between cleaved and uncleaved substrate. The Dbo-based assays can be adapted for high-throughput screening.

  7. Tuning peptide self-assembly by an in-tether chiral center

    Science.gov (United States)

    Hu, Kuan; Xiong, Wei; Li, Hu; Zhang, Pei-Yu; Yin, Feng; Zhang, Qianling; Jiang, Fan; Li, Zigang

    2018-01-01

    The self-assembly of peptides into ordered nanostructures is important for understanding both peptide molecular interactions and nanotechnological applications. However, because of the complexity and various self-assembling pathways of peptide molecules, design of self-assembling helical peptides with high controllability and tunability is challenging. We report a new self-assembling mode that uses in-tether chiral center-induced helical peptides as a platform for tunable peptide self-assembly with good controllability. It was found that self-assembling behavior was governed by in-tether substitutional groups, where chirality determined the formation of helical structures and aromaticity provided the driving force for self-assembly. Both factors were essential for peptide self-assembly to occur. Experiments and theoretical calculations indicate long-range crystal-like packing in the self-assembly, which was stabilized by a synergy of interpeptide π-π and π-sulfur interactions and hydrogen bond networks. In addition, the self-assembled peptide nanomaterials were demonstrated to be promising candidate materials for applications in biocompatible electrochemical supercapacitors.

  8. Cathepsin-Mediated Cleavage of Peptides from Peptide Amphiphiles Leads to Enhanced Intracellular Peptide Accumulation

    Energy Technology Data Exchange (ETDEWEB)

    Acar, Handan [Institute; Department; Samaeekia, Ravand [Institute; Department; Schnorenberg, Mathew R. [Institute; Department; Medical; Sasmal, Dibyendu K. [Institute; Huang, Jun [Institute; Tirrell, Matthew V. [Institute; Institute; LaBelle, James L. [Department

    2017-08-24

    Peptides synthesized in the likeness of their native interaction domain(s) are natural choices to target protein protein interactions (PPIs) due to their fidelity of orthostatic contact points between binding partners. Despite therapeutic promise, intracellular delivery of biofunctional peptides at concentrations necessary for efficacy remains a formidable challenge. Peptide amphiphiles (PAs) provide a facile method of intracellular delivery and stabilization of bioactive peptides. PAs consisting of biofunctional peptide headgroups linked to hydrophobic alkyl lipid-like tails prevent peptide hydrolysis and proteolysis in circulation, and PA monomers are internalized via endocytosis. However, endocytotic sequestration and steric hindrance from the lipid tail are two major mechanisms that limit PA efficacy to target intracellular PPIs. To address these problems, we have constructed a PA platform consisting of cathepsin-B cleavable PAs in which a selective p53-based inhibitory peptide is cleaved from its lipid tail within endosomes, allowing for intracellular peptide accumulation and extracellular recycling of the lipid moiety. We monitor for cleavage and follow individual PA components in real time using a resonance energy transfer (FRET)-based tracking system. Using this platform, components in real time using a Forster we provide a better understanding and quantification of cellular internalization, trafficking, and endosomal cleavage of PAs and of the ultimate fates of each component.

  9. De novo mutations in synaptic transmission genes including DNM1 cause epileptic encephalopathies

    DEFF Research Database (Denmark)

    2014-01-01

    in five individuals and de novo mutations in GABBR2, FASN, and RYR3 in two individuals each. Unlike previous studies, this cohort is sufficiently large to show a significant excess of de novo mutations in epileptic encephalopathy probands compared to the general population using a likelihood analysis (p...... = 8.2 × 10(-4)), supporting a prominent role for de novo mutations in epileptic encephalopathies. We bring statistical evidence that mutations in DNM1 cause epileptic encephalopathy, find suggestive evidence for a role of three additional genes, and show that at least 12% of analyzed individuals have...... analyzed exome-sequencing data of 356 trios with the "classical" epileptic encephalopathies, infantile spasms and Lennox Gastaut syndrome, including 264 trios previously analyzed by the Epi4K/EPGP consortium. In this expanded cohort, we find 429 de novo mutations, including de novo mutations in DNM1...

  10. Interference with RUNX1/ETO Leukemogenic Function by Cell-Penetrating Peptides Targeting the NHR2 Oligomerization Domain

    Directory of Open Access Journals (Sweden)

    Yvonne Bartel

    2013-01-01

    Full Text Available The leukemia-associated fusion protein RUNX1/ETO is generated by the chromosomal translocation t(8;21 which appears in about 12% of all de novo acute myeloid leukemias (AMLs. Essential for the oncogenic potential of RUNX1/ETO is the oligomerization of the chimeric fusion protein through the nervy homology region 2 (NHR2 within ETO. In previous studies, we have shown that the intracellular expression of peptides containing the NHR2 domain inhibits RUNX1/ETO oligomerization, thereby preventing cell proliferation and inducing differentiation of RUNX1/ETO transformed cells. Here, we show that introduction of a recombinant TAT-NHR2 fusion polypeptide into the RUNX1/ETO growth-dependent myeloid cell line Kasumi-1 results in decreased cell proliferation and increased numbers of apoptotic cells. This effect was highly specific and mediated by binding the TAT-NHR2 peptide to ETO sequences, as TAT-polypeptides containing the oligomerization domain of BCR did not affect cell proliferation or apoptosis in Kasumi-1 cells. Thus, the selective interference with NHR2-mediated oligomerization by peptides represents a challenging but promising strategy for the inhibition of the leukemogenic potential of RUNX1/ETO in t(8;21-positive leukemia.

  11. Antimicrobial peptide GH12 suppresses cariogenic virulence factors of Streptococcus mutans

    Science.gov (United States)

    Wang, Yufei; Wang, Xiuqing; Jiang, Wentao; Wang, Kun; Luo, Junyuan; Li, Wei; Zhou, Xuedong; Zhang, Linglin

    2018-01-01

    ABSTRACT Cariogenic virulence factors of Streptococcus mutans include acidogenicity, aciduricity, and extracellular polysaccharides (EPS) synthesis. The de novo designed antimicrobial peptide GH12 has shown bactericidal effects on S. mutans, but its interaction with virulence and regulatory systems of S. mutans remains to be elucidated. The objectives were to investigate the effects of GH12 on virulence factors of S. mutans, and further explore the function mechanisms at enzymatic and transcriptional levels. To avoid decrease in bacterial viability, we limited GH12 to subinhibitory levels. We evaluated effects of GH12 on acidogenicity of S. mutans by pH drop, lactic acid measurement and lactate dehydrogenase (LDH) assay, on aciduricity through survival rate at pH 5.0 and F1F0-ATPase assay, and on EPS synthesis using quantitative measurement, morphology observation, vertical distribution analyses and biomass calculation. Afterwards, we conducted quantitative real-time PCR to acquire the expression profile of related genes. GH12 at 1/2 MIC (4 mg/L) inhibited acid production, survival rate, EPS synthesis, and biofilm formation. The enzymatic activity of LDH and F1F0-ATPase was inhibited, and ldh, gtfBCD, vicR, liaR, and comDE genes were significantly downregulated. In conclusion, GH12 inhibited virulence factors of S. mutans, through reducing the activity of related enzymes, downregulating virulence genes, and inactivating specific regulatory systems. PMID:29503706

  12. Defining the maize transcriptome de novo using deep RNA-Seq

    Energy Technology Data Exchange (ETDEWEB)

    Martin, Jeffrey; Gross, Stephen; Choi, Cindy; Zhang, Tao; Lindquist, Erika; Wei, Chia-Lin; Wang, Zhong

    2011-06-01

    De novo assembly of the transcriptome is crucial for functional genomics studies in bioenergy research, since many of the organisms lack high quality reference genomes. In a previous study we successfully de novo assembled simple eukaryote transcriptomes exclusively from short Illumina RNA-Seq reads [1]. However, extensive alternative splicing, present in most of the higher eukaryotes, poses a significant challenge for current short read assembly processes. Furthermore, the size of next-generation datasets, often large for plant genomes, presents an informatics challenge. To tackle these challenges we present a combined experimental and informatics strategy for de novo assembly in higher eukaryotes. Using maize as a test case, preliminary results suggest our approach can resolve transcript variants and improve gene annotations.

  13. Defining the maize transcriptome de novo using deep RNA-Seq

    Energy Technology Data Exchange (ETDEWEB)

    Martin, Jeffrey; Gross, Stephen; Choi, Cindy; Zhang, Tao; Lindquist, Erika; Wei, Chia-Lin; Wang, Zhong

    2011-06-02

    De novo assembly of the transcriptome is crucial for functional genomics studies in bioenergy research, since many of the organisms lack high quality reference genomes. In a previous study we successfully de novo assembled simple eukaryote transcriptomes exclusively from short Illumina RNA-Seq reads [1]. However, extensive alternative splicing, present in most of the higher eukaryotes, poses a significant challenge for current short read assembly processes. Furthermore, the size of next-generation datasets, often large for plant genomes, presents an informatics challenge. To tackle these challenges we present a combined experimental and informatics strategy for de novo assembly in higher eukaryotes. Using maize as a test case, preliminary results suggest our approach can resolve transcript variants and improve gene annotations.

  14. An Approach Towards Structure Based Antimicrobial Peptide Design for Use in Development of Transgenic Plants: A Strategy for Plant Disease Management.

    Science.gov (United States)

    Ilyas, Humaira; Datta, Aritreyee; Bhunia, Anirban

    2017-01-01

    Antimicrobial peptides (AMPs), also known as host defense peptides (HDPs), are ubiquitous and vital components of innate defense response that present themselves as potential candidates for drug design, and aim to control plant and animal diseases. Though their application for plant disease management has long been studied with natural AMPs, cytotoxicity and stability related shortcomings for the development of transgenic plants limit their usage. Newer technologies like molecular modelling, NMR spectroscopy and combinatorial chemistry allow screening for potent candidates and provide new avenues for the generation of rationally designed synthetic AMPs with multiple biological functions. Such AMPs can be used for the control of plant diseases that lead to huge yield losses of agriculturally important crop plants, via generation of transgenic plants. Such approaches have gained significant attention in the past decade as a consequence of increasing antibiotic resistance amongst plant pathogens, and the shortcomings of existing strategies that include environmental contamination and human/animal health hazards amongst others. This review summarizes the recent trends and approaches used for employing AMPs, emphasizing on designed/modified ones, and their applications toward agriculture and food technology. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  15. A readily applicable strategy to convert peptides to peptoid-based therapeutics.

    Directory of Open Access Journals (Sweden)

    Minyoung Park

    Full Text Available Incorporation of unnatural amino acids and peptidomimetic residues into therapeutic peptides is highly efficacious and commonly employed, but generally requires laborious trial-and-error approaches. Previously, we demonstrated that C20 peptide has the potential to be a potential antiviral agent. Herein we report our attempt to improve the biological properties of this peptide by introducing peptidomimetics. Through combined alanine, proline, and sarcosine scans coupled with a competitive fluorescence polarization assay developed for identifying antiviral peptides, we enabled to pinpoint peptoid-tolerant peptide residues within C20 peptide. The synergistic benefits of combining these (and other commonly employed methods could lead to a easily applicable strategy for designing and refining therapeutically-attractive peptidomimetics.

  16. Design and biological activity of β-sheet breaker peptide conjugates

    International Nuclear Information System (INIS)

    Rocha, Sandra; Cardoso, Isabel; Boerner, Hans; Pereira, Maria Carmo; Saraiva, Maria Joao; Coelho, Manuel

    2009-01-01

    The sequence LPFFD (iAβ 5 ) prevents amyloid-β peptide (Aβ) fibrillogenesis and neurotoxicity, hallmarks of Alzheimer's disease (AD), as previously demonstrated. In this study iAβ 5 was covalently linked to poly(ethylene glycol) (PEG) and the activity of conjugates was assessed and compared to the activity of the peptide alone by in vitro studies. The conjugates were characterized by MALDI-TOF. Competition binding assays established that conjugates retained the ability to bind Aβ with similar strength as iAβ 5 . Transmission electron microscopy analysis showed that iAβ 5 conjugates inhibited amyloid fibril formation, which is in agreement with binding properties observed for the conjugates towards Aβ. The conjugates were also able to prevent amyloid-induced cell death, as evaluated by activation of caspase 3. These results demonstrated that the biological activity of iAβ 5 is not affected by the pegylation process.

  17. What peptides these deltorphins be.

    Science.gov (United States)

    Lazarus, L H; Bryant, S D; Cooper, P S; Salvadori, S

    1999-02-01

    The deltorphins are a class of highly selective delta-opioid heptapeptides from the skin of the Amazonian frogs Phyllomedusa sauvagei and P. bicolor. The first of these fascinating peptides came to light in 1987 by cloning of the cDNA of from frog skins, while the other members of this family were identified either by cDNA or isolation of the peptides. The distinctive feature of deltorphins is the presence of a naturally occurring D-enantiomer at the second position in their common N-terminal sequence, Tyr-D-Xaa-Phe, comparable to dermorphin, which is the prototype of a group of mu-selective opioids from the same source. The D-amino acid and the anionic residues, either Glu or Asp, as well as their unique amino acid compositions are responsible for the remarkable biostability, high delta-receptor affinity, bioactivity and peptide conformation. This review summarizes a decade of research from many laboratories that defined which residues and substituents in the deltorphins interact with the delta-receptor and characterized pharmacological and physiological activities in vitro and in vivo. It begins with a historical description of the topic and presents general schema for the synthesis of peptide analogues of deltorphins A, B and C as a means to document the methods employed in producing a myriad of analogues. Structure activity studies of the peptides and their pharmacological activities in vitro are detailed in abundantly tabulated data. A brief compendium of the current level of knowledge of the delta-receptor assists the reader to appreciate the rationale for the design of these analogues. Discussion of the conformation of these peptides addresses how structure leads to further hypotheses regarding ligand receptor interaction. The review ends with a broad discussion of the potential applications of these peptides in clinical and therapeutic settings.

  18. Functional tuning of the catalytic residue pKa in a de novo designed esterase.

    Science.gov (United States)

    Hiebler, Katharina; Lengyel, Zsófia; Castañeda, Carlos A; Makhlynets, Olga V

    2017-09-01

    AlleyCatE is a de novo designed esterase that can be allosterically regulated by calcium ions. This artificial enzyme has been shown to hydrolyze p-nitrophenyl acetate (pNPA) and 4-nitrophenyl-(2-phenyl)-propanoate (pNPP) with high catalytic efficiency. AlleyCatE was created by introducing a single-histidine residue (His 144 ) into a hydrophobic pocket of calmodulin. In this work, we explore the determinants of catalytic properties of AlleyCatE. We obtained the pK a value of the catalytic histidine using experimental measurements by NMR and pH rate profile and compared these values to those predicted from electrostatics pK a calculations (from both empirical and continuum electrostatics calculations). Surprisingly, the pK a value of the catalytic histidine inside the hydrophobic pocket of calmodulin is elevated as compared to the model compound pK a value of this residue in water. We determined that a short-range favorable interaction with Glu 127 contributes to the elevated pK a of His 144 . We have rationally modulated local electrostatic potential in AlleyCatE to decrease the pK a of its active nucleophile, His 144 , by 0.7 units. As a direct result of the decrease in the His 144 pK a value, catalytic efficiency of the enzyme increased by 45% at pH 6. This work shows that a series of simple NMR experiments that can be performed using low field spectrometers, combined with straightforward computational analysis, provide rapid and accurate guidance to rationally improve catalytic efficiency of histidine-promoted catalysis. Proteins 2017; 85:1656-1665. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

  19. Improved outcome with novel device for low-pressure PTCA in de novo and in-stent lesions

    International Nuclear Information System (INIS)

    Ischinger, Thomas A.; Solar, Ronald J.; Hitzke, Evelyn

    2003-01-01

    Purpose: Complex lesion morphology requiring the use of high pressure to effect lumen expansion and in-stent restenosis (ISR) remain two indications that challenge conventional PTCA balloons. We report on a new PTCA device that is designed to provide precise, low-pressure dilatation of both de novo and in-stent lesions. Methods: The FX miniRAIL catheter (FX) has an integral wire positioned external to a dilating balloon and a short, 12-mm guidewire lumen distal to the balloon. The balloon inflates against the guidewire and the external wire to prevent slippage and to introduce high focal longitudinal stresses at low inflation pressures. In this initial study, the FX was used in 37 lesions (25 de novo, 12 in-stent; vessel reference diameter=2.73±0.49 mm) in 30 patients. A stepwise inflation protocol and QCA were used to determine the balloon pressure at which the stenosis was resolved (stenosis resolution pressure, SRP). Results: All lesions (100%) were easily reached, crossed and dilated without complication. The SRP was 4.5±2.9 atm, and no balloon slippage was observed. Residual stenosis after FX was 26.39±13.29%. Minor dissections (Types A and B) were observed in eight lesions (21.6%). Target lesion revascularization (TLR) and target vessel revascularization (TVR) at follow-up (8.1±1.5 months) were 8.3% and 12.5%, respectively. Conclusion: The design of the FX is versatile and appears to provide for a safe, effective and improved low-pressure PTCA technique in de novo and in-stent lesions

  20. Novos paradigmas literários

    Directory of Open Access Journals (Sweden)

    Denise Azevedo Duarte Guimarães

    2005-12-01

    Full Text Available O artigo estuda a emergência de novos paradigmas literários, procurando refletir acerca das textualidades contemporâneas. Focaliza os hipertextos informatizados e a poesia multimídia, com o intuito de desvendar como estão sendo criados novos procedimentos expressivos e em que medida eles podem ser identificados com reflexões teóricas anteriores acerca do texto literário impresso. Remete a questões ligadas à leitura dos diferentes tipos de signos e aos modos como eles se integram para a constituição dessas novíssimas linguagens híbridas em novos suportes.El artículo estudia la emergencia de nuevos paradigmas literarios, procurando reflejar acerca de las textualidades contemporáneas. Enfoca los hipertextos informatizados y la poesía multimedia, intentando desvendar cómo están siendo creados nuevos procedimientos expresivos y en qué medida ellos pueden ser identificados a reflexiones teóricas anteriores acerca del texto literario impreso. Remite a cuestiones ligadas a la lectura de los diferentes tipos de signos y a los modos cómo ellos se interaccionan para la constitución de los novísimos lenguajes híbridos en nuevos supuestos.This article investigates the emergence of new literary paradigms as it tries to understand new contemporary textualities. It analyses some hypertexts and multimedia poetry trying to trace how new expressive procedures are being created. How can these new languages be identified and what are their relations to previous theories which dealt with the literary printed text? This study approaches questions linked to the reading of different types of signs and the modes they function towards the fabrication of these new hybrid languages.

  1. Bioactive Mimetics of Conotoxins and other Venom Peptides

    Directory of Open Access Journals (Sweden)

    Peter J. Duggan

    2015-10-01

    Full Text Available Ziconotide (Prialt®, a synthetic version of the peptide ω-conotoxin MVIIA found in the venom of a fish-hunting marine cone snail Conus magnus, is one of very few drugs effective in the treatment of intractable chronic pain. However, its intrathecal mode of delivery and narrow therapeutic window cause complications for patients. This review will summarize progress in the development of small molecule, non-peptidic mimics of Conotoxins and a small number of other venom peptides. This will include a description of how some of the initially designed mimics have been modified to improve their drug-like properties.

  2. A Public Trial De Novo

    DEFF Research Database (Denmark)

    Vedel, Jane Bjørn; Gad, Christopher

    2011-01-01

    This article addresses the concept of “industrial interests” and examines its role in a topical controversy about a large research grant from a private foundation, the Novo Nordisk Foundation, to the University of Copenhagen. The authors suggest that the debate took the form of a “public trial” w.......” The article ends with a discussion of some implications of the analysis, including that policy making, academic research, and public debates might benefit from more detailed accounts of interests and stakes.......This article addresses the concept of “industrial interests” and examines its role in a topical controversy about a large research grant from a private foundation, the Novo Nordisk Foundation, to the University of Copenhagen. The authors suggest that the debate took the form of a “public trial......” where the grant and close(r) intermingling between industry and public research was prosecuted and defended. First, the authors address how the grant was framed in the media. Second, they redescribe the case by introducing new “evidence” that, because of this framing, did not reach “the court...

  3. Peptides Trapping Dioxins: A Docking-Based Inverse Screening Approach

    Directory of Open Access Journals (Sweden)

    German Perez

    2013-01-01

    Full Text Available A rapid and cost-effective computational methodology for designing and rationalizing the selection of small peptides as receptors for dioxin-like compounds was proposed. The backbone of the dioxin Ah receptor binding site was used to design a series of penta- and hexapeptide libraries, with 1400 elements in total. Peptide flexibility was considered and 10 conformers were found to be a good option to represent peptide conformational space with fair speed-accuracy ratio. Each peptide conformer was treated as a possible receptor, generating a dedicated box and then running a docking process using as ligands a family of 76 dibenzo-p-dioxins and 113 dibenzofurans mono- and polychlorinated. Significant predictions were confirmed by comparing primary structure of top and bottom ranked peptides binding dioxins confirming that scrambled positions of the same amino acids gave completely different predicted binding. The hexapeptide EWFQPW, with the best binding score, was chosen as selective sorbent material in solid-phase extraction. The retention performances were tested using the 2,3,7,8-tetrachlorodibenzo-p-dioxin and two polychlorinated biphenyls in order to verify the hexapeptide specificity. The solid-phase extraction experimental procedure was optimized, and analytical parameters of hexapeptide sorbent material were compared with the resin without hexapeptide and a commercial reversed phase cartridge.

  4. Antimicrobial Peptides for Therapeutic Applications: A Review

    Directory of Open Access Journals (Sweden)

    Tsogbadrakh Mishig-Ochir

    2012-10-01

    Full Text Available Antimicrobial peptides (AMPs have been considered as potential therapeutic sources of future antibiotics because of their broad-spectrum activities and different mechanisms of action compared to conventional antibiotics. Although AMPs possess considerable benefits as new generation antibiotics, their clinical and commercial development still have some limitations, such as potential toxicity, susceptibility to proteases, and high cost of peptide production. In order to overcome those obstacles, extensive efforts have been carried out. For instance, unusual amino acids or peptido-mimetics are introduced to avoid the proteolytic degradation and the design of short peptides retaining antimicrobial activities is proposed as a solution for the cost issue. In this review, we focus on small peptides, especially those with less than twelve amino acids, and provide an overview of the relationships between their three-dimensional structures and antimicrobial activities. The efforts to develop highly active AMPs with shorter sequences are also described.

  5. Applications and Challenges for Use of Cell-Penetrating Peptides as Delivery Vectors for Peptide and Protein Cargos

    Directory of Open Access Journals (Sweden)

    Mie Kristensen

    2016-01-01

    Full Text Available The hydrophilic nature of peptides and proteins renders them impermeable to cell membranes. Thus, in order to successfully deliver peptide and protein-based therapeutics across the plasma membrane or epithelial and endothelial barriers, a permeation enhancing strategy must be employed. Cell-penetrating peptides (CPPs constitute a promising tool and have shown applications for peptide and protein delivery into cells as well as across various epithelia and the blood-brain barrier (BBB. CPP-mediated delivery of peptides and proteins may be pursued via covalent conjugation of the CPP to the cargo peptide or protein or via physical complexation obtained by simple bulk-mixing of the CPP with its cargo. Both approaches have their pros and cons, and which is the better choice likely relates to the physicochemical properties of the CPP and its cargo as well as the route of administration, the specific barrier and the target cell. Besides the physical barrier, a metabolic barrier must be taken into consideration when applying peptide-based delivery vectors, such as the CPPs, and stability-enhancing strategies are commonly employed to prolong the CPP half-life. The mechanisms by which CPPs translocate cell membranes are believed to involve both endocytosis and direct translocation, but are still widely investigated and discussed. The fact that multiple factors influence the mechanisms responsible for cellular CPP internalization and the lack of sensitive methods for detection of the CPP, and in some cases the cargo, further complicates the design and conduction of conclusive mechanistic studies.

  6. Peptide Logic Circuits Based on Chemoenzymatic Ligation for Programmable Cell Apoptosis.

    Science.gov (United States)

    Li, Yong; Sun, Sujuan; Fan, Lin; Hu, Shanfang; Huang, Yan; Zhang, Ke; Nie, Zhou; Yao, Shouzhou

    2017-11-20

    A novel and versatile peptide-based bio-logic system capable of regulating cell function is developed using sortase A (SrtA), a peptide ligation enzyme, as a generic processor. By modular peptide design, we demonstrate that mammalian cells apoptosis can be programmed by peptide-based logic operations, including binary and combination gates (AND, INHIBIT, OR, and AND-INHIBIT), and a complex sequential logic circuit (multi-input keypad lock). Moreover, a proof-of-concept peptide regulatory circuit was developed to analyze the expression profile of cell-secreted protein biomarkers and trigger cancer-cell-specific apoptosis. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  7. An Open-Source Automated Peptide Synthesizer Based on Arduino and Python.

    Science.gov (United States)

    Gali, Hariprasad

    2017-10-01

    The development of the first open-source automated peptide synthesizer, PepSy, using Arduino UNO and readily available components is reported. PepSy was primarily designed to synthesize small peptides in a relatively small scale (<100 µmol). Scripts to operate PepSy in a fully automatic or manual mode were written in Python. Fully automatic script includes functions to carry out resin swelling, resin washing, single coupling, double coupling, Fmoc deprotection, ivDde deprotection, on-resin oxidation, end capping, and amino acid/reagent line cleaning. Several small peptides and peptide conjugates were successfully synthesized on PepSy with reasonably good yields and purity depending on the complexity of the peptide.

  8. A Novel Soluble Peptide with pH-Responsive Membrane Insertion.

    Science.gov (United States)

    Nguyen, Vanessa P; Alves, Daiane S; Scott, Haden L; Davis, Forrest L; Barrera, Francisco N

    2015-11-03

    Several diseases, such as cancer, are characterized by acidification of the extracellular environment. Acidosis can be employed as a target to specifically direct therapies to the diseased tissue. We have used first principles to design an acidity-triggered rational membrane (ATRAM) peptide with high solubility in solution that is able to interact with lipid membranes in a pH-dependent fashion. Biophysical studies show that the ATRAM peptide binds to the surface of lipid membranes at pH 8.0. However, acidification leads to the peptide inserting into the lipid bilayer as a transmembrane α-helix. The insertion of ATRAM into membranes occurs at a moderately acidic pH (with a pK of 6.5), similar to the extracellular pH found in solid tumors. Studies with human cell lines showed a highly efficient pH-dependent membrane targeting, without causing toxicity. Here we show that it is possible to rationally design a soluble peptide that selectively targets cell membranes in acidic environments.

  9. Rational design of syn-safencin, a novel linear antimicrobial peptide derived from the circular bacteriocin safencin AS-48.

    Science.gov (United States)

    Fields, Francisco R; Carothers, Katelyn E; Balsara, Rashna D; Ploplis, Victoria A; Castellino, Francis J; Lee, Shaun W

    2018-06-01

    Bacteriocins hold unprecedented promise as a largely untapped source of antibiotic alternatives in the age of multidrug resistance. Here, we describe the first approach to systematically design variants of a novel AS-48 bacteriocin homologue, which we have termed safencin AS-48, from Bacillus safensis, to gain insights into engineering improved activity of bacteriocins. A library of synthetic peptides in which systematic amino acid substitutions to vary the periodicity and abundance of polar, acidic, aliphatic, and hydrophobic residues were generated for a total of 96 novel peptide variants of a single bacteriocin candidate. Using this method, we identified nine synthetic safencin (syn-safencin) variants with broad and potent antimicrobial activities with minimal inhibitory concentrations (MIC) as low as 250 nM against E. coli, P. aeruginosa, X. axonopodis, and S. pyogenes with minimal cytotoxicity to mammalian cells. It is anticipated that the strategies we have developed will serve as general guides for tuning the specificity of a given natural bacteriocin compound for therapeutic specificity.

  10. Carrier protein influences immunodominance of a known epitope: implication in peptide vaccine design.

    Science.gov (United States)

    Ghosh, Moumita; Solanki, Ashish K; Roy, Koushik; Dhoke, Reema R; Ashish; Roy, Syamal

    2013-09-23

    We investigated how the processing of a given antigen by antigen presenting cells (APC) is dictated by the conformation of the antigen and how this governs the immunodominance hierarchy. To address the question, a known immunodominant sequence of bacteriophage lambda repressor N-terminal sequence 12-26 [λR(12-26)] was engineered at the N and C termini of a heterologous leishmanial protein, Kinetoplastid membrane protein-11 (KMP-11); the resulting proteins were defined as N-KMP-11 and C-KMP-11 respectively. The presence of λR(12-26) in N-KMP-11 and C-KMP-11 was established by western blot analysis with antibody to λR(12-26) peptide. N-KMP-11 but not C-KMP-11 could stimulate the anti λR(12-26) T-cell clonal population very efficiently in the presence of APCs. Priming of BALB/c mice with N-KMP-11 or C-KMP-11 generated similar levels of anti-KMP-11 IgG, but anti-λR(12-26) specific IgG was observed only upon priming with N-KMP-11. Interestingly, uptake of both N-KMP-11 and C-KMP-11 by APCs was similar but catabolism of N-KMP-11 but not C-KMP-11 was biphasic and fast at the initial time point. Kratky plots of small angle X-ray scattering showed that while N-KMP-11 adopts flexible Gaussian type of topology, C-KMP-11 prefers Globular nature. To show that KMP-11 is not unique as a carrier protein, an epitope (SPITBTNLBTMBK) of Plasmodium yoelii (PY) apical membrane protein 1[AMA-1 (136-148)], is placed at the C and N terminals of a dominant T-cell epitope of ovalbumin protein OVA(323-339) and the resulting peptides are defined as PY-OVA and OVA-PY respectively. Interestingly, only OVA-PY could stimulate anti-OVA T-cells and produce IgG response upon priming of BALB/c mice with it. Thus for rational design of peptide vaccine it is important to place the dominant epitope appropriately in the context of the carrier protein. Copyright © 2013 Elsevier Ltd. All rights reserved.

  11. EThcD Discrimination of Isomeric Leucine/Isoleucine Residues in Sequencing of the Intact Skin Frog Peptides with Intramolecular Disulfide Bond

    Science.gov (United States)

    Samgina, Tatiana Yu; Kovalev, Sergey V.; Tolpina, Miriam D.; Trebse, Polonca; Torkar, Gregor; Lebedev, Albert T.

    2018-05-01

    Our scientific interests involve de novo sequencing of non-tryptic natural amphibian skin peptides including those with intramolecular S-S bond by means of exclusively mass spectrometry. Reliable discrimination of the isomeric leucine/isoleucine residues during peptide sequencing by means of mass spectrometry represents a bottleneck in the workflow for complete automation of the primary structure elucidation of these compounds. MS3 is capable of solving the problem. Earlier we demonstrated the advanced efficiency of ETD-HCD method to discriminate Leu/Ile in individual peptides by consecutive application of ETD to the polyprotonated peptides followed by HCD applied to the manually selected primary z-ions with the targeted isomeric residues at their N-termini and registration of the characteristic w-ions. Later this approach was extended to deal with several (4-7) broad band mass ranges, without special isolation of the primary z-ions. The present paper demonstrates an advanced version of this method when EThcD is applied in the whole mass range to a complex mixture of natural non-tryptic peptides without their separation and intermediate isolation of the targeted z-ions. The proposed EThcD method showed over 81% efficiency for the large natural peptides with intact disulfide ring, while the interfering process of radical site migration is suppressed. Due to higher speed and sensitivity, the proposed EThcD approach facilitates the analytical procedure and allows for the automation of the entire experiment and data processing. Moreover, in some cases it gives a chance to establish the nature of the residues in the intact intramolecular disulfide loops. [Figure not available: see fulltext.

  12. Structures and related properties of helical, disulfide-stabilized peptides

    Energy Technology Data Exchange (ETDEWEB)

    Pagel, Mark D. [Univ. of California, Berkeley, CA (United States). Dept. of Chemistry

    1993-11-01

    The three dimensional structure of several peptides were determined by NMR spectroscopy and distance geometry calculations. Each peptide formed a predictable, rigid structure, consisting of an α-helix, a "scaffold" region which packed along one face of the helix, and two disulfide bridges which covalently connect the helix and scaffold regions. The peptide Apa-M5 was designed to constrain the M5 peptide from MLCK in a helical geometry using the apamin disulfide scaffold. This scaffold constrains the N- terminal end of the helix with two disulfide bridges and a reverse turn. Like the M5 peptide, Apa-M5 was found to bind calmodulin in a Ca2+-dependent 1:1 stoichiometry. However, the dissociation constant of the (Apa-M5)-calmodulin complex, 107 nM, was 100-fold higher than the dissociation constant of the M5-calmodulin complex. This difference was due to a putative steric overlap between the Apa-M5 scaffold and calmodulin. The peptide Apa-Cro was designed to replace the large structural protein matrix of λ Cro with the apamin disulfide scaffold. However, Apa-Cro did not bind the consensus DNA operator half-site of λ Cro, probably due to a steric overlap between the Apa-Cro disulfide framework and the DNA. The amino acid sequence of the scaffold-disulfide bridge arrangement of the peptide Max was derived from the core sequence of scyllatoxin, which contains an α-helix constrained at the C-terminal end by two disulfide bridges and a two-stranded βsheet scaffold. Max was shown to fold with >84% yield to form a predictable, stable structure that is similar to scyllatoxin. The folding and stability properties of Max make this scaffold and disulfide bridge arrangement an ideal candidate for the development of hybrid sequence peptides. The dynamics of a fraying C-terminal end of the helix of the peptide Apa-AlaN was determined by analysis of 15N NMR relaxation properties.

  13. Recurrence risk in de novo structural chromosomal rearrangements.

    Science.gov (United States)

    Röthlisberger, Benno; Kotzot, Dieter

    2007-08-01

    According to the textbook of Gardner and Sutherland [2004], the standard on genetic counseling for chromosome abnormalities, the recurrence risk of de novo structural or combined structural and numeric chromosome rearrangements is less than 0.5-2% and takes into account recurrence by chance, gonadal mosaicism, and somatic-gonadal mosaicism. However, these figures are roughly estimated and neither any systematic study nor exact or evidence-based risk calculations are available. To address this question, an extensive literature search was performed and surprisingly only 29 case reports of recurrence of de novo structural or combined structural and numeric chromosomal rearrangements were found. Thirteen of them were with a trisomy 21 due to an i(21q) replacing one normal chromosome 21. In eight of them low-level mosaicism in one of the parents was found either in fibroblasts or in blood or in both. As a consequence of the low number of cases and theoretical considerations (clinical consequences, mechanisms of formation, etc.), the recurrence risk should be reduced to less than 1% for a de novo i(21q) and to even less than 0.3% for all other de novo structural or combined structural and numeric chromosomal rearrangements. As the latter is lower than the commonly accepted risk of approximately 0.3% for indicating an invasive prenatal diagnosis and as the risk of abortion of a healthy fetus after chorionic villous sampling or amniocentesis is higher than approximately 0.5%, invasive prenatal investigation in most cases is not indicated and should only be performed if explicitly asked by the parents subsequent to appropriate genetic counseling. (c) 2007 Wiley-Liss, Inc.

  14. Albumin-derived peptides efficiently reduce renal uptake of radiolabelled peptides

    International Nuclear Information System (INIS)

    Vegt, Erik; Eek, Annemarie; Oyen, Wim J.G.; Gotthardt, Martin; Boerman, Otto C.; Jong, Marion de

    2010-01-01

    In peptide-receptor radionuclide therapy (PRRT), the maximum activity dose that can safely be administered is limited by high renal uptake and retention of radiolabelled peptides. The kidney radiation dose can be reduced by coinfusion of agents that competitively inhibit the reabsorption of radiolabelled peptides, such as positively charged amino acids, Gelofusine, or trypsinised albumin. The aim of this study was to identify more specific and potent inhibitors of the kidney reabsorption of radiolabelled peptides, based on albumin. Albumin was fragmented using cyanogen bromide and six albumin-derived peptides with different numbers of electric charges were selected and synthesised. The effect of albumin fragments (FRALB-C) and selected albumin-derived peptides on the internalisation of 111 In-albumin, 111 In-minigastrin, 111 In-exendin and 111 In-octreotide by megalin-expressing cells was assessed. In rats, the effect of Gelofusine and albumin-derived peptides on the renal uptake and biodistribution of 111 In-minigastrin, 111 In-exendin and 111 In-octreotide was determined. FRALB-C significantly reduced the uptake of all radiolabelled peptides in vitro. The albumin-derived peptides showed different potencies in reducing the uptake of 111 In-albumin, 111 In-exendin and 111 In-minigastrin in vitro. The most efficient albumin-derived peptide (peptide 6), was selected for in vivo testing. In rats, 5 mg of peptide 6 very efficiently inhibited the renal uptake of 111 In-minigastrin, by 88%. Uptake of 111 In-exendin and 111 In-octreotide was reduced by 26 and 33%, respectively. The albumin-derived peptide 6 efficiently inhibited the renal reabsorption of 111 In-minigastrin, 111 In-exendin and 111 In-octreotide and is a promising candidate for kidney protection in PRRT. (orig.)

  15. Molecular evolution of a peptide GPCR ligand driven by artificial neural networks.

    Directory of Open Access Journals (Sweden)

    Sebastian Bandholtz

    Full Text Available Peptide ligands of G protein-coupled receptors constitute valuable natural lead structures for the development of highly selective drugs and high-affinity tools to probe ligand-receptor interaction. Currently, pharmacological and metabolic modification of natural peptides involves either an iterative trial-and-error process based on structure-activity relationships or screening of peptide libraries that contain many structural variants of the native molecule. Here, we present a novel neural network architecture for the improvement of metabolic stability without loss of bioactivity. In this approach the peptide sequence determines the topology of the neural network and each cell corresponds one-to-one to a single amino acid of the peptide chain. Using a training set, the learning algorithm calculated weights for each cell. The resulting network calculated the fitness function in a genetic algorithm to explore the virtual space of all possible peptides. The network training was based on gradient descent techniques which rely on the efficient calculation of the gradient by back-propagation. After three consecutive cycles of sequence design by the neural network, peptide synthesis and bioassay this new approach yielded a ligand with 70fold higher metabolic stability compared to the wild type peptide without loss of the subnanomolar activity in the biological assay. Combining specialized neural networks with an exploration of the combinatorial amino acid sequence space by genetic algorithms represents a novel rational strategy for peptide design and optimization.

  16. NovoTTF™-100A System (Tumor Treating Fields) transducer array layout planning for glioblastoma: a NovoTAL™ system user study.

    Science.gov (United States)

    Chaudhry, Aafia; Benson, Laura; Varshaver, Michael; Farber, Ori; Weinberg, Uri; Kirson, Eilon; Palti, Yoram

    2015-11-11

    Optune™, previously known as the NovoTTF-100A System™, generates Tumor Treating Fields (TTFields), an effective anti-mitotic therapy for glioblastoma. The system delivers intermediate frequency, alternating electric fields to the supratentorial brain. Patient therapy is personalized by configuring transducer array layout placement on the scalp to the tumor site using MRI measurements and the NovoTAL System. Transducer array layout mapping optimizes therapy by maximizing electric field intensity to the tumor site. This study evaluated physician performance in conducting transducer array layout mapping using the NovoTAL System compared with mapping performed by the Novocure in-house clinical team. Fourteen physicians (7 neuro-oncologists, 4 medical oncologists, and 3 neurosurgeons) evaluated five blinded cases of recurrent glioblastoma and performed head size and tumor location measurements using a standard Digital Imaging and Communications in Medicine reader. Concordance with Novocure measurement and intra- and inter-rater reliability were assessed using relevant correlation coefficients. The study criterion for success was a concordance correlation coefficient (CCC) >0.80. CCC for each physician versus Novocure on 20 MRI measurements was 0.96 (standard deviation, SD ± 0.03, range 0.90-1.00), indicating very high agreement between the two groups. Intra- and inter-rater reliability correlation coefficients were similarly high: 0.83 (SD ±0.15, range 0.54-1.00) and 0.80 (SD ±0.18, range 0.48-1.00), respectively. This user study demonstrated an excellent level of concordance between prescribing physicians and Novocure in-house clinical teams in performing transducer array layout planning. Intra-rater reliability was very high, indicating reproducible performance. Physicians prescribing TTFields, when trained on the NovoTAL System, can independently perform transducer array layout mapping required for the initiation and maintenance of patients on TTFields

  17. The application of 199Hg NMR and 199mHg perturbed angular correlation (PAC) spectroscopy to define the biological chemistry of HgII

    DEFF Research Database (Denmark)

    Iranzo, Olga; Thulstrup, Peter Waaben; Ryu, Seung-baek

    2007-01-01

    The use of de novo designed peptides is a powerful strategy to elucidate HgII-protein interactions and to gain insight into the chemistry of HgII in biological systems. Cysteine derivatives of the designed -helical peptides of the TRI family [Ac-G-(LaKbAcLdEeEfKg)4-G-NH2] bind HgII at high p...... to characterize the distinct species that are generated under different pH conditions and peptide TRI L9C/HgII ratios. These studies prove for the first time the formation of [Hg{(TRI L9C)2-(TRI L9C H)}], a dithiolate-HgII complex in the hydrophobic interior of the three-stranded coiled coil (TRI L9C)3. 199Hg NMR...

  18. A Terra em Transe: o cosmopolitismo às avessas do cinema novo

    Directory of Open Access Journals (Sweden)

    Angela Prysthon

    2008-11-01

    Full Text Available Usando como referencial teórico os estudos culturais, este artigo analisa o cinema novo brasileiro como parte de uma estratégia terceiro mundista de conceber a cultura. A partir da emergência do conceito de terceiro mundo e das lutas de descolonização nos anos 1950 e 1960, a ideologia cosmopolita foi sendo vista pelos intelectuais de esquerda como a versão cultural da aliança com as forças hegemônicas da Europa e dos Estados Unidos. O projeto do cinema novo chama a atenção por suas afinidades ideológicas com o terceiro mundismo, mas, paradoxalmente, trazendo à tona uma polí­tica cosmopolita da periferia. Palavras-chave cinema novo, identidade, cultura brasileira, terceiro mundismo, estudos culturais. Abstract Using the cultural studies theoretical framework, this paper analyzes the cinema novo movement in Brazil as a part of the Third World conception of culture. Following the creation of the term "Third World" and the international politics of colonial independence of the 1950s and 1960s, a cosmopolitan attitude was seen by the intellectuals of the left as a cultural version of the alliance with the hegemonic forces of Europe and North America. Even though the cinema novo project can be associated with the ideology of an united Third World ,it brings about, paradoxically, a very cosmopolitan politics of the periphery. Key words cinema novo, identity, Brazilian culture, third world, cultural studies.

  19. [Ala12]MCD peptide: a lead peptide to inhibitors of immunoglobulin E binding to mast cell receptors.

    Science.gov (United States)

    Buku, A; Condie, B A; Price, J A; Mezei, M

    2005-09-01

    An effort was made to discover mast cell degranulating (MCD) peptide analogs that bind with high affinity to mast cell receptors without triggering secretion of histamine or other mediators of the allergic reaction initiated by immunoglobulin E (IgE) after mast cell activation. Such compounds could serve as inhibitors of IgE binding to mast cell receptors. An alanine scan of MCD peptide reported previously showed that the analog [Ala12]MCD was 120-fold less potent in histamine-releasing activity and fivefold more potent in binding affinity to mast cell receptors than the parent MCD peptide. Because this analog showed marginal intrinsic activity and good binding affinity it was subsequently tested in the present study as an IgE inhibitor. In contrast to MCD peptide, [Ala12]MCD showed a 50% inhibition of IgE binding to the Fc epsilon RI alpha mast cell receptor by using rat basophilic leukemia (RBL-2H3) mast cells and fluorescence polarization. Furthermore, in a beta-hexosaminidase secretory assay, the peptide also showed a 50% inhibition of the secretion of this enzyme caused by IgE. An attempt was made to relate structural changes and biologic differences between the [Ala12]MCD analog and the parent MCD peptide. The present results show that [Ala12]MCD may provide a base for designing agents to prevent IgE/Fc epsilon RI alpha interactions and, consequently, allergic conditions.

  20. Eculizumab for drug-induced de novo posttransplantation thrombotic microangiopathy: A case report.

    Science.gov (United States)

    Safa, Kassem; Logan, Merranda S; Batal, Ibrahim; Gabardi, Steven; Rennke, Helmut G; Abdi, Reza

    2015-02-01

    De novo thrombotic microangiopathy (TMA) following renal transplantation is a severe complication associated with high rates of allograft failure. Several immunosuppressive agents are associated with TMA. Conventional approaches to managing this entity, such as withdrawal of the offending agent and/or plasmapheresis, often offer limited help, with high rates of treatment failure and graft loss. We herein report a case of drug induced de novo TMA successfully treated using the C5a inhibitor eculizumab in a renal transplant patient. This report highlights a potentially important role for eculizumab in settings where drug-induced de novo TMA is refractory to conventional therapies.

  1. Interactions of Bio-Inspired Membranes with Peptides and Peptide-Mimetic Nanoparticles

    Directory of Open Access Journals (Sweden)

    Michael Sebastiano

    2015-08-01

    Full Text Available Via Dissipative Particle Dynamics (DPD and implicit solvent coarse-grained (CG Molecular Dynamics (MD we examine the interaction of an amphiphilic cell-penetrating peptide PMLKE and its synthetic counterpart with a bio-inspired membrane. We use the DPD technique to investigate the interaction of peptide-mimetic nanoparticles, or nanopins, with a three-component membrane. The CG MD approach is used to investigate the interaction of a cell-penetrating peptide PMLKE with single-component membrane. We observe the spontaneous binding and subsequent insertion of peptide and nanopin in the membrane by using CG MD and DPD approaches, respectively. In addition, we find that the insertion of peptide and nanopins is mainly driven by the favorable enthalpic interactions between the hydrophobic components of the peptide, or nanopin, and the membrane. Our study provides insights into the mechanism underlying the interactions of amphiphilic peptide and peptide-mimetic nanoparticles with a membrane. The result of this study can be used to guide the functional integration of peptide and peptide-mimetic nanoparticles with a cell membrane.

  2. De novo autoimmune hepatitis after liver transplantation.

    Science.gov (United States)

    Lohse, Ansgar W; Weiler-Norman, Christina; Burdelski, Martin

    2007-10-01

    The Kings College group was the first to describe a clinical syndrome similar to autoimmune hepatitis in children and young adults transplanted for non-immune mediated liver diseases. They coined the term "de novo autoimmune hepatitis". Several other liver transplant centres confirmed this observation. Even though the condition is uncommon, patients with de novo AIH are now seen in most of the major transplant centres. The disease is usually characterized by features of acute hepatitis in otherwise stable transplant recipients. The most characteristic laboratory hallmark is a marked hypergammaglobulinaemia. Autoantibodies are common, mostly ANA. We described also a case of LKM1-positivity in a patients transplanted for Wilson's disease, however this patients did not develop clinical or histological features of AIH. Development of SLA/LP-autoantibodies is also not described. Therefore, serologically de novo AIH appears to correspond to type 1 AIH. Like classical AIH patients respond promptly to treatment with increased doses of prednisolone and azathioprine, while the calcineurin inhibitors cyclosporine or tacrolimus areof very limited value - which is not surprising, as almost all patients develop de novo AIH while receiving these drugs. Despite the good response to treatment, most patients remain a clinical challenge as complete stable remissions are uncommon and flares, relapses and chronic disease activity can often occur. Pathogenetically this syndrome is intriguing. It is not clear, if the immune response is directed against allo-antigens, neo-antigens in the liver, or self-antigens, possibly shared by donor and host cells. It is very likely that the inflammatory milieu due to alloreactive cells in the transplanted organ contribute to the disease process. Either leading to aberrant antigen presentation, or providing co-stimulatory signals leading to the breaking of self-tolerance. The development of this disease in the presence of treatment with calcineurin

  3. HIPdb: a database of experimentally validated HIV inhibiting peptides.

    Science.gov (United States)

    Qureshi, Abid; Thakur, Nishant; Kumar, Manoj

    2013-01-01

    Besides antiretroviral drugs, peptides have also demonstrated potential to inhibit the Human immunodeficiency virus (HIV). For example, T20 has been discovered to effectively block the HIV entry and was approved by the FDA as a novel anti-HIV peptide (AHP). We have collated all experimental information on AHPs at a single platform. HIPdb is a manually curated database of experimentally verified HIV inhibiting peptides targeting various steps or proteins involved in the life cycle of HIV e.g. fusion, integration, reverse transcription etc. This database provides experimental information of 981 peptides. These are of varying length obtained from natural as well as synthetic sources and tested on different cell lines. Important fields included are peptide sequence, length, source, target, cell line, inhibition/IC(50), assay and reference. The database provides user friendly browse, search, sort and filter options. It also contains useful services like BLAST and 'Map' for alignment with user provided sequences. In addition, predicted structure and physicochemical properties of the peptides are also included. HIPdb database is freely available at http://crdd.osdd.net/servers/hipdb. Comprehensive information of this database will be helpful in selecting/designing effective anti-HIV peptides. Thus it may prove a useful resource to researchers for peptide based therapeutics development.

  4. A randomized, double-blind, cross-over, phase IV trial of oros-methylphenidate (CONCERTA(®)) and generic novo-methylphenidate ER-C (NOVO-generic).

    Science.gov (United States)

    Fallu, Angelo; Dabouz, Farida; Furtado, Melissa; Anand, Leena; Katzman, Martin A

    2016-08-01

    Attention-deficit/hyperactivity disorder (ADHD) is a common neurobehavioral disorder with onset during childhood. Multiple aspects of a child's development are hindered, in both home and school settings, with negative impacts on social, emotional, and cognitive functioning. If left untreated, ADHD is commonly associated with poor academic achievement and low occupational status, as well as increased risk of substance abuse and delinquency. The objective of this study was to evaluate adult ADHD subject reported outcomes when switched from a stable dose of CONCERTA(®) to the same dose of generic Novo-methylphenidate ER-C(®). Randomized, double-blind, cross-over, phase IV trial consisted of two phases in which participants with a primary diagnosis of ADHD were randomized in a 1:1 ratio to 3 weeks of treatment with CONCERTA or generic Novo-Methylphenidate ER-C. Following 3 weeks of treatment, participants were crossed-over to receive the other treatment for an additional 3 weeks. Primary efficacy was assessed through the use of the Treatment Satisfaction Questionnaire for Medication, Version II (TSQM-II). Participants with ADHD treated with CONCERTA were more satisfied in terms of efficacy and side effects compared to those receiving an equivalent dose of generic Novo-Methylphenidate ER-C. All participants chose to continue with CONCERTA treatment at the conclusion of the study. Although CONCERTA and generic Novo-Methylphenidate ER-C have been deemed bioequivalent, however the present findings demonstrate clinically and statistically significant differences between generic and branded CONCERTA. Further investigation of these differences is warranted.

  5. Arginine de novo and nitric oxide production in disease states

    OpenAIRE

    Luiking, Yvette C.; Ten Have, Gabriella A. M.; Wolfe, Robert R.; Deutz, Nicolaas E. P.

    2012-01-01

    Arginine is derived from dietary protein intake, body protein breakdown, or endogenous de novo arginine production. The latter may be linked to the availability of citrulline, which is the immediate precursor of arginine and limiting factor for de novo arginine production. Arginine metabolism is highly compartmentalized due to the expression of the enzymes involved in arginine metabolism in various organs. A small fraction of arginine enters the NO synthase (NOS) pathway. Tetrahydrobiopterin ...

  6. [Plant signaling peptides. Cysteine-rich peptides].

    Science.gov (United States)

    Ostrowski, Maciej; Kowalczyk, Stanisław

    2015-01-01

    Recent bioinformatic and genetic analyses of several model plant genomes have revealed the existence of a highly abundant group of signaling peptides that are defined as cysteine-rich peptides (CRPs). CRPs are usually in size between 50 and 90 amino acid residues, they are positively charged, and they contain 4-16 cysteine residues that are important for the correct conformational folding. Despite the structural differences among CRP classes, members from each class have striking similarities in their molecular properties and function. The present review presents the recent progress in research on signaling peptides from several families including: EPF/EPFL, SP11/SCR, PrsS, RALF, LURE, and some other peptides belonging to CRP group. There is convincing evidence indicating multiple roles for these CRPs as signaling molecules during the plant life cycle, ranging from stomata development and patterning, self-incompatibility, pollen tube growth and guidance, reproductive processes, and nodule formation.

  7. Targeting Interleukin-4 Receptor Alpha by Hybrid Peptide for Novel Biliary Tract Cancer Therapy

    Directory of Open Access Journals (Sweden)

    Kahori Seto

    2014-01-01

    Full Text Available It is known that the interleukin-4 receptor α (IL-4Rα is highly expressed on the surface of various human solid tumors. We previously designed novel IL-4Rα-lytic hybrid peptide composed of binding peptide to IL-4Rα and cell-lytic peptide and reported that the designed IL-4Rα-lytic hybrid peptide exhibited cytotoxic and antitumor activity both in vitro and in vivo against the human pancreatic cancer cells expressing IL-4Rα. Here, we evaluated the antitumor activity of the IL-4Rα-lytic hybrid peptide as a novel molecular targeted therapy for human biliary tract cancer (BTC. The IL-4Rα-lytic hybrid peptide showed cytotoxic activity in six BTC cell lines with a concentration that killed 50% of all cells (IC50 as low as 5 μM. We also showed that IL-4Rα-lytic hybrid peptide in combination with gemcitabine exhibited synergistic cytotoxic activity in vitro. In addition, intravenous administration of IL-4Rα-lytic hybrid peptide significantly inhibited tumor growth in a xenograft model of human BTC in vivo. Taken together, these results indicated that the IL-4Rα-lytic hybrid peptide is a potent agent that might provide a novel therapy for patients with BTC.

  8. A novel BLyS antagonist peptide designed based on the 3-D complex structure of BCMA and BLyS

    International Nuclear Information System (INIS)

    Sun Jian; Feng Jiannan; Li Yan; Shen Beifen

    2006-01-01

    B lymphocyte stimulator (BLyS) is a member of tumor necrosis factor (TNF) family. Because of its roles in autoimmune diseases such as systemic lupus erythematosus (SLE), rheumatoid arthritis (RA), and Sjogren syndrome (SS), BLyS antagonists have been tested to treat SLE- and RA-like symptoms in mice and obtained optimistic results. So far, reported BLyS antagonists were mostly decoyed BLyS receptors or anti-BLyS antibodies. In this study, a novel BLyS antagonist peptide, PT, was designed based on the modeling 3-D complex structure of BCMA and BLyS. The interaction mode of PT with BLyS was analyzed theoretically. The results of competitive ELISA demonstrated that PT could inhibit the binding of BCMA-Fc and anti-BLyS antibody to BLyS in vitro. In addition, PT could partly block the proliferating activity of BLyS on mice splenocytes. The BLyS antagonizing activity of PT was significant (p < 0.05). This study highlights the possibility of using BLyS antagonist peptide to neutralize BLyS activity. Further optimization of PT with computer-guided molecular design method to enhance its biopotency may be useful in developing new BLyS antagonists to treat BLyS-related autoimmune diseases

  9. Affinity Purification and Comparative Biosensor Analysis of Citrulline-Peptide-Specific Antibodies in Rheumatoid Arthritis

    Directory of Open Access Journals (Sweden)

    Eszter Szarka

    2018-01-01

    Full Text Available Background: In rheumatoid arthritis (RA, anti-citrullinated protein/peptide antibodies (ACPAs are responsible for disease onset and progression, however, our knowledge is limited on ligand binding affinities of autoantibodies with different citrulline-peptide specificity. Methods: Citrulline-peptide-specific ACPA IgGs were affinity purified and tested by ELISA. Binding affinities of ACPA IgGs and serum antibodies were compared by surface plasmon resonance (SPR analysis. Bifunctional nanoparticles harboring a multi-epitope citrulline-peptide and a complement-activating peptide were used to induce selective depletion of ACPA-producing B cells. Results: KD values of affinity-purified ACPA IgGs varied between 10−6 and 10−8 M and inversely correlated with disease activity. Based on their cross-reaction with citrulline-peptides, we designed a novel multi-epitope peptide, containing Cit-Gly and Ala-Cit motifs in two–two copies, separated with a short, neutral spacer. This peptide detected antibodies in RA sera with 66% sensitivity and 98% specificity in ELISA and was recognized by 90% of RA sera, while none of the healthy samples in SPR. When coupled to nanoparticles, the multi-epitope peptide specifically targeted and depleted ACPA-producing B cells ex vivo. Conclusions: The unique multi-epitope peptide designed based on ACPA cross-reactivity might be suitable to develop better diagnostics and novel therapies for RA.

  10. 1,2,3-Triazole Rings as a Disulfide Bond Mimetic in Chimeric AGRP-Melanocortin Peptides: Design, Synthesis, and Functional Characterization.

    Science.gov (United States)

    Tala, Srinivasa R; Singh, Anamika; Lensing, Cody J; Schnell, Sathya M; Freeman, Katie T; Rocca, James R; Haskell-Luevano, Carrie

    2018-05-16

    The melanocortin system is involved in the regulation of complex physiological functions, including energy and weight homeostasis, feeding behavior, inflammation, sexual function, pigmentation, and exocrine gland function. The five melanocortin receptors that belong to the superfamily of G protein-coupled receptors (GPCRs) are regulated by endogenously expressed agonists and antagonists. The aim of this study was to explore the potential of replacing the disulfide bridge in chimeric AGRP-melanocortin peptide Tyr-c[Cys-His-d-Phe-Arg-Trp-Asn-Ala-Phe-Cys]-Tyr-NH 2 (1) with 1,2,3-triazole moieties. A series of 1,2,3-triazole-bridged peptidomimetics were designed, synthesized, and pharmacologically evaluated at the mouse melanocortin receptors. The ligands possessed nanomolar to micromolar agonist cAMP signaling potency. A key finding was that the disulfide bond in peptide 1 can be replaced with the monotriazole ring with minimal effect on the functional activity at the melanocortin receptors. The 1,5-disubstituted triazole-bridged peptide 6 showed equipotent functional activity at the mMC3R and modest 5-fold decreased agonist potency at the mMC4R compared to those of 1. Interestingly, the 1,4- and 1,5-disubstituted isomers of the triazole ring resulted in different selectivities at the receptor subtypes, indicating subtle structural features that may be exploited in the generation of selective melanocortin ligands. Introducing cyclic and acyclic bis-triazole moieties into chimeric AGRP template 1 generally decreased agonist activity. These results will be useful for the further design of neuronal chemical probes for the melanocortin receptors as well as in other receptor systems.

  11. Design of potent inhibitors of human RAD51 recombinase based on BRC motifs of BRCA2 protein: modeling and experimental validation of a chimera peptide.

    KAUST Repository

    Nomme, Julian; Renodon-Corniè re, Axelle; Asanomi, Yuya; Sakaguchi, Kazuyasu; Stasiak, Alicja Z; Stasiak, Andrzej; Norden, Bengt; Tran, Vinh; Takahashi, Masayuki

    2010-01-01

    We have previously shown that a 28-amino acid peptide derived from the BRC4 motif of BRCA2 tumor suppressor inhibits selectively human RAD51 recombinase (HsRad51). With the aim of designing better inhibitors for cancer treatment, we combined an in silico docking approach with in vitro biochemical testing to construct a highly efficient chimera peptide from eight existing human BRC motifs. We built a molecular model of all BRC motifs complexed with HsRad51 based on the crystal structure of the BRC4 motif-HsRad51 complex, computed the interaction energy of each residue in each BRC motif, and selected the best amino acid residue at each binding position. This analysis enabled us to propose four amino acid substitutions in the BRC4 motif. Three of these increased the inhibitory effect in vitro, and this effect was found to be additive. We thus obtained a peptide that is about 10 times more efficient in inhibiting HsRad51-ssDNA complex formation than the original peptide.

  12. Design of potent inhibitors of human RAD51 recombinase based on BRC motifs of BRCA2 protein: modeling and experimental validation of a chimera peptide.

    KAUST Repository

    Nomme, Julian

    2010-08-01

    We have previously shown that a 28-amino acid peptide derived from the BRC4 motif of BRCA2 tumor suppressor inhibits selectively human RAD51 recombinase (HsRad51). With the aim of designing better inhibitors for cancer treatment, we combined an in silico docking approach with in vitro biochemical testing to construct a highly efficient chimera peptide from eight existing human BRC motifs. We built a molecular model of all BRC motifs complexed with HsRad51 based on the crystal structure of the BRC4 motif-HsRad51 complex, computed the interaction energy of each residue in each BRC motif, and selected the best amino acid residue at each binding position. This analysis enabled us to propose four amino acid substitutions in the BRC4 motif. Three of these increased the inhibitory effect in vitro, and this effect was found to be additive. We thus obtained a peptide that is about 10 times more efficient in inhibiting HsRad51-ssDNA complex formation than the original peptide.

  13. Human peptide transporters

    DEFF Research Database (Denmark)

    Nielsen, Carsten Uhd; Brodin, Birger; Jørgensen, Flemming Steen

    2002-01-01

    Peptide transporters are epithelial solute carriers. Their functional role has been characterised in the small intestine and proximal tubules, where they are involved in absorption of dietary peptides and peptide reabsorption, respectively. Currently, two peptide transporters, PepT1 and PepT2, wh...

  14. Pb(II) and Hg(II) binding to $\\textit{de novo}$ designed proteins studied by $^{204m}$Pb- and $^{199m}$Hg-Perturbed Angular Correlation of $\\gamma$-rays (PAC) spectroscopy : Clues to heavy metal toxicity

    CERN Multimedia

    2002-01-01

    $\\textit{De novo}$ design of proteins combined with PAC spectroscopy offers a unique and powerful approach to the study of fundamental chemistry of heavy metal-protein interactions, and thus of the mechanisms underlying heavy metal toxicity. In this project we focus on Pb(II) and Hg(II) binding to designed three stranded coiled coil proteins with one or two binding sites, mimicking a variety of naturally occurring thiolate-rich metal ion binding sites in proteins. The $^{204m}$Pb- and $^{199m}$Hg-PAC experiments will complement data already recorded with EXAFS, NMR, UV-Vis and CD spectroscopies.

  15. Peptide dendrimers

    Czech Academy of Sciences Publication Activity Database

    Niederhafner, Petr; Šebestík, Jaroslav; Ježek, Jan

    2005-01-01

    Roč. 11, - (2005), 757-788 ISSN 1075-2617 R&D Projects: GA ČR(CZ) GA203/03/1362 Institutional research plan: CEZ:AV0Z40550506 Keywords : multiple antigen peptides * peptide dendrimers * synthetic vaccine * multipleantigenic peptides Subject RIV: CC - Organic Chemistry Impact factor: 1.803, year: 2005

  16. De novo FBXO11 mutations are associated with intellectual disability and behavioural anomalies.

    Science.gov (United States)

    Fritzen, Daniel; Kuechler, Alma; Grimmel, Mona; Becker, Jessica; Peters, Sophia; Sturm, Marc; Hundertmark, Hela; Schmidt, Axel; Kreiß, Martina; Strom, Tim M; Wieczorek, Dagmar; Haack, Tobias B; Beck-Wödl, Stefanie; Cremer, Kirsten; Engels, Hartmut

    2018-05-01

    Intellectual disability (ID) has an estimated prevalence of 1.5-2%. In most affected individuals, its genetic basis remains unclear. Whole exome sequencing (WES) studies have identified a multitude of novel causative gene defects and have shown that a large proportion of sporadic ID cases results from de novo mutations. Here, we present two unrelated individuals with similar clinical features and deleterious de novo variants in FBXO11 detected by WES. Individual 1, a 14-year-old boy, has mild ID as well as mild microcephaly, corrected cleft lip and alveolus, hyperkinetic disorder, mild brain atrophy and minor facial dysmorphism. WES detected a heterozygous de novo 1 bp insertion in the splice donor site of exon 3. Individual 2, a 3-year-old boy, showed ID and pre- and postnatal growth retardation, postnatal mild microcephaly, hyperkinetic and restless behaviour, as well as mild dysmorphism. WES detected a heterozygous de novo frameshift mutation. While ten individuals with ID and de novo variants in FBXO11 have been reported as part of larger studies, only one of the reports has some additional clinical data. Interestingly, the latter individual carries the identical mutation as our individual 2 and also displays ID, intrauterine growth retardation, microcephaly, behavioural anomalies, and dysmorphisms. Thus, we confirm deleterious de novo mutations in FBXO11 as a cause of ID and start the delineation of the associated clinical picture which may also comprise postnatal microcephaly or borderline small head size and behavioural anomalies.

  17. InverPep: A database of invertebrate antimicrobial peptides.

    Science.gov (United States)

    Gómez, Esteban A; Giraldo, Paula; Orduz, Sergio

    2017-03-01

    The aim of this work was to construct InverPep, a database specialised in experimentally validated antimicrobial peptides (AMPs) from invertebrates. AMP data contained in InverPep were manually curated from other databases and the scientific literature. MySQL was integrated with the development platform Laravel; this framework allows to integrate programming in PHP with HTML and was used to design the InverPep web page's interface. InverPep contains 18 separated fields, including InverPep code, phylum and species source, peptide name, sequence, peptide length, secondary structure, molar mass, charge, isoelectric point, hydrophobicity, Boman index, aliphatic index and percentage of hydrophobic amino acids. CALCAMPI, an algorithm to calculate the physicochemical properties of multiple peptides simultaneously, was programmed in PERL language. To date, InverPep contains 702 experimentally validated AMPs from invertebrate species. All of the peptides contain information associated with their source, physicochemical properties, secondary structure, biological activity and links to external literature. Most AMPs in InverPep have a length between 10 and 50 amino acids, a positive charge, a Boman index between 0 and 2 kcal/mol, and 30-50% hydrophobic amino acids. InverPep includes 33 AMPs not reported in other databases. Besides, CALCAMPI and statistical analysis of InverPep data is presented. The InverPep database is available in English and Spanish. InverPep is a useful database to study invertebrate AMPs and its information could be used for the design of new peptides. The user-friendly interface of InverPep and its information can be freely accessed via a web-based browser at http://ciencias.medellin.unal.edu.co/gruposdeinvestigacion/prospeccionydisenobiomoleculas/InverPep/public/home_en. Copyright © 2016 International Society for Chemotherapy of Infection and Cancer. Published by Elsevier Ltd. All rights reserved.

  18. Development and use of engineered peptide deformylase in chemoenzymatic peptide synthesis

    NARCIS (Netherlands)

    Di Toma, Claudia

    2012-01-01

    Deze thesis beschrijft het onderzoek naar potentieel van het gebruik van het peptide deformylase (PDF) in chemo enzymatische peptide synthese. PDF is geschikt voor selective N terminale deformylatie van bepaalde N-formyl-peptides zonder gelijktijdige hydrolyse van de peptide binding. Door de

  19. De novo transcriptome assembly associated with fumonisin production by the rice pathogen Fusarium fujikuroi

    Directory of Open Access Journals (Sweden)

    Keerthi S. Guruge

    2018-06-01

    Full Text Available The present study employed a next-generation sequencing method to assemble a de novo transcriptome database designed to distinguish gene expression changes exhibited by the fumonisin-producing fungus Fusarium fujikuroi when grown under ‘fumonisin-producing’ compared to ‘non-fumonisin-producing’ conditions. The raw data of this study have been deposited at DNA Data Bank of Japan (DDBJ under the accession ID DRA006146. Keywords: Fusarium fujikuroi, Fumonisin, Next-generation sequencing, Transcriptome, Gene-expression

  20. Spaced Seed Data Structures for De Novo Assembly

    Directory of Open Access Journals (Sweden)

    Inanç Birol

    2015-01-01

    Full Text Available De novo assembly of the genome of a species is essential in the absence of a reference genome sequence. Many scalable assembly algorithms use the de Bruijn graph (DBG paradigm to reconstruct genomes, where a table of subsequences of a certain length is derived from the reads, and their overlaps are analyzed to assemble sequences. Despite longer subsequences unlocking longer genomic features for assembly, associated increase in compute resources limits the practicability of DBG over other assembly archetypes already designed for longer reads. Here, we revisit the DBG paradigm to adapt it to the changing sequencing technology landscape and introduce three data structure designs for spaced seeds in the form of paired subsequences. These data structures address memory and run time constraints imposed by longer reads. We observe that when a fixed distance separates seed pairs, it provides increased sequence specificity with increased gap length. Further, we note that Bloom filters would be suitable to implicitly store spaced seeds and be tolerant to sequencing errors. Building on this concept, we describe a data structure for tracking the frequencies of observed spaced seeds. These data structure designs will have applications in genome, transcriptome and metagenome assemblies, and read error correction.

  1. The synergistic use of computation, chemistry and biology to discover novel peptide-based drugs: the time is right.

    Science.gov (United States)

    Audie, J; Boyd, C

    2010-01-01

    The case for peptide-based drugs is compelling. Due to their chemical, physical and conformational diversity, and relatively unproblematic toxicity and immunogenicity, peptides represent excellent starting material for drug discovery. Nature has solved many physiological and pharmacological problems through the use of peptides, polypeptides and proteins. If nature could solve such a diversity of challenging biological problems through the use of peptides, it seems reasonable to infer that human ingenuity will prove even more successful. And this, indeed, appears to be the case, as a number of scientific and methodological advances are making peptides and peptide-based compounds ever more promising pharmacological agents. Chief among these advances are powerful chemical and biological screening technologies for lead identification and optimization, methods for enhancing peptide in vivo stability, bioavailability and cell-permeability, and new delivery technologies. Other advances include the development and experimental validation of robust computational methods for peptide lead identification and optimization. Finally, scientific analysis, biology and chemistry indicate the prospect of designing relatively small peptides to therapeutically modulate so-called 'undruggable' protein-protein interactions. Taken together a clear picture is emerging: through the synergistic use of the scientific imagination and the computational, chemical and biological methods that are currently available, effective peptide therapeutics for novel targets can be designed that surpass even the proven peptidic designs of nature.

  2. The Sam-Sam interaction between Ship2 and the EphA2 receptor: design and analysis of peptide inhibitors.

    Science.gov (United States)

    Mercurio, Flavia Anna; Di Natale, Concetta; Pirone, Luciano; Iannitti, Roberta; Marasco, Daniela; Pedone, Emilia Maria; Palumbo, Rosanna; Leone, Marilisa

    2017-12-12

    The lipid phosphatase Ship2 represents a drug discovery target for the treatment of different diseases, including cancer. Its C-terminal sterile alpha motif domain (Ship2-Sam) associates with the Sam domain from the EphA2 receptor (EphA2-Sam). This interaction is expected to mainly induce pro-oncogenic effects in cells therefore, inhibition of the Ship2-Sam/EphA2-Sam complex may represent an innovative route to discover anti-cancer therapeutics. In the present work, we designed and analyzed several peptide sequences encompassing the interaction interface of EphA2-Sam for Ship2-Sam. Peptide conformational analyses and interaction assays with Ship2-Sam conducted through diverse techniques (CD, NMR, SPR and MST), identified a positively charged penta-amino acid native motif in EphA2-Sam, that once repeated three times in tandem, binds Ship2-Sam. NMR experiments show that the peptide targets the negatively charged binding site of Ship2-Sam for EphA2-Sam. Preliminary in vitro cell-based assays indicate that -at 50 µM concentration- it induces necrosis of PC-3 prostate cancer cells with more cytotoxic effect on cancer cells than on normal dermal fibroblasts. This work represents a pioneering study that opens further opportunities for the development of inhibitors of the Ship2-Sam/EphA2-Sam complex for therapeutic applications.

  3. Rapid and Weight-Independent Improvement of Glucose Tolerance Induced by a Peptide Designed to Elicit Apoptosis in Adipose Tissue Endothelium

    Science.gov (United States)

    Kim, Dong-Hoon; Sartor, Maureen A.; Bain, James R.; Sandoval, Darleen; Stevens, Robert D.; Medvedovic, Mario; Newgard, Christopher B.; Woods, Stephen C.; Seeley, Randy J.

    2012-01-01

    A peptide designed to induce apoptosis of endothelium in white adipose tissue (WAT) decreases adiposity. The goal of this work is to determine whether targeting of WAT endothelium results in impaired glucose regulation as a result of impaired WAT function. Glucose tolerance tests were performed on days 2 and 3 of treatment with vehicle (HF-V) or proapoptotic peptide (HF-PP) and mice pair-fed to HF-PP (HF-PF) in obese mice on a high-fat diet (HFD). Serum metabolic variables, including lipid profile, adipokines, individual fatty acids, and acylcarnitines, were measured. Microarray analysis was performed in epididymal fat of lean or obese mice treated with vehicle or proapoptotic peptide (PP). PP rapidly and potently improved glucose tolerance of obese mice in a weight- and food intake–independent manner. Serum insulin and triglycerides were decreased in HF-PP relative to HF-V. Levels of fatty acids and acylcarnitines were distinctive in HF-PP compared with HF-V or HF-PF. Microarray analysis in AT revealed that pathways involved in mitochondrial dysfunction, oxidative phosphorylation, and branched-chain amino acid degradation were changed by exposure to HFD and were reversed by PP administration. These studies suggest a novel role of the AT vasculature in glucose homeostasis and lipid metabolism. PMID:22733798

  4. Evolutionary combinatorial chemistry, a novel tool for SAR studies on peptide transport across the blood-brain barrier. Part 2. Design, synthesis and evaluation of a first generation of peptides.

    Science.gov (United States)

    Teixidó, Meritxell; Belda, Ignasi; Zurita, Esther; Llorà, Xavier; Fabre, Myriam; Vilaró, Senén; Albericio, Fernando; Giralt, Ernest

    2005-12-01

    The use of high-throughput methods in drug discovery allows the generation and testing of a large number of compounds, but at the price of providing redundant information. Evolutionary combinatorial chemistry combines the selection and synthesis of biologically active compounds with artificial intelligence optimization methods, such as genetic algorithms (GA). Drug candidates for the treatment of central nervous system (CNS) disorders must overcome the blood-brain barrier (BBB). This paper reports a new genetic algorithm that searches for the optimal physicochemical properties for peptide transport across the blood-brain barrier. A first generation of peptides has been generated and synthesized. Due to the high content of N-methyl amino acids present in most of these peptides, their syntheses were especially challenging due to over-incorporations, deletions and DKP formations. Distinct fragmentation patterns during peptide cleavage have been identified. The first generation of peptides has been studied by evaluation techniques such as immobilized artificial membrane chromatography (IAMC), a cell-based assay, log Poctanol/water calculations, etc. Finally, a second generation has been proposed. (c) 2005 European Peptide Society and John Wiley & Sons, Ltd.

  5. Peptide and protein delivery using new drug delivery systems.

    Science.gov (United States)

    Jain, Ashish; Jain, Aviral; Gulbake, Arvind; Shilpi, Satish; Hurkat, Pooja; Jain, Sanjay K

    2013-01-01

    Pharmaceutical and biotechnological research sorts protein drug delivery systems by importance based on their various therapeutic applications. The effective and potent action of the proteins/peptides makes them the drugs of choice for the treatment of numerous diseases. Major research issues in protein delivery include the stabilization of proteins in delivery devices and the design of appropriate target-specific protein carriers. Many efforts have been made for effective delivery of proteins/peptidal drugs through various routes of administrations for successful therapeutic effects. Nanoparticles made of biodegradable polymers such as poly lactic acid, polycaprolactone, poly(lactic-co-glycolic acid), the poly(fumaric-co-sebacic) anhydride chitosan, and modified chitosan, as well as solid lipids, have shown great potential in the delivery of proteins/peptidal drugs. Moreover, scientists also have used liposomes, PEGylated liposomes, niosomes, and aquasomes, among others, for peptidal drug delivery. They also have developed hydrogels and transdermal drug delivery systems for peptidal drug delivery. A receptor-mediated delivery system is another attractive strategy to overcome the limitation in drug absorption that enables the transcytosis of the protein across the epithelial barrier. Modification such as PEGnology is applied to various proteins and peptides of the desired protein and peptides also increases the circulating life, solubility and stability, pharmacokinetic properties, and antigenicity of protein. This review focuses on various approaches for effective protein/peptidal drug delivery, with special emphasis on insulin delivery.

  6. New Insights in the Design of Bioactive Peptides and Chelating Agents for Imaging and Therapy in Oncology

    Directory of Open Access Journals (Sweden)

    Anna Lucia Tornesello

    2017-08-01

    Full Text Available Many synthetic peptides have been developed for diagnosis and therapy of human cancers based on their ability to target specific receptors on cancer cell surface or to penetrate the cell membrane. Chemical modifications of amino acid chains have significantly improved the biological activity, the stability and efficacy of peptide analogues currently employed as anticancer drugs or as molecular imaging tracers. The stability of somatostatin, integrins and bombesin analogues in the human body have been significantly increased by cyclization and/or insertion of non-natural amino acids in the peptide sequences. Moreover, the overall pharmacokinetic properties of such analogues and others (including cholecystokinin, vasoactive intestinal peptide and neurotensin analogues have been improved by PEGylation and glycosylation. Furthermore, conjugation of those peptide analogues to new linkers and bifunctional chelators (such as AAZTA, TETA, TRAP, NOPO etc., produced radiolabeled moieties with increased half life and higher binding affinity to the cognate receptors. This review describes the most important and recent chemical modifications introduced in the amino acid sequences as well as linkers and new bifunctional chelators which have significantly improved the specificity and sensitivity of peptides used in oncologic diagnosis and therapy.

  7. New dendrimer - Peptide host - Guest complexes: Towards dendrimers as peptide carriers

    DEFF Research Database (Denmark)

    Boas, Ulrik; Sontjens, S.H.M.; Jensen, Knud Jørgen

    2002-01-01

    Adamantyl urea and adamantyl thiourea modified poly(propylene imine) dendrimers act as hosts for N-terminal tert-butoxycarbonyl (Boc)-protected peptides and form chloroform-soluble complexes. investigations with NMR spectroscopy show that the peptide is bound to the dendrimer by ionic interactions...... between the dendrimer outer shell tertiary amines and the C-terminal carboxylic acid of the peptide, and also through host-urea to peptide-amide hydrogen bonding. The hydrogen-bonding nature of the peptide dendrimer interactions was further confirmed by using Fourier transform IR spectroscopy, for which...... the NH- and CO-stretch signals of the peptide amide moieties shift towards lower wave-numbers upon complexation with the dendrimer. Spatial analysis of the complexes with NOESY spectroscopy generally shows close proximity of the N-terminal Boc group of the peptide to the peripheral adamantyl groups...

  8. Mimicking protein-protein interactions through peptide-peptide interactions: HIV-1 gp120 and CXCR4

    Directory of Open Access Journals (Sweden)

    Andrea eGross

    2013-09-01

    Full Text Available We have recently designed a soluble synthetic peptide that functionally mimics the HIV-1 coreceptor CXCR4, which is a chemokine receptor that belongs to the family of seven-transmembrane GPCRs. This CXCR4 mimetic peptide, termed CX4-M1, presents the three extracellular loops (ECLs of the receptor. In binding assays involving recombinant proteins, as well as in cellular infection assays, CX4-M1 was found to selectively recognize gp120 from HIV-1 strains that use CXCR4 for cell entry (X4 tropic HIV-1. Furthermore, anti-HIV-1 antibodies modulate this interaction in a molecular mechanism related to that of their impact on the gp120-CXCR4 interaction. We could now show that the selectivity of CX4-M1 pertains not only to gp120 from X4 tropic HIV-1, but also to synthetic peptides presenting the V3 loops of these gp120 proteins. The V3 loop is thought to be an essential part of the coreceptor binding site of gp120 that contacts the second ECL of the coreceptor. We were able to experimentally confirm this notion in binding assays using substitution analogs of CX4-M1 and the V3 loop peptides, respectively, as well as in cellular infection assays. These results indicate that interactions of the HIV-1 Env with coreceptors can be mimicked by synthetic peptides, which may be useful to explore these interactions at the molecular level in more detail.

  9. Design and synthesis of multiple antigenic peptides and their application for dengue diagnosis.

    Science.gov (United States)

    Rai, Reeta; Dubey, Sameer; Santosh, K V; Biswas, Ashutosh; Mehrotra, Vinit; Rao, D N

    2017-09-01

    Major difficulty in development of dengue diagnostics is availability of suitable antigens. To overcome this, we made an attempt to develop a peptide based diagnosis which offers significant advantage over other methods. With the help of in silico methods, two epitopes were selected from envelope protein and three from NS1 protein of dengue virus. These were synthesized in combination as three multiple antigenic peptides (MAPs). We have tested 157 dengue positive sera confirmed for NS1 antigen. MAP1 showed 96.81% sera positive for IgM and 68.15% positive for IgG. MAP2 detected 94.90% IgM and 59.23% IgG positive sera. MAP3 also detected 96.17% IgM and 59.87% IgG positive sera. To the best of our knowledge this is the first study describing the use of synthetic multiple antigenic peptides for the diagnosis of dengue infection. This study describes MAPs as a promising tool for the use in serodiagnosis of dengue. Copyright © 2017 International Alliance for Biological Standardization. Published by Elsevier Ltd. All rights reserved.

  10. A Brief Review of Computer-Assisted Approaches to Rational Design of Peptide Vaccines

    Directory of Open Access Journals (Sweden)

    Ashesh Nandy

    2016-05-01

    Full Text Available The growing incidences of new viral diseases and increasingly frequent viral epidemics have strained therapeutic and preventive measures; the high mutability of viral genes puts additional strains on developmental efforts. Given the high cost and time requirements for new drugs development, vaccines remain as a viable alternative, but there too traditional techniques of live-attenuated or inactivated vaccines have the danger of allergenic reactions and others. Peptide vaccines have, over the last several years, begun to be looked on as more appropriate alternatives, which are economically affordable, require less time for development and hold the promise of multi-valent dosages. The developments in bioinformatics, proteomics, immunogenomics, structural biology and other sciences have spurred the growth of vaccinomics where computer assisted approaches serve to identify suitable peptide targets for eventual development of vaccines. In this mini-review we give a brief overview of some of the recent trends in computer assisted vaccine development with emphasis on the primary selection procedures of probable peptide candidates for vaccine development.

  11. Derivatization of peptides as quaternary ammonium salts for sensitive detection by ESI-MS.

    Science.gov (United States)

    Cydzik, Marzena; Rudowska, Magdalena; Stefanowicz, Piotr; Szewczuk, Zbigniew

    2011-06-01

    A series of model peptides in the form of quaternary ammonium salts at the N-terminus was efficiently prepared by the solid-phase synthesis. Tandem mass spectrometric analysis of the peptide quaternary ammonium derivatives was shown to provide sequence confirmation and enhanced detection. We designed the 2-(1,4-diazabicyclo[2.2.2] octylammonium)acetyl quaternary ammonium group which does not suffer from neutral losses during MS/MS experiments. The presented quaternization of 1,4-diazabicyclo[2.2.2]octane (DABCO) by iodoacetylated peptides is relatively easy and compatible with standard solid-phase peptide synthesis. This methodology offers a novel sensitive approach to analyze peptides and other compounds. Copyright © 2011 European Peptide Society and John Wiley & Sons, Ltd.

  12. Peptide inhibition of human cytomegalovirus infection

    Directory of Open Access Journals (Sweden)

    Morris Cindy A

    2011-02-01

    M and 50 μM, respectively, and 60% at a concentration of 2.5 μM. While peptides 264-291 and 297-315, individually failed to inhibit viral infection, when combined, they showed 67% inhibition of HCMV infection at a concentration of 0.125 μM each. Conclusions Peptides designed to target putative fusogenic domains of gB provide a basis for the development of novel therapeutics that prevent HCMV infection.

  13. Morintides: cargo-free chitin-binding peptides from Moringa oleifera.

    Science.gov (United States)

    Kini, Shruthi G; Wong, Ka H; Tan, Wei Liang; Xiao, Tianshu; Tam, James P

    2017-03-31

    Hevein-like peptides are a family of cysteine-rich and chitin-binding peptides consisting of 29-45 amino acids. Their chitin-binding property is essential for plant defense against fungi. Based on the number of cysteine residues in their sequences, they are divided into three sub-families: 6C-, 8C- and 10C-hevein-like peptides. All three subfamilies contain a three-domain precursor comprising a signal peptide, a mature hevein-like peptide and a C-terminal domain comprising a hinge region with protein cargo in 8C- and 10C-hevein-like peptides. Here we report the isolation and characterization of two novel 8C-hevein-like peptides, designated morintides (mO1 and mO2), from the drumstick tree Moringa oleifera, a drought-resistant tree belonging to the Moringaceae family. Proteomic analysis revealed that morintides comprise 44 amino acid residues and are rich in cysteine, glycine and hydrophilic amino acid residues such as asparagine and glutamine. Morintides are resistant to thermal and enzymatic degradation, able to bind to chitin and inhibit the growth of phyto-pathogenic fungi. Transcriptomic analysis showed that they contain a three-domain precursor comprising an endoplasmic reticulum (ER) signal sequence, a mature peptide domain and a C-terminal domain. A striking feature distinguishing morintides from other 8C-hevein-like peptides is a short and protein-cargo-free C-terminal domain. Previously, a similar protein-cargo-free C-terminal domain has been observed only in ginkgotides, the 8C-hevein-like peptides from a gymnosperm Ginkgo biloba. Thus, morintides, with a cargo-free C-terminal domain, are a stand-alone class of 8C-hevein-like peptides from angiosperms. Our results expand the existing library of hevein-like peptides and shed light on molecular diversity within the hevein-like peptide family. Our work also sheds light on the anti-fungal activity and stability of 8C-hevein-like peptides.

  14. Software-aided approach to investigate peptide structure and metabolic susceptibility of amide bonds in peptide drugs based on high resolution mass spectrometry.

    Directory of Open Access Journals (Sweden)

    Tatiana Radchenko

    Full Text Available Interest in using peptide molecules as therapeutic agents due to high selectivity and efficacy is increasing within the pharmaceutical industry. However, most peptide-derived drugs cannot be administered orally because of low bioavailability and instability in the gastrointestinal tract due to protease activity. Therefore, structural modifications peptides are required to improve their stability. For this purpose, several in-silico software tools have been developed such as PeptideCutter or PoPS, which aim to predict peptide cleavage sites for different proteases. Moreover, several databases exist where this information is collected and stored from public sources such as MEROPS and ExPASy ENZYME databases. These tools can help design a peptide drug with increased stability against proteolysis, though they are limited to natural amino acids or cannot process cyclic peptides, for example. We worked to develop a new methodology to analyze peptide structure and amide bond metabolic stability based on the peptide structure (linear/cyclic, natural/unnatural amino acids. This approach used liquid chromatography / high resolution, mass spectrometry to obtain the analytical data from in vitro incubations. We collected experimental data for a set (linear/cyclic, natural/unnatural amino acids of fourteen peptide drugs and four substrate peptides incubated with different proteolytic media: trypsin, chymotrypsin, pepsin, pancreatic elastase, dipeptidyl peptidase-4 and neprilysin. Mass spectrometry data was analyzed to find metabolites and determine their structures, then all the results were stored in a chemically aware manner, which allows us to compute the peptide bond susceptibility by using a frequency analysis of the metabolic-liable bonds. In total 132 metabolites were found from the various in vitro conditions tested resulting in 77 distinct cleavage sites. The most frequent observed cleavage sites agreed with those reported in the literature. The

  15. Design of Decorated Self-Assembling Peptide Hydrogels as Architecture for Mesenchymal Stem Cells

    Directory of Open Access Journals (Sweden)

    Annj Zamuner

    2016-08-01

    Full Text Available Hydrogels from self-assembling ionic complementary peptides have been receiving a lot of interest from the scientific community as mimetic of the extracellular matrix that can offer three-dimensional supports for cell growth or can become vehicles for the delivery of stem cells, drugs or bioactive proteins. In order to develop a 3D “architecture” for mesenchymal stem cells, we propose the introduction in the hydrogel of conjugates obtained by chemoselective ligation between a ionic-complementary self-assembling peptide (called EAK and three different bioactive molecules: an adhesive sequence with 4 Glycine-Arginine-Glycine-Aspartic Acid-Serine-Proline (GRGDSP motifs per chain, an adhesive peptide mapped on h-Vitronectin and the growth factor Insulin-like Growth Factor-1 (IGF-1. The mesenchymal stem cell adhesion assays showed a significant increase in adhesion and proliferation for the hydrogels decorated with each of the synthesized conjugates; moreover, such functionalized 3D hydrogels support cell spreading and elongation, validating the use of this class of self-assembly peptides-based material as very promising 3D model scaffolds for cell cultures, at variance of the less realistic 2D ones. Furthermore, small amplitude oscillatory shear tests showed that the presence of IGF-1-conjugate did not alter significantly the viscoelastic properties of the hydrogels even though differences were observed in the nanoscale structure of the scaffolds obtained by changing their composition, ranging from long, well-defined fibers for conjugates with adhesion sequences to the compact and dense film for the IGF-1-conjugate.

  16. Catalysis by a de novo zinc-mediated protein interface: implications for natural enzyme evolution and rational enzyme engineering.

    Science.gov (United States)

    Der, Bryan S; Edwards, David R; Kuhlman, Brian

    2012-05-08

    Here we show that a recent computationally designed zinc-mediated protein interface is serendipitously capable of catalyzing carboxyester and phosphoester hydrolysis. Although the original motivation was to design a de novo zinc-mediated protein-protein interaction (called MID1-zinc), we observed in the homodimer crystal structure a small cleft and open zinc coordination site. We investigated if the cleft and zinc site at the designed interface were sufficient for formation of a primitive active site that can perform hydrolysis. MID1-zinc hydrolyzes 4-nitrophenyl acetate with a rate acceleration of 10(5) and a k(cat)/K(M) of 630 M(-1) s(-1) and 4-nitrophenyl phosphate with a rate acceleration of 10(4) and a k(cat)/K(M) of 14 M(-1) s(-1). These rate accelerations by an unoptimized active site highlight the catalytic power of zinc and suggest that the clefts formed by protein-protein interactions are well-suited for creating enzyme active sites. This discovery has implications for protein evolution and engineering: from an evolutionary perspective, three-coordinated zinc at a homodimer interface cleft represents a simple evolutionary path to nascent enzymatic activity; from a protein engineering perspective, future efforts in de novo design of enzyme active sites may benefit from exploring clefts at protein interfaces for active site placement.

  17. Nanoparticles and peptides: a fruitful liaison for biomimetic catalysis.

    Science.gov (United States)

    Stodulski, Maciej; Gulder, Tanja

    2012-11-05

    Inspired by nature: self-assembled peptide nanoparticles have been designed that accelerate ester hydrolysis (see picture; Cbz=carbobenzyloxy, NP=p-NO(2)-C(6) H(4)). The concerted interplay of the multivalent surface with the catalytically active peptide and the substrate at the same time combines several aspects decisive for the catalyst's efficiency, a property characteristic of enzymes. Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  18. Sequencing Lys-N Proteolytic Peptides by ESI and MALDI Tandem Mass Spectrometry

    Science.gov (United States)

    Dupré, Mathieu; Cantel, Sonia; Verdié, Pascal; Martinez, Jean; Enjalbal, Christine

    2011-02-01

    In this study, we explored the MS/MS behavior of various synthetic peptides that possess a lysine residue at the N-terminal position. These peptides were designed to mimic peptides produced upon proteolysis by the Lys-N enzyme, a metalloendopeptidase issued from a Japanese fungus Grifola frondosa that was recently investigated in proteomic studies as an alternative to trypsin digestion, as a specific cleavage at the amide X-Lys chain is obtained that provides N-terminal lysine peptide fragments. In contrast to tryptic peptides exhibiting a lysine or arginine residue solely at the C-terminal position, and are thus devoid of such basic amino acids within the sequence, these Lys-N proteolytic peptides can contain the highly basic arginine residue anywhere within the peptide chain. The fragmentation patterns of such sequences with the ESI-QqTOF and MALDI-TOF/TOF mass spectrometers commonly used in proteomic bottom-up experiments were investigated.

  19. Peptides and polypeptides as scaffolds for optoelectronics and biomaterials applications

    Science.gov (United States)

    Charati, Manoj B.

    Peptides and polypeptides are emerging as a new class of biomaterials due to their unique structural, physiochemical, mechanical, and biological properties. The development of peptide and protein-based biomaterials is driven by the convergence of convenient techniques for peptide/protein engineering and its importance in applications as smart biomaterials. The thesis is divided in two parts; the first part highlights the importance of incorporation of non-natural amino acids into peptides and proteins. In particular, incorporation on p-bromophenylalanine in short alpha-helical peptide templates to control the association of chromophores is discussed. In the second part, design of a multi-component, biocompatible polypeptide with superior elasticity is discussed. Part 1. Novel peptide templates to control association of chromophores. Tailor made peptide and protein materials have many versatile applications, as both conformation and functional group position can be controlled. Such control may have intriguing applications in the development of hybrid materials for electroactive applications. A critical need in fabricating devices from organic semiconducting materials is to achieve control over the conformation and distance between two conjugated chains. Controlling chromophore spacing and orientation with required precision over nanometer length scale poses a greater challenge. Here we propose a peptide based template to control the alignment of the methylstilbene and Oxa-PPV chromophores with desired orientations and spacing. The hybrid peptides were characterized via CD, exciton coupled CD, 1H NMR and photoluminescence experiments. It is observed that slight change in the orientation of molecules has pronounced effect on the photo-physical behavior of the molecules. Characterization of the hybrid peptides via circular dichroism (CD) confirmed the helical character of the designed peptides and indicated that inclusion of non-natural amino acids has significant

  20. Amide I SFG Spectral Line Width Probes the Lipid-Peptide and Peptide-Peptide Interactions at Cell Membrane In Situ and in Real Time.

    Science.gov (United States)

    Zhang, Baixiong; Tan, Junjun; Li, Chuanzhao; Zhang, Jiahui; Ye, Shuji

    2018-06-13

    The balance of lipid-peptide and peptide-peptide interactions at cell membrane is essential to a large variety of cellular processes. In this study, we have experimentally demonstrated for the first time that sum frequency generation vibrational spectroscopy can be used to probe the peptide-peptide and lipid-peptide interactions in cell membrane in situ and in real time by determination of the line width of amide I band of protein backbone. Using a "benchmark" model of α-helical WALP23, it is found that the dominated lipid-peptide interaction causes a narrow line width of the amide I band, whereas the peptide-peptide interaction can markedly broaden the line width. When WALP23 molecules insert into the lipid bilayer, a quite narrow line width of the amide I band is observed because of the lipid-peptide interaction. In contrast, when the peptide lies down on the bilayer surface, the line width of amide I band becomes very broad owing to the peptide-peptide interaction. In terms of the real-time change in the line width, the transition from peptide-peptide interaction to lipid-peptide interaction is monitored during the insertion of WALP23 into 1,2-dipalmitoyl- sn-glycero-3-phospho-(1'- rac-glycerol) (DPPG) lipid bilayer. The dephasing time of a pure α-helical WALP23 in 1-palmitoyl-2-oleoyl- sn-glycero-3-phospho-(1'- rac-glycerol) and DPPG bilayer is determined to be 2.2 and 0.64 ps, respectively. The peptide-peptide interaction can largely accelerate the dephasing time.

  1. De novo design of alpha-amylase inhibitor: A small linear mimetic of macromolecular proteinaceous ligands

    Czech Academy of Sciences Publication Activity Database

    Marešová, Lucie; Pavlík, Manfred; Horn, Martin; Mareš, Michael

    2005-01-01

    Roč. 12, č. 12 (2005), 1349-1357 ISSN 1074-5521 R&D Projects: GA ČR(CZ) GP203/02/P081; GA MŠk(CZ) OC D16.001 Institutional research plan: CEZ:AV0Z40550506 Keywords : amylase * peptide inhibitor * combinatorial chemistry Subject RIV: CE - Biochemistry Impact factor: 6.138, year: 2005

  2. Design, synthesis and in vitro evaluation of heterobivalent peptidic radioligands targeting both GRP- and VPAC1-Receptors concomitantly overexpressed on various malignancies - Is the concept feasible?

    Science.gov (United States)

    Lindner, Simon; Fiedler, Luise; Wängler, Björn; Bartenstein, Peter; Schirrmacher, Ralf; Wängler, Carmen

    2018-05-29

    Radiolabeled heterobivalent peptidic ligands (HBPLs), being able to address different receptors, are highly interesting tumor imaging agents as they can offer multiple advantages over monovalent peptide receptor ligands. However, few examples of radiolabeled HBPLs have been described so far. One promising approach is the combination of gastrin-releasing peptide receptor (GRPR)- and vasoactive intestinal peptide receptor subtype 1 (VPAC 1 R)-targeting peptides into one single radioligand since gastrinomas, prostate and breast cancer have been shown to concomitantly or complementarily overexpress both receptors. Here we report the design and synthesis of different HBPLs, comprising a GRPR-binding (BBN 7-14 ) and a VPAC 1 R-targeting (PACAP-27) peptide. The heterodimers were varied with regard to the distance between the peptide binders and the steric rigidity of the systems. We radiolabeled the HBPLs 19-23 as well as their monomeric reference standards 26 and 27 with 68 Ga, achieving radiochemical yields and purities of 95-99% and non-optimized molar activities of 25-61 GBq/μmol. We tested the stability of the radioligands and further evaluated them in vitro regarding their uptake in different prostate carcinoma cell lines (PC-3, DU-145 and VCaP cells). We found that the heterobivalent substances [ 68 Ga]19 - [ 68 Ga]23 showed comparable uptakes into the tumor cells to those of the respective monomers [ 68 Ga]26 and [ 68 Ga]27, indicating that both peptides are still able to address their target receptors. Furthermore, the obtained results indicate that in case of overall low receptor densities, heterobivalent peptides surpass peptide monomers in tumor cell uptake. Most importantly, it could be shown by blocking studies that both peptide parts of the HBPL [ 68 Ga]19 contributed to tumor cell uptake in VCaP cells, expressing both receptor types. Thus, we describe here the first examples of HBPLs being able to address the GRPR as well as the VPAC 1 R and have the

  3. De novo synthesis of adenine nucleotides in different skeletal muscle fiber types

    International Nuclear Information System (INIS)

    Tullson, P.C.; John-Alder, H.B.; Hood, D.A.; Terjung, R.L.

    1988-01-01

    Management of adenine nucleotide catabolism differs among skeletal muscle fiber types. This study evaluated whether there are corresponding differences in the rates of de novo synthesis of adenine nucleotide among fiber type sections of skeletal muscle using an isolated perfused rat hindquarter preparation. Label incorporation into adenine nucleotides from the [1-14C]glycine precursor was determined and used to calculate synthesis rates based on the intracellular glycine specific radioactivity. Results show that intracellular glycine is closely related to the direct precursor pool. Rates of de novo synthesis were highest in fast-twitch red muscle (57.0 +/- 4.0, 58.2 +/- 4.4 nmol.h-1.g-1; deep red gastrocnemius and vastus lateralis), relatively high in slow-twitch red muscle (47.0 +/- 3.1; soleus), and low in fast-twitch white muscle (26.1 +/- 2.0 and 21.6 +/- 2.3; superficial white gastrocnemius and vastus lateralis). Rates for four mixed muscles were intermediate, ranging between 32.3 and 37.3. Specific de novo synthesis rates exhibited a strong correlation (r = 0.986) with muscle section citrate synthase activity. Turnover rates (de novo synthesis rate/adenine nucleotide pool size) were highest in high oxidative muscle (0.82-1.06%/h), lowest in low oxidative muscle (0.30-0.35%/h), and intermediate in mixed muscle (0.44-0.55%/h). Our results demonstrate that differences in adenine nucleotide management among fiber types extends to the process of de novo adenine nucleotide synthesis

  4. Wegener's granulomatosis occurring de novo during pregnancy.

    Science.gov (United States)

    Alfhaily, F; Watts, R; Leather, A

    2009-01-01

    Wegener's granulomatosis (WG) is rarely diagnosed during the reproductive years and uncommonly manifests for the first time during pregnancy. We report a case of de novo WG presenting at 30 weeks gestation with classical symptoms of WG (ENT, pulmonary). The diagnosis was confirmed by radiological, laboratory, and histological investigations. With a multidisciplinary approach, she had a successful vaginal delivery of a healthy baby. She was treated successfully by a combination of steroids, azathioprine and intravenous immunoglobulin in the active phase of disease for induction of remission and by azathioprine and steroids for maintenance of remission. The significant improvement in her symptoms allowed us to continue her pregnancy to 37 weeks when delivery was electively induced. Transplacental transmission of PR3-ANCA occurred but the neonate remained well. This case of de novo WG during pregnancy highlights the seriousness of this disease and the challenge in management of such patients.

  5. A Quantitative Tool to Distinguish Isobaric Leucine and Isoleucine Residues for Mass Spectrometry-Based De Novo Monoclonal Antibody Sequencing

    Science.gov (United States)

    Poston, Chloe N.; Higgs, Richard E.; You, Jinsam; Gelfanova, Valentina; Hale, John E.; Knierman, Michael D.; Siegel, Robert; Gutierrez, Jesus A.

    2014-07-01

    De novo sequencing by mass spectrometry (MS) allows for the determination of the complete amino acid (AA) sequence of a given protein based on the mass difference of detected ions from MS/MS fragmentation spectra. The technique relies on obtaining specific masses that can be attributed to characteristic theoretical masses of AAs. A major limitation of de novo sequencing by MS is the inability to distinguish between the isobaric residues leucine (Leu) and isoleucine (Ile). Incorrect identification of Ile as Leu or vice versa often results in loss of activity in recombinant antibodies. This functional ambiguity is commonly resolved with costly and time-consuming AA mutation and peptide sequencing experiments. Here, we describe a set of orthogonal biochemical protocols, which experimentally determine the identity of Ile or Leu residues in monoclonal antibodies (mAb) based on the selectivity that leucine aminopeptidase shows for n-terminal Leu residues and the cleavage preference for Leu by chymotrypsin. The resulting observations are combined with germline frequencies and incorporated into a logistic regression model, called Predictor for Xle Sites (PXleS) to provide a statistical likelihood for the identity of Leu at an ambiguous site. We demonstrate that PXleS can generate a probability for an Xle site in mAbs with 96% accuracy. The implementation of PXleS precludes the expression of several possible sequences and, therefore, reduces the overall time and resources required to go from spectra generation to a biologically active sequence for a mAb when an Ile or Leu residue is in question.

  6. Design of a peptidic turn with high affinity for HgII

    DEFF Research Database (Denmark)

    Pires, Sara; Habjanic, Jelena; Sezer, Murat

    2012-01-01

    A four amino acid peptide containing the ß-turn template dPro-Pro in the middle and two cysteines (Cys) in the terminal positions (CdPPC) has been synthesized and its mercury(II) coordination properties studied using different spectroscopic methods. The UV-vis, CD, (199m)Hg PAC, and Raman...... spectroscopic studies indicate the binding of Hg(II) to the two Cys, forming the dithiolatemercury(II) complex Hg(CdPPC). Electrospray ionization mass spectrometry corroborates the 1:1 complex formation. A log K = 40.0 was determined for the formation constant of the Hg(CdPPC) complex using competition...... potentiometric studies. Replacement of the dPro-Pro motif by a Pro-Pro unit generated a peptide (CPPC) capable of forming a similar species [Hg(CPPC)] but showing a lower affinity for Hg(II) (at least 3-3.5 orders of magnitude lower). The introduction of the dPro-Pro motif is crucial to induce the folding...

  7. Taylor Dispersion Analysis as a promising tool for assessment of peptide-peptide interactions.

    Science.gov (United States)

    Høgstedt, Ulrich B; Schwach, Grégoire; van de Weert, Marco; Østergaard, Jesper

    2016-10-10

    Protein-protein and peptide-peptide (self-)interactions are of key importance in understanding the physiochemical behavior of proteins and peptides in solution. However, due to the small size of peptide molecules, characterization of these interactions is more challenging than for proteins. In this work, we show that protein-protein and peptide-peptide interactions can advantageously be investigated by measurement of the diffusion coefficient using Taylor Dispersion Analysis. Through comparison to Dynamic Light Scattering it was shown that Taylor Dispersion Analysis is well suited for the characterization of protein-protein interactions of solutions of α-lactalbumin and human serum albumin. The peptide-peptide interactions of three selected peptides were then investigated in a concentration range spanning from 0.5mg/ml up to 80mg/ml using Taylor Dispersion Analysis. The peptide-peptide interactions determination indicated that multibody interactions significantly affect the PPIs at concentration levels above 25mg/ml for the two charged peptides. Relative viscosity measurements, performed using the capillary based setup applied for Taylor Dispersion Analysis, showed that the viscosity of the peptide solutions increased with concentration. Our results indicate that a viscosity difference between run buffer and sample in Taylor Dispersion Analysis may result in overestimation of the measured diffusion coefficient. Thus, Taylor Dispersion Analysis provides a practical, but as yet primarily qualitative, approach to assessment of the colloidal stability of both peptide and protein formulations. Copyright © 2016 Elsevier B.V. All rights reserved.

  8. Komodo dragon-inspired synthetic peptide DRGN-1 promotes wound-healing of a mixed-biofilm infected wound

    OpenAIRE

    M.C. Chung, Ezra; Dean, Scott N.; Propst, Crystal N.; Bishop, Barney M.; van Hoek, Monique L.

    2017-01-01

    Cationic antimicrobial peptides are multifunctional molecules that have a high potential as therapeutic agents. We have identified a histone H1-derived peptide from the Komodo dragon (Varanus komodoensis), called VK25. Using this peptide as inspiration, we designed a synthetic peptide called DRGN-1. We evaluated the antimicrobial and anti-biofilm activity of both peptides against Pseudomonas aeruginosa and Staphylococcus aureus. DRGN-1, more than VK25, exhibited potent antimicrobial and anti-...

  9. Peptide-oligonucleotide conjugates as nanoscale building blocks for assembly of an artificial three-helix protein mimic

    DEFF Research Database (Denmark)

    Lou, Chenguang; Martos-Maldonado, Manuel C.; Madsen, Charlotte Stahl

    2016-01-01

    Peptide-based structures can be designed to yield artificial proteins with specific folding patterns and functions. Template-based assembly of peptide units is one design option, but the use of two orthogonal self-assembly principles, oligonucleotide triple helix and a coiled coil protein domain ...

  10. De novo ORFs in Drosophila are important to organismal fitness and evolved rapidly from previously non-coding sequences.

    Directory of Open Access Journals (Sweden)

    Josephine A Reinhardt

    Full Text Available How non-coding DNA gives rise to new protein-coding genes (de novo genes is not well understood. Recent work has revealed the origins and functions of a few de novo genes, but common principles governing the evolution or biological roles of these genes are unknown. To better define these principles, we performed a parallel analysis of the evolution and function of six putatively protein-coding de novo genes described in Drosophila melanogaster. Reconstruction of the transcriptional history of de novo genes shows that two de novo genes emerged from novel long non-coding RNAs that arose at least 5 MY prior to evolution of an open reading frame. In contrast, four other de novo genes evolved a translated open reading frame and transcription within the same evolutionary interval suggesting that nascent open reading frames (proto-ORFs, while not required, can contribute to the emergence of a new de novo gene. However, none of the genes arose from proto-ORFs that existed long before expression evolved. Sequence and structural evolution of de novo genes was rapid compared to nearby genes and the structural complexity of de novo genes steadily increases over evolutionary time. Despite the fact that these genes are transcribed at a higher level in males than females, and are most strongly expressed in testes, RNAi experiments show that most of these genes are essential in both sexes during metamorphosis. This lethality suggests that protein coding de novo genes in Drosophila quickly become functionally important.

  11. Persistent hyperthyroidism and de novo Graves' ophthalmopathy after total thyroidectomy.

    Science.gov (United States)

    Tay, Wei Lin; Loh, Wann Jia; Lee, Lianne Ai Ling; Chng, Chiaw Ling

    2017-01-01

    We report a patient with Graves' disease who remained persistently hyperthyroid after a total thyroidectomy and also developed de novo Graves' ophthalmopathy 5 months after surgery. She was subsequently found to have a mature cystic teratoma containing struma ovarii after undergoing a total hysterectomy and salpingo-oophorectomy for an incidental ovarian lesion. It is important to investigate for other causes of primary hyperthyroidism when thyrotoxicosis persists after total thyroidectomy.TSH receptor antibody may persist after total thyroidectomy and may potentially contribute to the development of de novo Graves' ophthalmopathy.

  12. A therapeutic exploratory study to determine the efficacy and safety of calcineurin-inhibitor-free de-novo immunosuppression after liver transplantation: CILT

    Directory of Open Access Journals (Sweden)

    Lorf Thomas

    2010-04-01

    Full Text Available Abstract Background Immunosuppression with calcineurin inhibitors (CNI increases the risk of renal dysfunction after orthotopic liver transplantation (OLT. Controlled trials have shown improvement of renal function in patients that received delayed and/or reduced-dose CNI after OLT. Delaying immunosuppression with CNI in combination with induction therapy does not increase the risk of acute rejection but reduces the incidence of acute renal dysfunction. Based on this clinical data this study protocol was designed to assess the efficacy and safety of calcineurin-inhibitor-free de-novo immunosuppression after liver transplantation. Methods/Design A prospective therapeutic exploratory, non-placebo controlled, two stage monocenter trial in a total of 29 liver transplant patients was designed to assess the safety and efficacy of de-novo CNI-free immunosuppression with basiliximab, mycophenolate sodium, prednisolone and everolimus. The primary endpoint is the rate of steroid resistant rejections. Secondary endpoints are the incidence of acute rejection, kidney function (assessed by incidence and duration of renal replacement therapy, incidence of chronic renal failure, and measurement glomerular filtration rate, liver allograft function (assessed by measurement of AST, ALT, total bilirubin, AP, GGT, treatment failure, (i. e., re-introduction of CNI, incidence of adverse events, and mortality up to one year after OLT. Discussion This prospective, two-stage, single-group pilot study represents an intermediate element of the research chain. If the data of the phase II study corroborates safety of de-novo CNI-free immunosuppressive regimen this should be confirmed in a randomized, prospective, controlled double-blinded clinical trial. The exploratory data from this trial may then also facilitate the design (e. g. sample size calculation of this phase III trial. Trial registration number NCT00890253 (clinicaltrials.gov

  13. Antitumor activity of novel chimeric peptides derived from cyclinD/CDK4 and the protein transduction domain 4.

    Science.gov (United States)

    Wang, Haili; Chen, Xi; Chen, Yanping; Sun, Lei; Li, Guodong; Zhai, Mingxia; Zhai, Wenjie; Kang, Qiaozhen; Gao, Yanfeng; Qi, Yuanming

    2013-02-01

    CyclinD1/CDK4 and cyclinD3/CDK4 complexes are key regulators of the cell progression and therefore constitute promising targets for the design of anticancer agents. In the present study, the key peptide motifs were selected from these two complexes. Chimeric peptides with these peptides conjugated to the protein transduction domain 4 (PTD4) were designed and synthesized. The chimeric peptides, PTD4-D1, PTD4-D3, PTD4-K4 exhibited significant anti-proliferation effects on cancer cell lines. These peptides could compete with the cyclinD/CDK4 complex and induce the G1/S phase arrest and apoptosis of cancer cells. In the tumor challenge experiment, these peptides showed potent antitumor effects with no significant side effects. Our results suggested that these peptides could be served as novel leading compounds with potent antitumor activity.

  14. Elucidation of Peptide-Directed Palladium Surface Structure for Biologically Tunable Nanocatalysts

    Energy Technology Data Exchange (ETDEWEB)

    Bedford, Nicholas M.; Ramezani-Dakhel, Hadi; Slocik, Joseph M.; Briggs, Beverly D.; Ren, Yang; Frenkel, Anatoly I.; Petkov, Valeri; Heinz, Hendrik; Naik, Rajesh R.; Knecht, Mark R.

    2015-05-01

    Peptide-enabled synthesis of inorganic nanostructures represents an avenue to access catalytic materials with tunable and optimized properties. This is achieved via peptide complexity and programmability that is missing in traditional ligands for catalytic nanomaterials. Unfortunately, there is limited information available to correlate peptide sequence to particle structure and catalytic activity to date. As such, the application of peptide-enabled nanocatalysts remains limited to trial and error approaches. In this paper, a hybrid experimental and computational approach is introduced to systematically elucidate biomolecule-dependent structure/function relationships for peptide-capped Pd nanocatalysts. Synchrotron X-ray techniques were used to uncover substantial particle surface structural disorder, which was dependent upon the amino acid sequence of the peptide capping ligand. Nanocatalyst configurations were then determined directly from experimental data using reverse Monte Carlo methods and further refined using molecular dynamics simulation, obtaining thermodynamically stable peptide-Pd nanoparticle configurations. Sequence-dependent catalytic property differences for C-C coupling and olefin hydrogenation were then eluddated by identification of the catalytic active sites at the atomic level and quantitative prediction of relative reaction rates. This hybrid methodology provides a clear route to determine peptide-dependent structure/function relationships, enabling the generation of guidelines for catalyst design through rational tailoring of peptide sequences

  15. Mass Spectrometry Analysis Coupled with de novo Sequencing Reveals Amino Acid Substitutions in Nucleocapsid Protein from Influenza A Virus

    Directory of Open Access Journals (Sweden)

    Zijian Li

    2014-02-01

    Full Text Available Amino acid substitutions in influenza A virus are the main reasons for both antigenic shift and virulence change, which result from non-synonymous mutations in the viral genome. Nucleocapsid protein (NP, one of the major structural proteins of influenza virus, is responsible for regulation of viral RNA synthesis and replication. In this report we used LC-MS/MS to analyze tryptic digestion of nucleocapsid protein of influenza virus (A/Puerto Rico/8/1934 H1N1, which was isolated and purified by SDS poly-acrylamide gel electrophoresis. Thus, LC-MS/MS analyses, coupled with manual de novo sequencing, allowed the determination of three substituted amino acid residues R452K, T423A and N430T in two tryptic peptides. The obtained results provided experimental evidence that amino acid substitutions resulted from non-synonymous gene mutations could be directly characterized by mass spectrometry in proteins of RNA viruses such as influenza A virus.

  16. Selecting Superior De Novo Transcriptome Assemblies: Lessons Learned by Leveraging the Best Plant Genome.

    Directory of Open Access Journals (Sweden)

    Loren A Honaas

    Full Text Available Whereas de novo assemblies of RNA-Seq data are being published for a growing number of species across the tree of life, there are currently no broadly accepted methods for evaluating such assemblies. Here we present a detailed comparison of 99 transcriptome assemblies, generated with 6 de novo assemblers including CLC, Trinity, SOAP, Oases, ABySS and NextGENe. Controlled analyses of de novo assemblies for Arabidopsis thaliana and Oryza sativa transcriptomes provide new insights into the strengths and limitations of transcriptome assembly strategies. We find that the leading assemblers generate reassuringly accurate assemblies for the majority of transcripts. At the same time, we find a propensity for assemblers to fail to fully assemble highly expressed genes. Surprisingly, the instance of true chimeric assemblies is very low for all assemblers. Normalized libraries are reduced in highly abundant transcripts, but they also lack 1000s of low abundance transcripts. We conclude that the quality of de novo transcriptome assemblies is best assessed through consideration of a combination of metrics: 1 proportion of reads mapping to an assembly 2 recovery of conserved, widely expressed genes, 3 N50 length statistics, and 4 the total number of unigenes. We provide benchmark Illumina transcriptome data and introduce SCERNA, a broadly applicable modular protocol for de novo assembly improvement. Finally, our de novo assembly of the Arabidopsis leaf transcriptome revealed ~20 putative Arabidopsis genes lacking in the current annotation.

  17. Role of de novo biosynthesis in ecosystem scale monoterpene emissions from a boreal Scots pine forest

    Directory of Open Access Journals (Sweden)

    R. Taipale

    2011-08-01

    Full Text Available Monoterpene emissions from Scots pine have traditionally been assumed to originate as evaporation from specialized storage pools. More recently, the significance of de novo emissions, originating directly from monoterpene biosynthesis, has been recognized. To study the role of biosynthesis at the ecosystem scale, we measured monoterpene emissions from a Scots pine dominated forest in southern Finland using the disjunct eddy covariance method combined with proton transfer reaction mass spectrometry. The interpretation of the measurements was based on a correlation analysis and a hybrid emission algorithm describing both de novo and pool emissions. During the measurement period May–August 2007, the monthly medians of daytime emissions were 200, 290, 180, and 200 μg m−2 h−1. The emissions were partly light dependent, probably due to de novo biosynthesis. The emission potential for both de novo and pool emissions exhibited a decreasing summertime trend. The ratio of the de novo emission potential to the total emission potential varied between 30 % and 46 %. Although the monthly changes were not significant, the ratio always differed statistically from zero, suggesting that the role of de novo biosynthesis was observable. Given the uncertainties in this study, we conclude that more accurate estimates of the contribution of de novo emissions are required for improving monoterpene emission algorithms for Scots pine dominated forests.

  18. Functional Hybrid Biomaterials based on Peptide-Polymer Conjugates for Nanomedicine

    Science.gov (United States)

    Shu, Jessica Yo

    The focus of this dissertation is the design, synthesis and characterization of hybrid functional biomaterials based on peptide-polymer conjugates for nanomedicine. Generating synthetic materials with properties comparable to or superior than those found in nature has been a "holy grail" for the materials community. Man-made materials are still rather simplistic when compared to the chemical and structural complexity of a cell. Peptide-polymer conjugates have the potential to combine the advantages of the biological and synthetic worlds---that is they can combine the precise chemical structure and diverse functionality of biomolecules with the stability and processibility of synthetic polymers. As a new family of soft matter, they may lead to materials with novel properties that have yet to be realized with either of the components alone. In order for peptide-polymer conjugates to reach their full potential as useful materials, the structure and function of the peptide should be maintained upon polymer conjugation. The success in achieving desirable, functional assemblies relies on fundamentally understanding the interactions between each building block and delicately balancing and manipulating these interactions to achieve targeted assemblies without interfering with designed structures and functionalities. Such fundamental studies of peptide-polymer interactions were investigated as the nature of the polymer (hydrophilic vs. hydrophobic) and the site of its conjugation (end-conjugation vs. side-conjugation) were varied. The fundamental knowledge gained was then applied to the design of amphiphiles that self-assemble to form stable functional micelles. The micelles exhibited exceptional monodispersity and long-term stability, which is atypical of self-assembled systems. Thus such micelles based on amphiphilic peptide-polymer conjugates may meet many current demands in nanomedicine, in particular for drug delivery of hydrophobic anti-cancer therapeutics. Lastly

  19. Probabilistic consensus scoring improves tandem mass spectrometry peptide identification.

    Science.gov (United States)

    Nahnsen, Sven; Bertsch, Andreas; Rahnenführer, Jörg; Nordheim, Alfred; Kohlbacher, Oliver

    2011-08-05

    Database search is a standard technique for identifying peptides from their tandem mass spectra. To increase the number of correctly identified peptides, we suggest a probabilistic framework that allows the combination of scores from different search engines into a joint consensus score. Central to the approach is a novel method to estimate scores for peptides not found by an individual search engine. This approach allows the estimation of p-values for each candidate peptide and their combination across all search engines. The consensus approach works better than any single search engine across all different instrument types considered in this study. Improvements vary strongly from platform to platform and from search engine to search engine. Compared to the industry standard MASCOT, our approach can identify up to 60% more peptides. The software for consensus predictions is implemented in C++ as part of OpenMS, a software framework for mass spectrometry. The source code is available in the current development version of OpenMS and can easily be used as a command line application or via a graphical pipeline designer TOPPAS.

  20. A de novo designed 11 kDa polypeptide: model for amyloidogenic intrinsically disordered proteins.

    Science.gov (United States)

    Topilina, Natalya I; Ermolenkov, Vladimir V; Sikirzhytski, Vitali; Higashiya, Seiichiro; Lednev, Igor K; Welch, John T

    2010-07-01

    A de novo polypeptide GH(6)[(GA)(3)GY(GA)(3)GE](8)GAH(6) (YE8) has a significant number of identical weakly interacting beta-strands with the turns and termini functionalized by charged amino acids to control polypeptide folding and aggregation. YE8 exists in a soluble, disordered form at neutral pH but is responsive to changes in pH and ionic strength. The evolution of YE8 secondary structure has been successfully quantified during all stages of polypeptide fibrillation by deep UV resonance Raman (DUVRR) spectroscopy combined with other morphological, structural, spectral, and tinctorial characterization. The YE8 folding kinetics at pH 3.5 are strongly dependent on polypeptide concentration with a lag phase that can be eliminated by seeding with a solution of folded fibrillar YE8. The lag phase of polypeptide folding is concentration dependent leading to the conclusion that beta-sheet folding of the 11-kDa amyloidogenic polypeptide is completely aggregation driven.

  1. Evaluation of Novel Antimicrobial Peptides as Topical Anti-Infectives with Broad-Spectrum Activity against Combat-Related Bacterial and Fungal Wound Infections

    Science.gov (United States)

    2017-10-01

    that the dHDPs kill by disrupting membrane function . This is consistent with the amphipathic properties of the dHDPs and is a mechanism to which...recalcitrant biofilm are major obstacles in treating wounds. Antimicrobial peptides ( AMPs ), also known as host defense peptides, are evolutionarily highly...Designed antimicrobial peptides (dAMPs) are synthesized peptides that have been rationally designed based on sequences found in naturally occurring AMPs

  2. De novo nonsense mutations in ASXL1 cause Bohring-Opitz syndrome

    DEFF Research Database (Denmark)

    Hoischen, Alexander; van Bon, Bregje W M; Rodríguez-Santiago, Benjamín

    2011-01-01

    Bohring-Opitz syndrome is characterized by severe intellectual disability, distinctive facial features and multiple congenital malformations. We sequenced the exomes of three individuals with Bohring-Opitz syndrome and in each identified heterozygous de novo nonsense mutations in ASXL1, which...... is required for maintenance of both activation and silencing of Hox genes. In total, 7 out of 13 subjects with a Bohring-Opitz phenotype had de novo ASXL1 mutations, suggesting that the syndrome is genetically heterogeneous....

  3. Novel angiotensin-converting enzyme (ACE) inhibitory peptides derived from boneless chicken leg meat.

    Science.gov (United States)

    Terashima, Masaaki; Baba, Takako; Ikemoto, Narumi; Katayama, Midori; Morimoto, Tomoko; Matsumura, Saki

    2010-06-23

    Four peptides that inhibit angiotensin-converting enzyme (ACE) were separated from the hydorlysate of boneless chicken leg meat digested with artificial gastric juice (pepsin). Two peptides were identified as the peptides encrypted in myosin heavy chain. The peptide P1 (MNVKHWPWMK) corresponds to the amino acid sequence from amino acids 825 to 834 of myosin heavy chain, and the peptide P4 (VTVNPYKWLP) corresponds to the amino acid sequence from amino acids 125 to 135 of myosin heavy chain. They are novel ACE inhibitory peptides derived from chicken, and IC(50) values of P1 and P4 were determined as 228 and 5.5 microM, respectively. Although these values were much larger than 0.022 microM for captopril, a typical synthetic ACE inhibitor, they are comparable to IC(50) values reported for various ACE inhibitory peptides derived from foods. Because the peptide P4 has a relatively low IC(50) value, it is a good starting substance for designing food supplements for hypertensive patients.

  4. Connecting peptide (c-peptide) and the duration of diabetes mellitus ...

    African Journals Online (AJOL)

    Objective: C-peptide is derived from proinsulin and it is secreted in equimolar concentration with insulin. Plasma C-peptide is more stable than insulin and it provides an indirect measure of insulin secretory reserve and beta cell function. To determine relationship between C-peptide and duration of diabetes mellitus, age, ...

  5. Induced resistance to the antimicrobial peptide lactoferricin B in Staphylococcus aureus.

    Science.gov (United States)

    Samuelsen, Orjan; Haukland, Hanne H; Jenssen, Håvard; Krämer, Manuela; Sandvik, Kjersti; Ulvatne, Hilde; Vorland, Lars H

    2005-06-20

    This study was designed to investigate inducible intrinsic resistance against lactoferricin B in Staphylococcus aureus. Serial passage of seven S. aureus strains in medium with increasing concentrations of peptide resulted in an induced resistance at various levels in all strains. The induced resistance was unstable and decreased relatively rapidly during passages in peptide free medium but the minimum inhibitory concentration remained elevated after thirty passages. Cross-resistance to penicillin G and low-level cross-resistance to the antimicrobial peptides indolicidin and Ala(8,13,18)-magainin-II amide [corrected] was observed. No cross-resistance was observed to the human cathelicidin LL-37. In conclusion, this study shows that S. aureus has intrinsic resistance mechanisms against antimicrobial peptides that can be induced upon exposure, and that this may confer low-level cross-resistance to other antimicrobial peptides.

  6. De novo triiodothyronine formation from thyrocytes activated by thyroid-stimulating hormone.

    Science.gov (United States)

    Citterio, Cintia E; Veluswamy, Balaji; Morgan, Sarah J; Galton, Valerie A; Banga, J Paul; Atkins, Stephen; Morishita, Yoshiaki; Neumann, Susanne; Latif, Rauf; Gershengorn, Marvin C; Smith, Terry J; Arvan, Peter

    2017-09-15

    The thyroid gland secretes primarily tetraiodothyronine (T 4 ), and some triiodothyronine (T 3 ). Under normal physiological circumstances, only one-fifth of circulating T 3 is directly released by the thyroid, but in states of hyperactivation of thyroid-stimulating hormone receptors (TSHRs), patients develop a syndrome of relative T 3 toxicosis. Thyroidal T 4 production results from iodination of thyroglobulin (TG) at residues Tyr 5 and Tyr 130 , whereas thyroidal T 3 production may originate in several different ways. In this study, the data demonstrate that within the carboxyl-terminal portion of mouse TG, T 3 is formed de novo independently of deiodination from T 4 We found that upon iodination in vitro , de novo T 3 formation in TG was decreased in mice lacking TSHRs. Conversely, de novo T 3 that can be formed upon iodination of TG secreted from PCCL3 (rat thyrocyte) cells was augmented from cells previously exposed to increased TSH, a TSHR agonist, a cAMP analog, or a TSHR-stimulating antibody. We present data suggesting that TSH-stimulated TG phosphorylation contributes to enhanced de novo T 3 formation. These effects were reversed within a few days after removal of the hyperstimulating conditions. Indeed, direct exposure of PCCL3 cells to human serum from two patients with Graves' disease, but not control sera, led to secretion of TG with an increased intrinsic ability to form T 3 upon in vitro iodination. Furthermore, TG secreted from human thyrocyte cultures hyperstimulated with TSH also showed an increased intrinsic ability to form T 3 Our data support the hypothesis that TG processing in the secretory pathway of TSHR-hyperstimulated thyrocytes alters the structure of the iodination substrate in a way that enhances de novo T 3 formation, contributing to the relative T 3 toxicosis of Graves' disease.

  7. Peptide-Carrier Conjugation

    DEFF Research Database (Denmark)

    Hansen, Paul Robert

    2015-01-01

    To produce antibodies against synthetic peptides it is necessary to couple them to a protein carrier. This chapter provides a nonspecialist overview of peptide-carrier conjugation. Furthermore, a protocol for coupling cysteine-containing peptides to bovine serum albumin is outlined....

  8. Highly selective enrichment of phosphorylated peptides from peptide mixtures using titanium dioxide microcolumns

    DEFF Research Database (Denmark)

    Larsen, Martin Røssel; Thingholm, Tine E; Jensen, Ole N

    2005-01-01

    based on TiO2microcolumns and peptide loading in 2,5-dihydroxybenzoic acid (DHB). The effect of DHB was a very efficient reduction in the binding of nonphosphorylated peptides to TiO2 while retaining its high binding affinity for phosphorylated peptides. Thus, inclusion of DHB dramatically increased...... the selectivity of the enrichment of phosphorylated peptides by TiO2. We demonstrated that this new procedure was more selective for binding phosphorylated peptides than IMAC using MALDI mass spectrometry. In addition, we showed that LC-ESI-MSMS was biased toward monophosphorylated peptides, whereas MALDI MS...... was not. Other substituted aromatic carboxylic acids were also capable of specifically reducing binding of nonphosphorylated peptides, whereas phosphoric acid reduced binding of both phosphorylated and nonphosphorylated peptides. A putative mechanism for this intriguing effect is presented....

  9. Novel short antibacterial and antifungal peptides with low cytotoxicity: Efficacy and action mechanisms

    Energy Technology Data Exchange (ETDEWEB)

    Qi, Xiaobao; Zhou, Chuncai; Li, Peng [School of Chemical and Biomedical Engineering, Nanyang Technological University, 62 Nanyang Drive, 637459 Singapore (Singapore); Xu, Weixin [School of Biological Sciences, Nanyang Technological University, 60 Nanyang Drive, 637551 Singapore (Singapore); Cao, Ye; Ling, Hua; Ning Chen, Wei; Ming Li, Chang; Xu, Rong [School of Chemical and Biomedical Engineering, Nanyang Technological University, 62 Nanyang Drive, 637459 Singapore (Singapore); Lamrani, Mouad [Menicon Co., Ltd. Immeuble Espace Cordeliers, 2, rue President Carnot, 69002 Lyon (France); Mu, Yuguang, E-mail: ygmu@ntu.edu.sg [School of Biological Sciences, Nanyang Technological University, 60 Nanyang Drive, 637551 Singapore (Singapore); Leong, Susanna Su Jan [School of Chemical and Biomedical Engineering, Nanyang Technological University, 62 Nanyang Drive, 637459 Singapore (Singapore); Wook Chang, Matthew, E-mail: matthewchang@ntu.edu.sg [School of Chemical and Biomedical Engineering, Nanyang Technological University, 62 Nanyang Drive, 637459 Singapore (Singapore); Chan-Park, Mary B., E-mail: mbechan@ntu.edu.sg [School of Chemical and Biomedical Engineering, Nanyang Technological University, 62 Nanyang Drive, 637459 Singapore (Singapore)

    2010-07-30

    Research highlights: {yields} Short antimicrobial peptides with nine and eleven residues were developed. {yields} These peptides show strong bactericidal activity against clinically important bacterial and fungal pathogens. {yields} These peptides exhibit high stability in the presence of salts, and low cytotoxicity. {yields} These peptides exert their action by disrupting membrane lipids. -- Abstract: Short antimicrobial peptides with nine and eleven residues were developed against several clinically important bacterial and fungal pathogens (specifically Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus, Candida albicans, and Fusarium solani). Twelve analogues of previously reported peptides BP76 (KKLFKKILKFL) and Pac-525 (KWRRWVRWI) were designed, synthesized, and tested for their antimicrobial activities. Two of our eleven amino acid peptides, P11-5 (GKLFKKILKIL) and P11-6 (KKLIKKILKIL), have very low MICs of 3.1-12.5 {mu}g ml{sup -1} against all five pathogens. The MICs of these two peptides against S. aureus, C. albicans and F. solani are four to ten times lower than the corresponding MICs of the reference peptide BP76. P9-4 (KWRRWIRWL), our newly designed nine-amino acid analogue, also has particularly low MICs of 3.1-6.2 {mu}g ml{sup -1} against four of the tested pathogens; these MICs are two to eight times lower than those reported for Pac-525 (6.2-50 {mu}g ml{sup -1}).These new peptides (P11-5, P11-6 and P9-4) also exhibit improved stability in the presence of salts, and have low cytotoxicity as shown by the hemolysis and MTT assays. From the results of field-emission scanning electron microscopy, membrane depolarization and dye-leakage assays, we propose that these peptides exert their action by disrupting membrane lipids. Molecular dynamics simulation studies confirm that P11-6 peptide maintains relatively stable helical structure and exerts more perturbation action on the order of acyl tail of lipid bilayer.

  10. Peptides with Dual Antimicrobial and Anticancer Activities

    Science.gov (United States)

    Felício, Mário R.; Silva, Osmar N.; Gonçalves, Sônia; Santos, Nuno C.; Franco, Octávio L.

    2017-02-01

    In recent years, the number of people suffering from cancer and multi-resistant infections has increased, such that both diseases are already seen as current and future major causes of death. Moreover, chronic infections are one of the main causes of cancer, due to the instability in the immune system that allows cancer cells to proliferate. Likewise, the physical debility associated with cancer or with anticancer therapy itself often paves the way for opportunistic infections. It is urgent to develop new therapeutic methods, with higher efficiency and lower side effects. Antimicrobial peptides (AMPs) are found in the innate immune system of a wide range of organisms. Identified as the most promising alternative to conventional molecules used nowadays against infections, some of them have been shown to have dual activity, both as antimicrobial and anticancer peptides (ACPs). Highly cationic and amphipathic, they have demonstrated efficacy against both conditions, with the number of nature-driven or synthetically designed peptides increasing year by year. With similar properties, AMPs that can also act as ACPs are viewed as future chemotherapeutic drugs, with the advantage of low propensity to resistance, which started this paradigm in the pharmaceutical market. These peptides have already been described as molecules presenting killing mechanisms at the membrane level, but also acting towards intracellular targets, which increases their success comparatively to specific one-target drugs. This review will approach the desirable characteristics of small peptides that demonstrated dual activity against microbial infections and cancer, as well as the peptides engaged in clinical trials.

  11. Antimicrobial peptide evolution in the Asiatic honey bee Apis cerana.

    Directory of Open Access Journals (Sweden)

    Peng Xu

    Full Text Available The Asiatic honeybee, Apis cerana Fabricius, is an important honeybee species in Asian countries. It is still found in the wild, but is also one of the few bee species that can be domesticated. It has acquired some genetic advantages and significantly different biological characteristics compared with other Apis species. However, it has been less studied, and over the past two decades, has become a threatened species in China. We designed primers for the sequences of the four antimicrobial peptide cDNA gene families (abaecin, defensin, apidaecin, and hymenoptaecin of the Western honeybee, Apis mellifera L. and identified all the antimicrobial peptide cDNA genes in the Asiatic honeybee for the first time. All the sequences were amplified by reverse transcriptase-polymerase chain reaction (RT-PCR. In all, 29 different defensin cDNA genes coding 7 different defensin peptides, 11 different abaecin cDNA genes coding 2 different abaecin peptides, 13 different apidaecin cDNA genes coding 4 apidaecin peptides and 34 different hymenoptaecin cDNA genes coding 13 different hymenoptaecin peptides were cloned and identified from the Asiatic honeybee adult workers. Detailed comparison of these four antimicrobial peptide gene families with those of the Western honeybee revealed that there are many similarities in the quantity and amino acid components of peptides in the abaecin, defensin and apidaecin families, while many more hymenoptaecin peptides are found in the Asiatic honeybee than those in the Western honeybee (13 versus 1. The results indicated that the Asiatic honeybee adult generated more variable antimicrobial peptides, especially hymenoptaecin peptides than the Western honeybee when stimulated by pathogens or injury. This suggests that, compared to the Western honeybee that has a longer history of domestication, selection on the Asiatic honeybee has favored the generation of more variable antimicrobial peptides as protection against pathogens.

  12. Antimicrobial peptide evolution in the Asiatic honey bee Apis cerana.

    Science.gov (United States)

    Xu, Peng; Shi, Min; Chen, Xue-Xin

    2009-01-01

    The Asiatic honeybee, Apis cerana Fabricius, is an important honeybee species in Asian countries. It is still found in the wild, but is also one of the few bee species that can be domesticated. It has acquired some genetic advantages and significantly different biological characteristics compared with other Apis species. However, it has been less studied, and over the past two decades, has become a threatened species in China. We designed primers for the sequences of the four antimicrobial peptide cDNA gene families (abaecin, defensin, apidaecin, and hymenoptaecin) of the Western honeybee, Apis mellifera L. and identified all the antimicrobial peptide cDNA genes in the Asiatic honeybee for the first time. All the sequences were amplified by reverse transcriptase-polymerase chain reaction (RT-PCR). In all, 29 different defensin cDNA genes coding 7 different defensin peptides, 11 different abaecin cDNA genes coding 2 different abaecin peptides, 13 different apidaecin cDNA genes coding 4 apidaecin peptides and 34 different hymenoptaecin cDNA genes coding 13 different hymenoptaecin peptides were cloned and identified from the Asiatic honeybee adult workers. Detailed comparison of these four antimicrobial peptide gene families with those of the Western honeybee revealed that there are many similarities in the quantity and amino acid components of peptides in the abaecin, defensin and apidaecin families, while many more hymenoptaecin peptides are found in the Asiatic honeybee than those in the Western honeybee (13 versus 1). The results indicated that the Asiatic honeybee adult generated more variable antimicrobial peptides, especially hymenoptaecin peptides than the Western honeybee when stimulated by pathogens or injury. This suggests that, compared to the Western honeybee that has a longer history of domestication, selection on the Asiatic honeybee has favored the generation of more variable antimicrobial peptides as protection against pathogens.

  13. A microbially derived tyrosine-sulfated peptide mimics a plant peptide hormone.

    Science.gov (United States)

    Pruitt, Rory N; Joe, Anna; Zhang, Weiguo; Feng, Wei; Stewart, Valley; Schwessinger, Benjamin; Dinneny, José R; Ronald, Pamela C

    2017-07-01

    The biotrophic pathogen Xanthomonas oryzae pv. oryzae (Xoo) produces a sulfated peptide named RaxX, which shares similarity to peptides in the PSY (plant peptide containing sulfated tyrosine) family. We hypothesize that RaxX mimics the growth-stimulating activity of PSY peptides. Root length was measured in Arabidopsis and rice treated with synthetic RaxX peptides. We also used comparative genomic analyses and reactive oxygen species burst assays to evaluate the activity of RaxX and PSY peptides. Here we found that a synthetic sulfated RaxX derivative comprising 13 residues (RaxX13-sY), highly conserved between RaxX and PSY, induces root growth in Arabidopsis and rice in a manner similar to that triggered by PSY. We identified residues that are required for activation of immunity mediated by the rice XA21 receptor but that are not essential for root growth induced by PSY. Finally, we showed that a Xanthomonas strain lacking raxX is impaired in virulence. These findings suggest that RaxX serves as a molecular mimic of PSY peptides to facilitate Xoo infection and that XA21 has evolved the ability to recognize and respond specifically to the microbial form of the peptide. © 2017 UT-Battelle LLC. New Phytologist © 2017 New Phytologist Trust.

  14. Memetic algorithms for de novo motif-finding in biomedical sequences.

    Science.gov (United States)

    Bi, Chengpeng

    2012-09-01

    The objectives of this study are to design and implement a new memetic algorithm for de novo motif discovery, which is then applied to detect important signals hidden in various biomedical molecular sequences. In this paper, memetic algorithms are developed and tested in de novo motif-finding problems. Several strategies in the algorithm design are employed that are to not only efficiently explore the multiple sequence local alignment space, but also effectively uncover the molecular signals. As a result, there are a number of key features in the implementation of the memetic motif-finding algorithm (MaMotif), including a chromosome replacement operator, a chromosome alteration-aware local search operator, a truncated local search strategy, and a stochastic operation of local search imposed on individual learning. To test the new algorithm, we compare MaMotif with a few of other similar algorithms using simulated and experimental data including genomic DNA, primary microRNA sequences (let-7 family), and transmembrane protein sequences. The new memetic motif-finding algorithm is successfully implemented in C++, and exhaustively tested with various simulated and real biological sequences. In the simulation, it shows that MaMotif is the most time-efficient algorithm compared with others, that is, it runs 2 times faster than the expectation maximization (EM) method and 16 times faster than the genetic algorithm-based EM hybrid. In both simulated and experimental testing, results show that the new algorithm is compared favorably or superior to other algorithms. Notably, MaMotif is able to successfully discover the transcription factors' binding sites in the chromatin immunoprecipitation followed by massively parallel sequencing (ChIP-Seq) data, correctly uncover the RNA splicing signals in gene expression, and precisely find the highly conserved helix motif in the transmembrane protein sequences, as well as rightly detect the palindromic segments in the primary micro

  15. Tumor penetrating peptides

    Directory of Open Access Journals (Sweden)

    Tambet eTeesalu

    2013-08-01

    Full Text Available Tumor-homing peptides can be used to deliver drugs into tumors. Phage library screening in live mice has recently identified homing peptides that specifically recognize the endothelium of tumor vessels, extravasate, and penetrate deep into the extravascular tumor tissue. The prototypic peptide of this class, iRGD (CRGDKGPDC, contains the integrin-binding RGD motif. RGD mediates tumor homing through binding to αv integrins, which are selectively expressed on various cells in tumors, including tumor endothelial cells. The tumor-penetrating properties of iRGD are mediated by a second sequence motif, R/KXXR/K. This C-end Rule (or CendR motif is active only when the second basic residue is exposed at the C-terminus of the peptide. Proteolytic processing of iRGD in tumors activates the cryptic CendR motif, which then binds to neuropilin-1 activating an endocytic bulk transport pathway through tumor tissue. Phage screening has also yielded tumor-penetrating peptides that function like iRGD in activating the CendR pathway, but bind to a different primary receptor. Moreover, novel tumor-homing peptides can be constructed from tumor-homing motifs, CendR elements and protease cleavage sites. Pathologies other than tumors can be targeted with tissue-penetrating peptides, and the primary receptor can also be a vascular zip code of a normal tissue. The CendR technology provides a solution to a major problem in tumor therapy, poor penetration of drugs into tumors. The tumor-penetrating peptides are capable of taking a payload deep into tumor tissue in mice, and they also penetrate into human tumors ex vivo. Targeting with these peptides specifically increases the accumulation in tumors of a variety of drugs and contrast agents, such as doxorubicin, antibodies and nanoparticle-based compounds. Remarkably the drug to be targeted does not have to be coupled to the peptide; the bulk transport system activated by the peptide sweeps along any compound that is

  16. In-silico screening and validation of high-affinity tetra-peptide inhibitor of Leishmania donovani O-acetyl serine sulfhydrylase (OASS).

    Science.gov (United States)

    Kant, Vishnu; Vijayakumar, Saravanan; Sahoo, Ganesh Chandra; Chaudhery, Shailendra S; Das, Pradeep

    2018-02-07

    OASS is a specific enzyme that helps Leishmania parasite to survive the oxidative stress condition in human macrophages. SAT C-terminal peptides in several organisms, including Leishmania, were reported to inhibit or reduce the activity of OASS. Small peptide and small molecules mimicking the SAT C-terminal residues are designed and tested for the inhibition of OASS in different organisms. Hence, in this study, all the possible tetra-peptide combinations were designed and screened based on the docking ability with Leishmania donovani OASS (Ld-OASS). The top ranked peptides were further validated for the stability using 50 ns molecular dynamic simulation. In order to identify the better binding capability of the peptides, the top peptides complexed with Ld-OASS were also subjected to molecular dynamic simulation. The docking and simulation results favored the peptide EWSI to possess greater advantage than previously reported peptide (DWSI) in binding with Ld-OASS active site. Also, screening of non-peptide inhibitor of Asinex Biodesign library based on the shape similarity of EWSI and DWSI was performed. The top similar molecules of each peptides were docked on to Ld-OASS active site and subsequently simulated for 20 ns. The results suggested that the ligand that shares high shape similarity with EWSI possess better binding capability than the ligand that shares high shape similarity with DWSI. This study revealed that the tetra-peptide EWSI had marginal advantage over DWSI in binding with Ld-OASS, thereby providing basis for defining a pharmacophoric scaffold for the design of peptidomimetic inhibitors as well as non-peptide inhibitors of Ld-OASS.

  17. The impact of employee satisfaction on productivity in Tiskarna Novo mesto, Ltd.

    Directory of Open Access Journals (Sweden)

    Simona Cimperman

    2016-06-01

    Full Text Available Research Question: Does employee satisfaction, impact on productivity? How are these two variables associated? What is the job satisfaction in Tiskarna Novo mesto, Ltd. What needs to be done to make employees more satisfied at work and, consequently, more productive? Purpose: The purpose of the study is to determine what are the factors that influence employee satisfaction Tiskarna Novo mesto, Ltd. and check the connection between work satisfaction and employee productivity. The aim of the research is to examine what is the level of job satisfaction of employees in Tiskarna Novo mesto, Ltd. And find our reasons and factors that prevent employees were satisfied in the workplace. Method: In this study we used a descriptive method and the method of combining the study of domestic and foreign literature. Pending the results we have come to interview employees in the Tiskarna Novo mesto, Ltd. Results: We conducted a survey among employees in Tiskarna Novo mesto, Ltd and we came to the conclusion that the employees are medium satisfied – the average grade point job satisfaction of employees was 3.1 (evaluated on a 5-point Likert scale. The worst assessed was factor in job satisfaction opportunity for advancement and educational opportunities. We have found out that factors like receiving praise and awards as well as good interpersonal relations are those that affect good on job satisfaction, on the other hand conflict is the one that reduces job satisfaction. The existence of links between work satisfaction and productivity were not found (r = -0.061. Organization: The organization and managers, it is important to know which are the factors by which employees are satisfied or dissatisfied. Results of the research will give managers a clear picture of the factors of satisfaction / dissatisfaction and opinion on productivity. Society: The employees it means a lot to have your job satisfaction and consequently they are more productive. Originality: The

  18. Interplay between De Novo Biosynthesis and Sequestration of Cyanogenic Glucosides in Arthropods

    DEFF Research Database (Denmark)

    Fürstenberg-Hägg, Joel

    (Zygaenidae, Lepidoptera) both sequester (take up and accumulate) the CNglcs linamarin and lotaustralin from their food plants (Fabacea) and biosynthesize them de novo from valine and isoleucine. The presented research demonstrates that de novo biosynthesis of CNglcs in Z. filipendulae is dependent...

  19. Biopharmaceuticals: From peptide to drug

    Science.gov (United States)

    Hannappel, Margarete

    2017-08-01

    Biologics are therapeutic proteins or peptides that are produced by means of biological processes within living organisms and cells. They are highly specific molecules and play a crucial role as therapeutics for the treatment of severe and chronic diseases (e.g. cancer, rheumatoid arthritis, diabetes, autoimmune disorders). The development of new biologics and biologics-based drugs gains more and more importance in the fight against various diseases. A short overview on biotherapeutical drug development is given. Cone snails are a large group of poisonous, predatory sea snails with more than 700 species. They use a very powerful venom which rapidly inactivates and paralyzes their prey. Most bioactive venom components are small peptides (conotoxins, conopeptides) which are precisely directed towards a specific target (e.g. ion channel, receptors). Due to their small size, their precision and speed of action, naturally occurring cone snail venom peptides represent an attractive source for the identification and design of novel biological drug entities. The Jagna cone snail project is an encouraging initiative to map the ecological variety of cone snails around the island of Bohol (Philippines) and to conserve the biological information for potential future application.

  20. A multiple multicomponent approach to chimeric peptide-peptoid podands.

    Science.gov (United States)

    Rivera, Daniel G; León, Fredy; Concepción, Odette; Morales, Fidel E; Wessjohann, Ludger A

    2013-05-10

    The success of multi-armed, peptide-based receptors in supramolecular chemistry traditionally is not only based on the sequence but equally on an appropriate positioning of various peptidic chains to create a multivalent array of binding elements. As a faster, more versatile and alternative access toward (pseudo)peptidic receptors, a new approach based on multiple Ugi four-component reactions (Ugi-4CR) is proposed as a means of simultaneously incorporating several binding and catalytic elements into organizing scaffolds. By employing α-amino acids either as the amino or acid components of the Ugi-4CRs, this multiple multicomponent process allows for the one-pot assembly of podands bearing chimeric peptide-peptoid chains as appended arms. Tripodal, bowl-shaped, and concave polyfunctional skeletons are employed as topologically varied platforms for positioning the multiple peptidic chains formed by Ugi-4CRs. In a similar approach, steroidal building blocks with several axially-oriented isocyano groups are synthesized and utilized to align the chimeric chains with conformational constrains, thus providing an alternative to the classical peptido-steroidal receptors. The branched and hybrid peptide-peptoid appendages allow new possibilities for both rational design and combinatorial production of synthetic receptors. The concept is also expandable to other multicomponent reactions. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  1. Electric fields control the orientation of peptides irreversibly immobilized on radical-functionalized surfaces.

    Science.gov (United States)

    Martin, Lewis J; Akhavan, Behnam; Bilek, Marcela M M

    2018-01-24

    Surface functionalization of an implantable device with bioactive molecules can overcome adverse biological responses by promoting specific local tissue integration. Bioactive peptides have advantages over larger protein molecules due to their robustness and sterilizability. Their relatively small size presents opportunities to control the peptide orientation on approach to a surface to achieve favourable presentation of bioactive motifs. Here we demonstrate control of the orientation of surface-bound peptides by tuning electric fields at the surface during immobilization. Guided by computational simulations, a peptide with a linear conformation in solution is designed. Electric fields are used to control the peptide approach towards a radical-functionalized surface. Spontaneous, irreversible immobilization is achieved when the peptide makes contact with the surface. Our findings show that control of both peptide orientation and surface concentration is achieved simply by varying the solution pH or by applying an electric field as delivered by a small battery.

  2. Hypotensive Effects and Angiotensin-Converting Enzyme Inhibitory Peptides of Reishi (Ganoderma lingzhi Auto-Digested Extract

    Directory of Open Access Journals (Sweden)

    Hai-Bang Tran

    2014-08-01

    Full Text Available Reishi (Ganoderma lingzhi has been used as a traditional medicine for millennia. However, relatively little is known about this mushroom’s proteins and their bioactivities. In this study, we used reishi’s own proteases to hydrolyze its protein and obtained auto-digested reishi (ADR extract. The extract was subjected to in vitro assays and administered to spontaneous hypertensive rats (SHRs to determine its potential for use as a hypotensive medication. Bioassay-guided fractionation and de novo sequencing were used for identifying the active compounds. After 4 h administration of ADR, the systolic pressure of SHRs significantly decreased to 34.3 mmHg (19.5% change and the effect was maintained up to 8 h of administration, with the decrease reaching as low as 26.8 mmHg (15% reduction–compare to base line a decrease of 26.8 mmHg is less than a decrease of 34.3 mmHg so it should give a smaller % reduction. Eleven peptides were identified and four of them showed potent inhibition against ACE with IC50 values ranging from 73.1 μM to 162.7 μM. The results showed that ADR could be a good source of hypotensive peptides that could be used for antihypertensive medication or incorporation into functional foods.

  3. Peptide Nucleic Acids

    DEFF Research Database (Denmark)

    2003-01-01

    A novel class of compounds, known as peptide nucleic acids, bind complementary ssDNA and RNA strands more strongly than a corresponding DNA. The peptide nucleic acids generally comprise ligands such as naturally occurring DNA bases attached to a peptide backbone through a suitable linker....

  4. Peptide Nucleic Acids

    DEFF Research Database (Denmark)

    1998-01-01

    A novel class of compounds, known as peptide nucleic acids, bind complementary ssDNA and RNA strands more strongly than a corresponding DNA. The peptide nucleic acids generally comprise ligands such as naturally occurring DNA bases attached to a peptide backbone through a suitable linker....

  5. PeptideAtlas

    Data.gov (United States)

    U.S. Department of Health & Human Services — PeptideAtlas is a multi-organism, publicly accessible compendium of peptides identified in a large set of tandem mass spectrometry proteomics experiments. Mass...

  6. Synthetic peptide vaccines: palmitoylation of peptide antigens by a thioester bond increases immunogenicity

    DEFF Research Database (Denmark)

    Beekman, N.J.C.M.; Schaaper, W.M.M.; Tesser, G.I.

    1997-01-01

    Synthetic peptides have frequently been used to immunize animals. However, peptides less than about 20 to 30 amino acids long are poor immunogens. In general, to increase its immunogenicity, the presentation of the peptide should be improved, and molecular weight needs to be increased. Many...... or an amide bond. It was found that these S-palmitoylated peptides were much more immunogenic than N-palmitoylated peptides and at least similar to KLH-conjugated peptides with respect to appearance and magnitude of induced antibodies (canine parvovirus) or immunocastration effect (gonadotropin...

  7. Novo Jornalismo: fronteiras litero-factuais em A sangue Frio e em Radical Chique

    Directory of Open Access Journals (Sweden)

    Francisco Aquinei Timóteo Queirós

    2012-12-01

    Full Text Available A pesquisa busca analisar de que forma fato e ficção se entrecruzam no “movimento” do Novo Jornalismo, a partir das obras A sangue Frio e Radical Chique e o Novo Jornalismo, de Truman Capote e Tom Wolfe, respectivamente. Pretende-se, a partir da investigação do corpus em estudo, revelar os aspectos que aproximam o fato jornalístico, a notícia e a reportagem às técnicas literárias do romance, do conto e da crônica. O estudo investiga o Novo Jornalismo sob o viés de textos centrais das áreas de teoria literária e estudos jornalísticos utilizando autores como Mikhail Bakhtin, Hayden White, Paul Ricoeur, Muniz Sodré; além de referenciar outros escritores que, como Tom Wolfe e Truman Capote, fizeram parte de um grande movimento renovador do jornalismo literário nos anos 1950, 1960 e 1970 chamado, genericamente, de Novo Jornalismo.

  8. Demanda dos principais metais e novos materiais : analise de tendencias

    OpenAIRE

    Wilson Trigueiro de Sousa

    1990-01-01

    Resumo: Neste trabalho são analisadas algumas tendências na área de novos materiais na tentativa de obter um melhor entendimento das repercussões das atuais inovações tecnológicas para o setor mineral. Inicialmente são revisados os principais estudos sobre as mudanças ocorridas por volta de 1972/74 no comportamento da demanda dos metais mais importantes. Entre as possíveis causas, está o progresso técnico, que tornou possível o surgimento de novos materiais e o aperfeiçoamento de outros em us...

  9. Piezoelectric peptide-based nanogenerator enhanced by single-electrode triboelectric nanogenerator

    Directory of Open Access Journals (Sweden)

    Vu Nguyen

    2017-07-01

    Full Text Available Peptide has recently been demonstrated as a sustainable and smart material for piezoelectric energy conversion. Although the power output was improved compared to other biomaterials, the use of a piezoelectric device alone can only capture the energy from the minute deformation in materials. In comparison, the triboelectric effect can convert mechanical energy from large motion. Consequently, utilizing both piezoelectric and triboelectric effects is of significant research interest due to their complementary energy conversion mechanisms. Here we demonstrated a hybrid nanogenerator that combined a peptide-based piezoelectric nanogenerator with a single-electrode triboelectric nanogenerator. Our device structure enabled the voltage and current outputs of each individual type of nanogenerator to be superposed in the hybrid nanogenerator, producing overall constructive outputs. The design of our device also enabled a simplified configuration of hybrid nanogenerator. This study is important not only for the enhancement of peptide-based piezoelectric device but also for the future design of hybrid piezoelectric and triboelectric nanogenerators.

  10. Sensitivity-Informed De Novo Programming for Many-Objective Water Portfolio Planning Under Uncertainty

    Science.gov (United States)

    Kasprzyk, J. R.; Reed, P. M.; Kirsch, B. R.; Characklis, G. W.

    2009-12-01

    Risk-based water supply management presents severe cognitive, computational, and social challenges to planning in a changing world. Decision aiding frameworks must confront the cognitive biases implicit to risk, the severe uncertainties associated with long term planning horizons, and the consequent ambiguities that shape how we define and solve water resources planning and management problems. This paper proposes and demonstrates a new interactive framework for sensitivity informed de novo programming. The theoretical focus of our many-objective de novo programming is to promote learning and evolving problem formulations to enhance risk-based decision making. We have demonstrated our proposed de novo programming framework using a case study for a single city’s water supply in the Lower Rio Grande Valley (LRGV) in Texas. Key decisions in this case study include the purchase of permanent rights to reservoir inflows and anticipatory thresholds for acquiring transfers of water through optioning and spot leases. A 10-year Monte Carlo simulation driven by historical data is used to provide performance metrics for the supply portfolios. The three major components of our methodology include Sobol globoal sensitivity analysis, many-objective evolutionary optimization and interactive tradeoff visualization. The interplay between these components allows us to evaluate alternative design metrics, their decision variable controls and the consequent system vulnerabilities. Our LRGV case study measures water supply portfolios’ efficiency, reliability, and utilization of transfers in the water supply market. The sensitivity analysis is used interactively over interannual, annual, and monthly time scales to indicate how the problem controls change as a function of the timescale of interest. These results have been used then to improve our exploration and understanding of LRGV costs, vulnerabilities, and the water portfolios’ critical reliability constraints. These results

  11. Sequencing and de novo assembly of 150 genomes from Denmark as a population reference

    DEFF Research Database (Denmark)

    Maretty, Lasse; Jensen, Jacob Malte; Petersen, Bent

    2017-01-01

    or by performing local assembly. However, these approaches are biased against discovery of structural variants and variation in the more complex parts of the genome. Hence, large-scale de novo assembly is needed. Here we show that it is possible to construct excellent de novo assemblies from high......-coverage sequencing with mate-pair libraries extending up to 20 kilobases. We report de novo assemblies of 150 individuals (50 trios) from the GenomeDenmark project. The quality of these assemblies is similar to those obtained using the more expensive long-read technology. We use the assemblies to identify a rich set...

  12. Self-assembled peptide-based nanostructures: Smart nanomaterials toward targeted drug delivery.

    Science.gov (United States)

    Habibi, Neda; Kamaly, Nazila; Memic, Adnan; Shafiee, Hadi

    2016-02-01

    Self-assembly of peptides can yield an array of well-defined nanostructures that are highly attractive nanomaterials for many biomedical applications such as drug delivery. Some of the advantages of self-assembled peptide nanostructures over other delivery platforms include their chemical diversity, biocompatibility, high loading capacity for both hydrophobic and hydrophilic drugs, and their ability to target molecular recognition sites. Furthermore, these self-assembled nanostructures could be designed with novel peptide motifs, making them stimuli-responsive and achieving triggered drug delivery at disease sites. The goal of this work is to present a comprehensive review of the most recent studies on self-assembled peptides with a focus on their "smart" activity for formation of targeted and responsive drug-delivery carriers.

  13. Hydrophobic and electrostatic interactions between cell penetrating peptides and plasmid DNA are important for stable non-covalent complexation and intracellular delivery.

    Science.gov (United States)

    Upadhya, Archana; Sangave, Preeti C

    2016-10-01

    Cell penetrating peptides are useful tools for intracellular delivery of nucleic acids. Delivery of plasmid DNA, a large nucleic acid, poses a challenge for peptide mediated transport. The paper investigates and compares efficacy of five novel peptide designs for complexation of plasmid DNA and subsequent delivery into cells. The peptides were designed to contain reported DNA condensing agents and basic cell penetrating sequences, octa-arginine (R 8 ) and CHK 6 HC coupled to cell penetration accelerating peptides such as Bax inhibitory mutant peptide (KLPVM) and a peptide derived from the Kaposi fibroblast growth factor (kFGF) membrane translocating sequence. A tryptophan rich peptide, an analogue of Pep-3, flanked with CH 3 on either ends was also a part of the study. The peptides were analysed for plasmid DNA complexation, protection of peptide-plasmid DNA complexes against DNase I, serum components and competitive ligands by simple agarose gel electrophoresis techniques. Hemolysis of rat red blood corpuscles (RBCs) in the presence of the peptides was used as a measure of peptide cytotoxicity. Plasmid DNA delivery through the designed peptides was evaluated in two cell lines, human cervical cancer cell line (HeLa) and (NIH/3 T3) mouse embryonic fibroblasts via expression of the secreted alkaline phosphatase (SEAP) reporter gene. The importance of hydrophobic sequences in addition to cationic sequences in peptides for non-covalent plasmid DNA complexation and delivery has been illustrated. An alternative to the employment of fatty acid moieties for enhanced gene transfer has been proposed. Comparison of peptides for plasmid DNA complexation and delivery of peptide-plasmid DNA complexes to cells estimated by expression of a reporter gene, SEAP. Copyright © 2016 European Peptide Society and John Wiley & Sons, Ltd. Copyright © 2016 European Peptide Society and John Wiley & Sons, Ltd.

  14. “Specificity Determinants” Improve Therapeutic Indices of Two Antimicrobial Peptides Piscidin 1 and Dermaseptin S4 Against the Gram-negative Pathogens Acinetobacter baumannii and Pseudomonas aeruginosa

    Directory of Open Access Journals (Sweden)

    Ziqing Jiang

    2014-03-01

    Full Text Available A new class of antimicrobial agents with lower rates of resistance and different targets is urgently needed because of the rapidly increasing resistance to classical antibiotics. Amphipathic cationic α-helical antimicrobial peptides (AMPs represent such a class of compounds. In our previous studies, using a 26-residue de novo designed antimicrobial peptide, we proposed the concept of “specificity determinant(s”: positively charged residue(s in the center of the non-polar face of AMPs that could decrease hemolytic activity/toxicity but increase or maintain the same level of antimicrobial activity to increase dramatically the therapeutic index. In the current study, we used d-enantiomers of two AMPs, Piscidin 1 isolated from fish and dermaseptin S4 isolated from frog. We substituted different positions in the center of the hydrophobic face with one or two lysine residue(s (one or two “specificity determinant(s”. This simple modification not only maintained or improved antimicrobial activity against Gram-negative pathogens Acinetobacter baumannii (11 strains and Pseudomonas aeruginosa (6 strains, but also dramatically decreased hemolytic activity of human red blood cells, as predicted. Therapeutic indices improved by 55-fold and 730-fold for piscidin 1 (I9K and dermaseptin S4 (L7K, A14K, respectively, against A. baumannii. Similarly, the therapeutic indices improved 32-fold and 980-fold for piscidin 1 (I9K and dermaseptin S4 (L7K, A14K, respectively, against P. aeruginosa.

  15. Diversity-oriented peptide stapling

    DEFF Research Database (Denmark)

    Tran, Thu Phuong; Larsen, Christian Ørnbøl; Røndbjerg, Tobias

    2017-01-01

    as a powerful method for peptide stapling. However, to date CuAAC stapling has not provided a simple method for obtaining peptides that are easily diversified further. In the present study, we report a new diversity-oriented peptide stapling (DOPS) methodology based on CuAAC chemistry. Stapling of peptides...

  16. Automated solid-phase peptide synthesis to obtain therapeutic peptides

    Directory of Open Access Journals (Sweden)

    Veronika Mäde

    2014-05-01

    Full Text Available The great versatility and the inherent high affinities of peptides for their respective targets have led to tremendous progress for therapeutic applications in the last years. In order to increase the drugability of these frequently unstable and rapidly cleared molecules, chemical modifications are of great interest. Automated solid-phase peptide synthesis (SPPS offers a suitable technology to produce chemically engineered peptides. This review concentrates on the application of SPPS by Fmoc/t-Bu protecting-group strategy, which is most commonly used. Critical issues and suggestions for the synthesis are covered. The development of automated methods from conventional to essentially improved microwave-assisted instruments is discussed. In order to improve pharmacokinetic properties of peptides, lipidation and PEGylation are described as covalent conjugation methods, which can be applied by a combination of automated and manual synthesis approaches. The synthesis and application of SPPS is described for neuropeptide Y receptor analogs as an example for bioactive hormones. The applied strategies represent innovative and potent methods for the development of novel peptide drug candidates that can be manufactured with optimized automated synthesis technologies.

  17. Novel chimeric peptide with enhanced cell specificity and anti-inflammatory activity.

    Science.gov (United States)

    Kim, Young-Min; Kim, Nam-Hong; Lee, Jong-Wan; Jang, Jin-Sun; Park, Yung-Hoon; Park, Seong-Cheol; Jang, Mi-Kyeong

    2015-07-31

    An antimicrobial peptide (AMP), Hn-Mc, was designed by combining the N-terminus of HPA3NT3 and the C-terminus of melittin. This chimeric AMP exhibited potent antibacterial activity with low minimal inhibitory concentrations (MICs), ranging from 1 to 2 μM against four drug-susceptible bacteria and ten drug-resistant bacteria. Moreover, the hemolysis and cytotoxicity was reduced significantly compared to those of the parent peptides, highlighting its high cell selectivity. The morphological changes in the giant unilamellar vesicles and bacterial cell surfaces caused by the Hn-Mc peptide suggested that it killed the microbial cells by damaging the membrane envelope. An in vivo study also demonstrated the antibacterial activity of the Hn-Mc peptide in a mouse model infected with drug-resistant bacteria. In addition, the chimeric peptide inhibited the expression of lipopolysaccharide (LPS)-induced cytokines in RAW 264.7 cells by preventing the interaction between LPS and Toll-like receptors. These results suggest that this chimeric peptide is an antimicrobial and anti-inflammatory candidate as a pharmaceutic agent. Copyright © 2015 Elsevier Inc. All rights reserved.

  18. Production of peptide antisera specific for mouse and rat proinsulin C-peptide 2

    DEFF Research Database (Denmark)

    Blume, N; Madsen, O D; Kofod, Hans

    1990-01-01

    for antibody binding to the immunizing antigen. Antisera to C-peptide 2, stained islet beta-cells on mouse and rat, but not monkey pancreas sections in immunocytochemical analysis. Preabsorption to the synthetic C-peptide 2, but not the synthetic mouse and rat C-peptide 1 abolished staining. In conclusion we......Mice and rats have two functional non-allelic insulin genes. By using a synthetic peptide representing a common sequence in mouse and rat C-peptide 2 as antigen, we have produced rabbit antisera specific for an epitope which is not present in mouse or rat C-peptide 1. Long-term immunization did...... not seem to increase the end point titre as tested in direct ELISA. The specificity of the antiserum was determined by competitive ELISA and histochemistry on pancreas sections. Only the synthetic C-peptide 2, but not the homologous synthetic C-peptide 1 from mouse and rat competed efficiently in ELISA...

  19. Coiled-coil forming peptides for the induction of silver nanoparticles

    International Nuclear Information System (INIS)

    Božič Abram, Sabina; Aupič, Jana; Dražić, Goran; Gradišar, Helena; Jerala, Roman

    2016-01-01

    Biopolymers with defined sequence patterns offer an attractive alternative for the formation of silver nanoparticle (AgNP). A set of coiled-coil dimer forming peptides was tested for their AgNP formation ability. Seventeen of those peptides mediated the formation of AgNPs in aqueous solution at neutral pH, while the formation of a coiled-coil dimer inhibited the nanoparticle generation. A QSAR regression model on the relationship between sequence and function suggests that in this peptide type the patterns KXQQ and KXEE are favorable, whereas Ala residues appear to have an inhibitory effect. UV–VIS spectra of the obtained nanoparticles gave a peak at around 420 nm, typical for AgNPs in the size range around 40 nm, which was confirmed by dynamic light scattering and transmission electron microscopy. Peptide-induced AgNPs exhibited good antibacterial activity, even after a 15 min contact time, while they had low toxicity to human cells at the same concentrations. These results show that our designed peptides generate AgNPs with antibacterial activity at mild conditions and might be used for antibacterial coatings. - Highlights: • 17 of the 30 tested coiled-coil forming peptides induce AgNP formation. • Coiled-coil dimer formation suppresses AgNP generation of individual peptides. • Size of the peptide-induced silver nanoparticles is around 40 nm. • QSAR analysis points to the importance of KXQQ and KXEE motifs for AgNP generation. • Peptide-induced silver nanoparticles exhibit antibacterial activity.

  20. Exploring the role of peptides in polymer-based gene delivery.

    Science.gov (United States)

    Sun, Yanping; Yang, Zhen; Wang, Chunxi; Yang, Tianzhi; Cai, Cuifang; Zhao, Xiaoyun; Yang, Li; Ding, Pingtian

    2017-09-15

    Polymers are widely studied as non-viral gene vectors because of their strong DNA binding ability, capacity to carry large payload, flexibility of chemical modifications, low immunogenicity, and facile processes for manufacturing. However, high cytotoxicity and low transfection efficiency substantially restrict their application in clinical trials. Incorporating functional peptides is a promising approach to address these issues. Peptides demonstrate various functions in polymer-based gene delivery systems, such as targeting to specific cells, breaching membrane barriers, facilitating DNA condensation and release, and lowering cytotoxicity. In this review, we systematically summarize the role of peptides in polymer-based gene delivery, and elaborate how to rationally design polymer-peptide based gene delivery vectors. Polymers are widely studied as non-viral gene vectors, but suffer from high cytotoxicity and low transfection efficiency. Incorporating short, bioactive peptides into polymer-based gene delivery systems can address this issue. Peptides demonstrate various functions in polymer-based gene delivery systems, such as targeting to specific cells, breaching membrane barriers, facilitating DNA condensation and release, and lowering cytotoxicity. In this review, we highlight the peptides' roles in polymer-based gene delivery, and elaborate how to utilize various functional peptides to enhance the transfection efficiency of polymers. The optimized peptide-polymer vectors should be able to alter their structures and functions according to biological microenvironments and utilize inherent intracellular pathways of cells, and consequently overcome the barriers during gene delivery to enhance transfection efficiency. Copyright © 2017 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

  1. Coiled-coil forming peptides for the induction of silver nanoparticles

    Energy Technology Data Exchange (ETDEWEB)

    Božič Abram, Sabina [Department of Synthetic Biology and Immunology, National Institute of Chemistry, Hajdrihova 19, 1000 Ljubljana (Slovenia); Graduate School of Biomedicine, University of Ljubljana, Ljubljana 1000 (Slovenia); Aupič, Jana [Department of Synthetic Biology and Immunology, National Institute of Chemistry, Hajdrihova 19, 1000 Ljubljana (Slovenia); Doctoral Programme in Chemical Sciences, Faculty of Chemistry and Chemical Technology, University of Ljubljana, Ljubljana 1000 (Slovenia); Dražić, Goran [Laboratory for Materials Chemistry, National Institute of Chemistry, Hajdrihova 19, 1000 Ljubljana (Slovenia); Gradišar, Helena [Department of Synthetic Biology and Immunology, National Institute of Chemistry, Hajdrihova 19, 1000 Ljubljana (Slovenia); EN-FIST, Centre of Excellence, Trg Osvobodilne fronte 13, Ljubljana 1000 (Slovenia); Jerala, Roman, E-mail: roman.jerala@ki.si [Department of Synthetic Biology and Immunology, National Institute of Chemistry, Hajdrihova 19, 1000 Ljubljana (Slovenia); EN-FIST, Centre of Excellence, Trg Osvobodilne fronte 13, Ljubljana 1000 (Slovenia)

    2016-04-08

    Biopolymers with defined sequence patterns offer an attractive alternative for the formation of silver nanoparticle (AgNP). A set of coiled-coil dimer forming peptides was tested for their AgNP formation ability. Seventeen of those peptides mediated the formation of AgNPs in aqueous solution at neutral pH, while the formation of a coiled-coil dimer inhibited the nanoparticle generation. A QSAR regression model on the relationship between sequence and function suggests that in this peptide type the patterns KXQQ and KXEE are favorable, whereas Ala residues appear to have an inhibitory effect. UV–VIS spectra of the obtained nanoparticles gave a peak at around 420 nm, typical for AgNPs in the size range around 40 nm, which was confirmed by dynamic light scattering and transmission electron microscopy. Peptide-induced AgNPs exhibited good antibacterial activity, even after a 15 min contact time, while they had low toxicity to human cells at the same concentrations. These results show that our designed peptides generate AgNPs with antibacterial activity at mild conditions and might be used for antibacterial coatings. - Highlights: • 17 of the 30 tested coiled-coil forming peptides induce AgNP formation. • Coiled-coil dimer formation suppresses AgNP generation of individual peptides. • Size of the peptide-induced silver nanoparticles is around 40 nm. • QSAR analysis points to the importance of KXQQ and KXEE motifs for AgNP generation. • Peptide-induced silver nanoparticles exhibit antibacterial activity.

  2. Conformational determinants of phosphotyrosine peptides complexed with the Src SH2 domain.

    Directory of Open Access Journals (Sweden)

    Joseph Nachman

    2010-06-01

    Full Text Available The inhibition of specific SH2 domain mediated protein-protein interactions as an effective chemotherapeutic approach in the treatment of diseases remains a challenge. That different conformations of peptide-ligands are preferred by different SH2 domains is an underappreciated observation from the structural analysis of phosphotyrosine peptide binding to SH2 domains that may aid in future drug design. To explore the nature of ligand binding, we use simulated annealing (SA to sample the conformational space of phosphotyrosine-containing peptides complexed with the Src SH2 domain. While in good agreement with the crystallographic and NMR studies of high-affinity phosphopeptide-SH2 domain complexes, the results suggest that the structural basis for phopsphopeptide- Src SH2 interactions is more complex than the "two-pronged plug two-hole socket" model. A systematic study of peptides of type pYEEX, where pY is phosphotyrosine and X is a hydrophobic residue, indicates that these peptides can assume two conformations, one extended and one helical, representing the balance between the interaction of residue X with the hydrophobic hole on the surface of the Src SH2 domain, and its contribution to the inherent tendency of the two glutamic acids to form an alpha-helix. In contrast, a beta-turn conformation, almost identical to that observed in the crystal structure of pYVNV bound to the Grb2 SH2 domain, predominates for pYXNX peptides, even in the presence of isoleucine at the third position. While peptide binding affinities, as measured by fluorescence polarization, correlate with the relative proportion of extended peptide conformation, these results suggest a model where all three residues C-terminal to the phosphotyrosine determine the conformation of the bound phosphopeptide. The information obtained in this work can be used in the design of specific SH2 domain inhibitors.

  3. On the performance of de novo pathway enrichment

    DEFF Research Database (Denmark)

    Batra, Richa; Alcaraz, Nicolas; Gitzhofer, Kevin

    2017-01-01

    De novo pathway enrichment is a powerful approach to discover previously uncharacterized molecular mechanisms in addition to already known pathways. To achieve this, condition-specific functional modules are extracted from large interaction networks. Here, we give an overview of the state...

  4. Purification and use of E. coli peptide deformylase for peptide deprotection in chemoenzymatic peptide synthesis

    NARCIS (Netherlands)

    Di Toma, Claudia; Sonke, Theo; Quaedflieg, Peter J.; Janssen, Dick B.

    Peptide deformylases (PDFs) catalyze the removal of the formyl group from the N-terminal methionine residue in nascent polypeptide chains in prokaryotes. Its deformylation activity makes PDF an attractive candidate for the biocatalytic deprotection of formylated peptides that are used in

  5. Dynamic peptide libraries for the discovery of supramolecular nanomaterials

    Science.gov (United States)

    Pappas, Charalampos G.; Shafi, Ramim; Sasselli, Ivan R.; Siccardi, Henry; Wang, Tong; Narang, Vishal; Abzalimov, Rinat; Wijerathne, Nadeesha; Ulijn, Rein V.

    2016-11-01

    Sequence-specific polymers, such as oligonucleotides and peptides, can be used as building blocks for functional supramolecular nanomaterials. The design and selection of suitable self-assembling sequences is, however, challenging because of the vast combinatorial space available. Here we report a methodology that allows the peptide sequence space to be searched for self-assembling structures. In this approach, unprotected homo- and heterodipeptides (including aromatic, aliphatic, polar and charged amino acids) are subjected to continuous enzymatic condensation, hydrolysis and sequence exchange to create a dynamic combinatorial peptide library. The free-energy change associated with the assembly process itself gives rise to selective amplification of self-assembling candidates. By changing the environmental conditions during the selection process, different sequences and consequent nanoscale morphologies are selected.

  6. Antioxidant activity of yoghurt peptides: Part 2 – Characterisationof peptide fractions

    DEFF Research Database (Denmark)

    Farvin, Sabeena; Baron, Caroline; Nielsen, Nina Skall

    2010-01-01

    the peptides identified contained at least one proline residue. Some of the identified peptides included the hydrophobic amino acid residues Val or Leu at the N-terminus and Pro, His or Tyr in the amino acid sequence, which is characteristic of antioxidant peptides. In addition, the yoghurt contained...

  7. Ligand-regulated peptide aptamers.

    Science.gov (United States)

    Miller, Russell A

    2009-01-01

    The peptide aptamer approach employs high-throughput selection to identify members of a randomized peptide library displayed from a scaffold protein by virtue of their interaction with a target molecule. Extending this approach, we have developed a peptide aptamer scaffold protein that can impart small-molecule control over the aptamer-target interaction. This ligand-regulated peptide (LiRP) scaffold, consisting of the protein domains FKBP12, FRB, and GST, binds to the cell-permeable small-molecule rapamycin and the binding of this molecule can prevent the interaction of the randomizable linker region connecting FKBP12 with FRB. Here we present a detailed protocol for the creation of a peptide aptamer plasmid library, selection of peptide aptamers using the LiRP scaffold in a yeast two-hybrid system, and the screening of those peptide aptamers for a ligand-regulated interaction.

  8. Transferência do fator caturra para o cultivar Mundo Novo de Coffea arabica Transfer of the CT gene to Mundo Novo cultivar

    Directory of Open Access Journals (Sweden)

    A. Carvalho

    1972-01-01

    Full Text Available No presente trabalho são relatados os estudos realizados visando à introdução do gene Ct (caturra que contribui para reduzir a altura da planta, no cultivar Mundo" Novo de Coffea arabica.Estudaram-se, em ensaios de produtividade, as populações Fv F.,, F3 e F4. Nessas populações e principalmente entre os descendentes dos "caféeiros H 2077-2-5 e H 2077-2-12, foram selecionadas plantas homozigotas para os alelos Ct e também para os alelos responsáveis pela cor do fruto xc ou Xc. Essas combinações foram denominadas 'Catuaí Amarelo' e 'Catuaí Vermelho', respectivamente, e suas características são apresentadas. Os novos cultivares vêm-se mostrando de interesse econômico para as regiões cafeeiras não somente pelo porte pequeno, mas também pela produtividade, pelo vigor vegetativo e pela precocidade.The successful transfer of the Ct gene for short internode to the tall cultivar of Coffea arábica'Mundo Novo' is reported. Individual selections were carried out in the F1, F2, F3 and F4 generations. It was found that early selection in the F2 generation was quite effective. A remarkably good correlation was found between productitivity of F2 plants and the yield of the F3 and F4 generations. Plants of the F4 generation have shown reasonable uniformity and high yield in several trials. The new selections showed to be early producers. Two new cultivars were released namely 'Catuaí Amarelo' and 'Catuaí Vermelho'. The former has yellow fruits whereas the latter has red fruits. The plants are much shorter that the ones of Mundo Novo. The new cultivars have a very strong secondary and tertiary branching. Because of these characteristics Catuaí Amarelo and Catuaí Vermelho are being planted in large scale replacing the tall cultivars.

  9. Multimerized CHR-derived peptides as HIV-1 fusion inhibitors.

    Science.gov (United States)

    Nomura, Wataru; Hashimoto, Chie; Suzuki, Takaharu; Ohashi, Nami; Fujino, Masayuki; Murakami, Tsutomu; Yamamoto, Naoki; Tamamura, Hirokazu

    2013-08-01

    To date, several HIV-1 fusion inhibitors based on the carboxy-terminal leucine/isoleucine heptad repeat (CHR) region of an HIV-1 envelope protein gp41 have been discovered. We have shown that a synthetic peptide mimetic of a trimer form of the CHR-derived peptide C34 has potent inhibitory activity against the HIV-1 fusion mechanism, compared to a monomer C34 peptide. The present study revealed that a dimeric form of C34 is evidently structurally critical for fusion inhibitors, and that the activity of multimerized CHR-derived peptides in fusion inhibition is affected by the properties of the unit peptides C34, SC34EK, and T20. The fluorescence-based study suggested that the N36-interactive sites of the C34 trimer, including hydrophobic residues, are exposed outside the trimer and that trimerization of C34 caused a remarkable increase in fusion inhibitory activity. The present results could be useful in the design of fusion inhibitors against viral infections which proceed via membrane fusion with host cells. Copyright © 2013 The Authors. Published by Elsevier Ltd.. All rights reserved.

  10. De Novo Glutamine Synthesis

    Science.gov (United States)

    He, Qiao; Shi, Xinchong; Zhang, Linqi; Yi, Chang; Zhang, Xuezhen

    2016-01-01

    Purpose: The aim of this study was to investigate the role of de novo glutamine (Gln) synthesis in the proliferation of C6 glioma cells and its detection with 13N-ammonia. Methods: Chronic Gln-deprived C6 glioma (0.06C6) cells were established. The proliferation rates of C6 and 0.06C6 cells were measured under the conditions of Gln deprivation along with or without the addition of ammonia or glutamine synthetase (GS) inhibitor. 13N-ammonia uptake was assessed in C6 cells by gamma counting and in rats with C6 and 0.06C6 xenografts by micro–positron emission tomography (PET) scanning. The expression of GS in C6 cells and xenografts was assessed by Western blotting and immunohistochemistry, respectively. Results: The Gln-deprived C6 cells showed decreased proliferation ability but had a significant increase in GS expression. Furthermore, we found that low concentration of ammonia was sufficient to maintain the proliferation of Gln-deprived C6 cells, and 13N-ammonia uptake in C6 cells showed Gln-dependent decrease, whereas inhibition of GS markedly reduced the proliferation of C6 cells as well as the uptake of 13N-ammoina. Additionally, microPET/computed tomography exhibited that subcutaneous 0.06C6 xenografts had higher 13N-ammonia uptake and GS expression in contrast to C6 xenografts. Conclusion: De novo Gln synthesis through ammonia–glutamate reaction plays an important role in the proliferation of C6 cells. 13N-ammonia can be a potential metabolic PET tracer for Gln-dependent tumors. PMID:27118759

  11. Antibacterial activity of synthetic peptides derived from lactoferricin against Escherichia coli ATCC 25922 and Enterococcus faecalis ATCC 29212.

    Science.gov (United States)

    León-Calvijo, María A; Leal-Castro, Aura L; Almanzar-Reina, Giovanni A; Rosas-Pérez, Jaiver E; García-Castañeda, Javier E; Rivera-Monroy, Zuly J

    2015-01-01

    Peptides derived from human and bovine lactoferricin were designed, synthesized, purified, and characterized using RP-HPLC and MALDI-TOF-MS. Specific changes in the sequences were designed as (i) the incorporation of unnatural amino acids in the sequence, the (ii) reduction or (iii) elongation of the peptide chain length, and (iv) synthesis of molecules with different number of branches containing the same sequence. For each peptide, the antibacterial activity against Escherichia coli ATCC 25922 and Enterococcus faecalis ATCC 29212 was evaluated. Our results showed that Peptides I.2 (RWQWRWQWR) and I.4 ((RRWQWR)4K2Ahx2C2) exhibit bigger or similar activity against E. coli (MIC 4-33 μM) and E. faecalis (MIC 10-33 μM) when they were compared with lactoferricin protein (LF) and some of its derivate peptides as II.1 (FKCRRWQWRMKKLGA) and IV.1 (FKCRRWQWRMKKLGAPSITCVRRAE). It should be pointed out that Peptides I.2 and I.4, containing the RWQWR motif, are short and easy to synthesize; our results demonstrate that it is possible to design and obtain synthetic peptides that exhibit enhanced antibacterial activity using a methodology that is fast and low-cost and that allows obtaining products with a high degree of purity and high yield.

  12. Antibacterial Activity of Synthetic Peptides Derived from Lactoferricin against Escherichia coli ATCC 25922 and Enterococcus faecalis ATCC 29212

    Directory of Open Access Journals (Sweden)

    María A. León-Calvijo

    2015-01-01

    Full Text Available Peptides derived from human and bovine lactoferricin were designed, synthesized, purified, and characterized using RP-HPLC and MALDI-TOF-MS. Specific changes in the sequences were designed as (i the incorporation of unnatural amino acids in the sequence, the (ii reduction or (iii elongation of the peptide chain length, and (iv synthesis of molecules with different number of branches containing the same sequence. For each peptide, the antibacterial activity against Escherichia coli ATCC 25922 and Enterococcus faecalis ATCC 29212 was evaluated. Our results showed that Peptides I.2 (RWQWRWQWR and I.4 ((RRWQWR4K2Ahx2C2 exhibit bigger or similar activity against E. coli (MIC 4–33 μM and E. faecalis (MIC 10–33 μM when they were compared with lactoferricin protein (LF and some of its derivate peptides as II.1 (FKCRRWQWRMKKLGA and IV.1 (FKCRRWQWRMKKLGAPSITCVRRAE. It should be pointed out that Peptides I.2 and I.4, containing the RWQWR motif, are short and easy to synthesize; our results demonstrate that it is possible to design and obtain synthetic peptides that exhibit enhanced antibacterial activity using a methodology that is fast and low-cost and that allows obtaining products with a high degree of purity and high yield.

  13. De Novo Discovery of Structured ncRNA Motifs in Genomic Sequences

    DEFF Research Database (Denmark)

    Ruzzo, Walter L; Gorodkin, Jan

    2014-01-01

    De novo discovery of "motifs" capturing the commonalities among related noncoding ncRNA structured RNAs is among the most difficult problems in computational biology. This chapter outlines the challenges presented by this problem, together with some approaches towards solving them, with an emphas...... on an approach based on the CMfinder CMfinder program as a case study. Applications to genomic screens for novel de novo structured ncRNA ncRNA s, including structured RNA elements in untranslated portions of protein-coding genes, are presented.......De novo discovery of "motifs" capturing the commonalities among related noncoding ncRNA structured RNAs is among the most difficult problems in computational biology. This chapter outlines the challenges presented by this problem, together with some approaches towards solving them, with an emphasis...

  14. A distributive peptide cyclase processes multiple microviridin core peptides within a single polypeptide substrate.

    Science.gov (United States)

    Zhang, Yi; Li, Kunhua; Yang, Guang; McBride, Joshua L; Bruner, Steven D; Ding, Yousong

    2018-05-03

    Ribosomally synthesized and post-translationally modified peptides (RiPPs) are an important family of natural products. Their biosynthesis follows a common scheme in which the leader peptide of a precursor peptide guides the modifications of a single core peptide. Here we describe biochemical studies of the processing of multiple core peptides within a precursor peptide, rare in RiPP biosynthesis. In a cyanobacterial microviridin pathway, an ATP-grasp ligase, AMdnC, installs up to two macrolactones on each of the three core peptides within AMdnA. The enzyme catalysis occurs in a distributive fashion and follows an unstrict N-to-C overall directionality, but a strict order in macrolactonizing each core peptide. Furthermore, AMdnC is catalytically versatile to process unnatural substrates carrying one to four core peptides, and kinetic studies provide insights into its catalytic properties. Collectively, our results reveal a distinct biosynthetic logic of RiPPs, opening up the possibility of modular production via synthetic biology approaches.

  15. Superior Antifouling Performance of a Zwitterionic Peptide Compared to an Amphiphilic, Non-Ionic Peptide.

    Science.gov (United States)

    Ye, Huijun; Wang, Libing; Huang, Renliang; Su, Rongxin; Liu, Boshi; Qi, Wei; He, Zhimin

    2015-10-14

    The aim of this study was to explore the influence of amphiphilic and zwitterionic structures on the resistance of protein adsorption to peptide self-assembled monolayers (SAMs) and gain insight into the associated antifouling mechanism. Two kinds of cysteine-terminated heptapeptides were studied. One peptide had alternating hydrophobic and hydrophilic residues with an amphiphilic sequence of CYSYSYS. The other peptide (CRERERE) was zwitterionic. Both peptides were covalently attached onto gold substrates via gold-thiol bond formation. Surface plasmon resonance analysis results showed that both peptide SAMs had ultralow or low protein adsorption amounts of 1.97-11.78 ng/cm2 in the presence of single proteins. The zwitterionic peptide showed relatively higher antifouling ability with single proteins and natural complex protein media. We performed molecular dynamics simulations to understand their respective antifouling behaviors. The results indicated that strong surface hydration of peptide SAMs contributes to fouling resistance by impeding interactions with proteins. Compared to the CYSYSYS peptide, more water molecules were predicted to form hydrogen-bonding interactions with the zwitterionic CRERERE peptide, which is in agreement with the antifouling test results. These findings reveal a clear relation between peptide structures and resistance to protein adsorption, facilitating the development of novel peptide-containing antifouling materials.

  16. in silico Whole Genome Sequencer & Analyzer (iWGS): a computational pipeline to guide the design and analysis of de novo genome sequencing studies

    Science.gov (United States)

    The availability of genomes across the tree of life is highly biased toward vertebrates, pathogens, human disease models, and organisms with relatively small and simple genomes. Recent progress in genomics has enabled the de novo decoding of the genome of virtually any organism, greatly expanding it...

  17. Antimicrobial Peptides in Reptiles

    Science.gov (United States)

    van Hoek, Monique L.

    2014-01-01

    Reptiles are among the oldest known amniotes and are highly diverse in their morphology and ecological niches. These animals have an evolutionarily ancient innate-immune system that is of great interest to scientists trying to identify new and useful antimicrobial peptides. Significant work in the last decade in the fields of biochemistry, proteomics and genomics has begun to reveal the complexity of reptilian antimicrobial peptides. Here, the current knowledge about antimicrobial peptides in reptiles is reviewed, with specific examples in each of the four orders: Testudines (turtles and tortosises), Sphenodontia (tuataras), Squamata (snakes and lizards), and Crocodilia (crocodilans). Examples are presented of the major classes of antimicrobial peptides expressed by reptiles including defensins, cathelicidins, liver-expressed peptides (hepcidin and LEAP-2), lysozyme, crotamine, and others. Some of these peptides have been identified and tested for their antibacterial or antiviral activity; others are only predicted as possible genes from genomic sequencing. Bioinformatic analysis of the reptile genomes is presented, revealing many predicted candidate antimicrobial peptides genes across this diverse class. The study of how these ancient creatures use antimicrobial peptides within their innate immune systems may reveal new understandings of our mammalian innate immune system and may also provide new and powerful antimicrobial peptides as scaffolds for potential therapeutic development. PMID:24918867

  18. Peptides Displayed as High Density Brush Polymers Resist Proteolysis and Retain Bioactivity

    Science.gov (United States)

    2015-01-01

    We describe a strategy for rendering peptides resistant to proteolysis by formulating them as high-density brush polymers. The utility of this approach is demonstrated by polymerizing well-established cell-penetrating peptides (CPPs) and showing that the resulting polymers are not only resistant to proteolysis but also maintain their ability to enter cells. The scope of this design concept is explored by studying the proteolytic resistance of brush polymers composed of peptides that are substrates for either thrombin or a metalloprotease. Finally, we demonstrate that the proteolytic susceptibility of peptide brush polymers can be tuned by adjusting the density of the polymer brush and offer in silico models to rationalize this finding. We contend that this strategy offers a plausible method of preparing peptides for in vivo use, where rapid digestion by proteases has traditionally restricted their utility. PMID:25314576

  19. In silico panning for a non-competitive peptide inhibitor

    Directory of Open Access Journals (Sweden)

    Ikebukuro Kazunori

    2007-01-01

    Full Text Available Abstract Background Peptide ligands have tremendous therapeutic potential as efficacious drugs. Currently, more than 40 peptides are available in the market for a drug. However, since costly and time-consuming synthesis procedures represent a problem for high-throughput screening, novel procedures to reduce the time and labor involved in screening peptide ligands are required. We propose the novel approach of 'in silico panning' which consists of a two-stage screening, involving affinity selection by docking simulation and evolution of the peptide ligand using genetic algorithms (GAs. In silico panning was successfully applied to the selection of peptide inhibitor for water-soluble quinoprotein glucose dehydrogenase (PQQGDH. Results The evolution of peptide ligands for a target enzyme was achieved by combining a docking simulation with evolution of the peptide ligand using genetic algorithms (GAs, which mimic Darwinian evolution. Designation of the target area as next to the substrate-binding site of the enzyme in the docking simulation enabled the selection of a non-competitive inhibitor. In all, four rounds of selection were carried out on the computer; the distribution of the docking energy decreased gradually for each generation and improvements in the docking energy were observed over the four rounds of selection. One of the top three selected peptides with the lowest docking energy, 'SERG' showed an inhibitory effect with Ki value of 20 μM. PQQGDH activity, in terms of the Vmax value, was 3-fold lower than that of the wild-type enzyme in the presence of this peptide. The mechanism of the SERG blockage of the enzyme was identified as non-competitive inhibition. We confirmed the specific binding of the peptide, and its equilibrium dissociation constant (KD value was calculated as 60 μM by surface plasmon resonance (SPR analysis. Conclusion We demonstrate an effective methodology of in silico panning for the selection of a non

  20. Isoform-Selective Disruption of AKAP-Localized PKA Using Hydrocarbon Stapled Peptides

    Science.gov (United States)

    2015-01-01

    A-kinase anchoring proteins (AKAPs) play an important role in the spatial and temporal regulation of protein kinase A (PKA) by scaffolding critical intracellular signaling complexes. Here we report the design of conformationally constrained peptides that disrupt interactions between PKA and AKAPs in an isoform-selective manner. Peptides derived from the A Kinase Binding (AKB) domain of several AKAPs were chemically modified to contain an all-hydrocarbon staple and target the docking/dimerization domain of PKA-R, thereby occluding AKAP interactions. The peptides are cell-permeable against diverse human cell lines, are highly isoform-selective for PKA-RII, and can effectively inhibit interactions between AKAPs and PKA-RII in intact cells. These peptides can be applied as useful reagents in cell-based studies to selectively disrupt AKAP-localized PKA-RII activity and block AKAP signaling complexes. In summary, the novel hydrocarbon-stapled peptides developed in this study represent a new class of AKAP disruptors to study compartmentalized RII-regulated PKA signaling in cells. PMID:24422448

  1. Synthesis and evaluation of amphiphilic peptides as nanostructures and drug delivery tools

    Science.gov (United States)

    Sayeh, Naser Ali

    conjugates although one limitation lies in the effort of controlling the rate of drug release. The encapsulated or complexed drugs tend to be released rapidly (before reaching the target site) and in the dendrimer--drug conjugates, it is the chemical linkage that controls the drug release. Thus, future studies in this field are urgently required to create more efficient and stable biomaterials. Peptides are considered as efficient vectors for achieving optimal cellular uptake. The potential use of peptides as drug delivery vectors received much attention by the discovery of several cell-penetrating peptides (CPPs). The first CPPs discovered in 1988, that were sequences from HIV-1 encoded TAT protein, TAT (48--60), and penetrated very efficiently through cell membranes of cultured mammalian cells. CPPs are a class of diverse peptides, typically with 8--25 amino acids, and unlike most peptides, they can cross the cellular membrane with more efficiency. CPPs have also shown to undergo self-assembly and generate nanostructures. The generation of self-assembled peptides and nanostructures occur through various types of interactions between functional groups of amino acid residues, such as electrostatic, hydrophobic, and hydrogen bonding. Appropriate design and functionalization of peptides are critical for generating nanostructures. Chemically CPPs are classified into two major groups: linear and cyclic peptides. It has been previously reported that linear peptides containing hydrophilic and hydrophobic amino acids could act as membrane protein stabilizers. These compounds are short hydrophilic or amphiphilic peptides that have positively charged amino acids, such as arginine, lysine or histidine, which can interact with the negative charge phospholipids layer on the cell membrane and translocate the cargo into the cells. Conjugation to cationic linear CPPs, such as TAT, penetratin, or oligoarginine efficiently improves the cellular uptake of large hydrophilic molecules, but the

  2. Sub-nanometer-resolution imaging of peptide nanotubes in water using frequency modulation atomic force microscopy

    Energy Technology Data Exchange (ETDEWEB)

    Sugihara, Tomoki; Hayashi, Itsuho; Onishi, Hiroshi [Department of Chemistry, Graduate School of Science, Kobe University, 1-1 Rokkodai-cho, Nada-ku, Kobe 657-8501 (Japan); Kimura, Kenjiro, E-mail: kimura@gold.kobe-u.ac.jp [Department of Chemistry, Graduate School of Science, Kobe University, 1-1 Rokkodai-cho, Nada-ku, Kobe 657-8501 (Japan); Tamura, Atsuo [Department of Chemistry, Graduate School of Science, Kobe University, 1-1 Rokkodai-cho, Nada-ku, Kobe 657-8501 (Japan)

    2013-06-20

    Highlights: ► Peptide nanotubes were aligned on highly oriented pyrolytic graphite surface. ► We visualized sub-nanometer-scale structure on peptide nanotube surface in water. ► We observed hydration structure at a peptide nanotube/water interface. - Abstract: Peptide nanotubes are self-assembled fibrous materials composed of cyclic polypeptides. Recently, various aspects of peptide nanotubes have been studied, in particular the utility of different methods for making peptide nanotubes with diverse designed functions. In order to investigate the relationship between formation, function and stability, it is essential to analyze the precise structure of peptide nanotubes. Atomic-scale surface imaging in liquids was recently achieved using frequency modulation atomic force microscopy with improved force sensing. Here we provide a precise surface structural analysis of peptide nanotubes in water without crystallizing them obtained by imaging the nanotubes at the sub-nanometer scale in water. In addition, the local hydration structure around the peptide nanotubes was observed at the nanotube/water interface.

  3. Novel heparan sulfate-binding peptides for blocking herpesvirus entry.

    Directory of Open Access Journals (Sweden)

    Pranay Dogra

    Full Text Available Human cytomegalovirus (HCMV infection can lead to congenital hearing loss and mental retardation. Upon immune suppression, reactivation of latent HCMV or primary infection increases morbidity in cancer, transplantation, and late stage AIDS patients. Current treatments include nucleoside analogues, which have significant toxicities limiting their usefulness. In this study we screened a panel of synthetic heparin-binding peptides for their ability to prevent CMV infection in vitro. A peptide designated, p5+14 exhibited ~ 90% reduction in murine CMV (MCMV infection. Because negatively charged, cell-surface heparan sulfate proteoglycans (HSPGs, serve as the attachment receptor during the adsorption phase of the CMV infection cycle, we hypothesized that p5+14 effectively competes for CMV adsorption to the cell surface resulting in the reduction in infection. Positively charged Lys residues were required for peptide binding to cell-surface HSPGs and reducing viral infection. We show that this inhibition was not due to a direct neutralizing effect on the virus itself and that the peptide blocked adsorption of the virus. The peptide also inhibited infection of other herpesviruses: HCMV and herpes simplex virus 1 and 2 in vitro, demonstrating it has broad-spectrum antiviral activity. Therefore, this peptide may offer an adjunct therapy for the treatment of herpes viral infections and other viruses that use HSPGs for entry.

  4. Driving engineering of novel antimicrobial peptides from simulations of peptide-micelle interactions

    DEFF Research Database (Denmark)

    Khandelia, Himanshu; Langham, Allison A; Kaznessis, Yiannis N

    2006-01-01

    Simulations of antimicrobial peptides in membrane mimics can provide the high resolution, atomistic picture that is necessary to decipher which sequence and structure components are responsible for activity and toxicity. With such detailed insight, engineering new sequences that are active but non...... peptides and their interaction with membrane mimics. In this article, we discuss the promise and the challenges of widely used models and detail our recent work on peptide-micelle simulations as an attractive alternative to peptide-bilayer simulations. We detail our results with two large structural...... classes of peptides, helical and beta-sheet and demonstrate how simulations can assist in engineering of novel antimicrobials with therapeutic potential....

  5. The effect of a beta-lactamase inhibitor peptide on bacterial membrane structure and integrity: a comparative study.

    Science.gov (United States)

    Alaybeyoglu, Begum; Uluocak, Bilge Gedik; Akbulut, Berna Sariyar; Ozkirimli, Elif

    2017-05-01

    Co-administration of beta-lactam antibiotics and beta-lactamase inhibitors has been a favored treatment strategy against beta-lactamase-mediated bacterial antibiotic resistance, but the emergence of beta-lactamases resistant to current inhibitors necessitates the discovery of novel non-beta-lactam inhibitors. Peptides derived from the Ala46-Tyr51 region of the beta-lactamase inhibitor protein are considered as potent inhibitors of beta-lactamase; unfortunately, peptide delivery into the cell limits their potential. The properties of cell-penetrating peptides could guide the design of beta-lactamase inhibitory peptides. Here, our goal is to modify the peptide with the sequence RRGHYY that possesses beta-lactamase inhibitory activity under in vitro conditions. Inspired by the work on the cell-penetrating peptide pVEC, our approach involved the addition of the N-terminal hydrophobic residues, LLIIL, from pVEC to the inhibitor peptide to build a chimera. These residues have been reported to be critical in the uptake of pVEC. We tested the potential of RRGHYY and its chimeric derivative as a beta-lactamase inhibitory peptide on Escherichia coli cells and compared the results with the action of the antimicrobial peptide melittin, the beta-lactam antibiotic ampicillin, and the beta-lactamase inhibitor potassium clavulanate to get mechanistic details on their action. Our results show that the addition of LLIIL to the N-terminus of the beta-lactamase inhibitory peptide RRGHYY increases its membrane permeabilizing potential. Interestingly, the addition of this short stretch of hydrophobic residues also modified the inhibitory peptide such that it acquired antimicrobial property. We propose that addition of the hydrophobic LLIIL residues to the peptide N-terminus offers a promising strategy to design novel antimicrobial peptides in the battle against antibiotic resistance. Copyright © 2017 European Peptide Society and John Wiley & Sons, Ltd. Copyright © 2017 European

  6. Peptide Pattern Recognition for high-throughput protein sequence analysis and clustering

    DEFF Research Database (Denmark)

    Busk, Peter Kamp

    2017-01-01

    Large collections of protein sequences with divergent sequences are tedious to analyze for understanding their phylogenetic or structure-function relation. Peptide Pattern Recognition is an algorithm that was developed to facilitate this task but the previous version does only allow a limited...... number of sequences as input. I implemented Peptide Pattern Recognition as a multithread software designed to handle large numbers of sequences and perform analysis in a reasonable time frame. Benchmarking showed that the new implementation of Peptide Pattern Recognition is twenty times faster than...... the previous implementation on a small protein collection with 673 MAP kinase sequences. In addition, the new implementation could analyze a large protein collection with 48,570 Glycosyl Transferase family 20 sequences without reaching its upper limit on a desktop computer. Peptide Pattern Recognition...

  7. Use of artificial intelligence in the design of small peptide antibiotics effective against a broad spectrum of highly antibiotic-resistant superbugs.

    Science.gov (United States)

    Cherkasov, Artem; Hilpert, Kai; Jenssen, Håvard; Fjell, Christopher D; Waldbrook, Matt; Mullaly, Sarah C; Volkmer, Rudolf; Hancock, Robert E W

    2009-01-16

    Increased multiple antibiotic resistance in the face of declining antibiotic discovery is one of society's most pressing health issues. Antimicrobial peptides represent a promising new class of antibiotics. Here we ask whether it is possible to make small broad spectrum peptides employing minimal assumptions, by capitalizing on accumulating chemical biology information. Using peptide array technology, two large random 9-amino-acid peptide libraries were iteratively created using the amino acid composition of the most active peptides. The resultant data was used together with Artificial Neural Networks, a powerful machine learning technique, to create quantitative in silico models of antibiotic activity. On the basis of random testing, these models proved remarkably effective in predicting the activity of 100,000 virtual peptides. The best peptides, representing the top quartile of predicted activities, were effective against a broad array of multidrug-resistant "Superbugs" with activities that were equal to or better than four highly used conventional antibiotics, more effective than the most advanced clinical candidate antimicrobial peptide, and protective against Staphylococcus aureus infections in animal models.

  8. Novo Jornalismo: fronteiras litero-factuais em A sangue Frio e em Radical Chique

    Directory of Open Access Journals (Sweden)

    Francisco Aquinei Timóteo Queirós

    2012-03-01

    Full Text Available http://dx.doi.org/10.5007/1984-784X.2012v12n18p130 A pesquisa busca analisar de que forma fato e ficção se entrecruzam no “movimento” do Novo Jornalismo, a partir das obras A sangue Frio e Radical Chique e o Novo Jornalismo, de Truman Capote e Tom Wolfe, respectivamente. Pretende-se, a partir da investigação do corpus em estudo, revelar os aspectos que aproximam o fato jornalístico, a notícia e a reportagem às técnicas literárias do romance, do conto e da crônica. O estudo investiga o Novo Jornalismo sob o viés de textos centrais das áreas de teoria literária e estudos jornalísticos utilizando autores como Mikhail Bakhtin, Hayden White, Paul Ricoeur, Muniz Sodré; além de referenciar outros escritores que, como Tom Wolfe e Truman Capote, fizeram parte de um grande movimento renovador do jornalismo literário nos anos 1950, 1960 e 1970 chamado, genericamente, de Novo Jornalismo.

  9. Facebook - Um novo espaço autobiográfico?

    Directory of Open Access Journals (Sweden)

    Maria Tereza Lima

    2015-07-01

    Full Text Available O arigo "Facebook - Um novo espaço autobiográfico?" tem como objetivo central investigar como a perspectiva autobiográfica e biográfica se configura em uma rede social. Levando em consideração esse novo espaço de exteriorização da memória, analisamos as escolhas de uma pessoa ao postar os mais diversos gêneros textuais no Facebook e verificamos até que ponto tais fragmentos textuais narram a história de um indivíduo. Quais textos são postados? O que foi escolhido e o que foi excluído desse perfil? O autor trava um pacto de leitura com o leitor? Se levarmos em consideração que os textos postados nessa rede social são textos produzidos pelo próprio autor do perfil e de autores diversos, como configuraremos esses espaços virtuais? Autobiográficos e biográficos? Quem escreve a página virtual é o próprio autor do perfil ou múltiplos autores? Com as redes sociais, surge um novo modelo de autobiografia e de biógrafo? Esses e tantos outros questionamentos nortearam nossas investigações e permitiram-nos conhecer um pouco mais sobre as estratégias autobiográficas dos autores virtuais contemporâneos.

  10. Structure-Based Drug Design of Small Molecule Peptide Deformylase Inhibitors to Treat Cancer

    Directory of Open Access Journals (Sweden)

    Jian Gao

    2016-03-01

    Full Text Available Human peptide deformylase (HsPDF is an important target for anticancer drug discovery. In view of the limited HsPDF, inhibitors were reported, and high-throughput virtual screening (HTVS studies based on HsPDF for developing new PDF inhibitors remain to be reported. We reported here on diverse small molecule inhibitors with excellent anticancer activities designed based on HTVS and molecular docking studies using the crystal structure of HsPDF. The compound M7594_0037 exhibited potent anticancer activities against HeLa, A549 and MCF-7 cell lines with IC50s of 35.26, 29.63 and 24.63 μM, respectively. Molecular docking studies suggested that M7594_0037 and its three derivatives could interact with HsPDF by several conserved hydrogen bonds. Moreover, the pharmacokinetic and toxicity properties of M7594_0037 and its derivatives were predicted using the OSIRIS property explorer. Thus, M7594_0037 and its derivatives might represent a promising scaffold for the further development of novel anticancer drugs.

  11. Multivalent display of the antimicrobial peptides BP100 and BP143

    Directory of Open Access Journals (Sweden)

    Imma Güell

    2012-12-01

    Full Text Available Carbohydrates are considered as promising templates for the display of multiple copies of antimicrobial peptides. Herein, we describe the design and synthesis of chimeric structures containing two or four copies of the antimicrobial peptides KKLFKKILKYL-NH2 (BP100 and KKLfKKILKYL-NH2 (BP143 attached to the carbohydrate template cyclodithioerythritol (cDTE or α-D-galactopyranoside (Galp. The synthesis involved the preparation of the corresponding peptide aldehyde followed by coupling to an aminooxy-functionalized carbohydrate template. After purification, the multivalent display systems were obtained in high purities (90–98% and in good yields (42–64%. These compounds were tested against plant and human pathogenic bacteria and screened for their cytotoxicity on eukaryotic cells. They showed lower MIC values than the parent peptides against the bacteria analyzed. In particular, the carbopeptides derived from cDTE and Galp, which contained two or four copies of BP100, respectively, were 2- to 8-fold more active than the monomeric peptide against the phytopathogenic bacteria. These results suggest that preassembling antimicrobial peptides to multimeric structures is not always associated with a significant improvement of the activity. In contrast, the carbopeptides synthesized were active against human red blood cells pointing out that peptide preassembly is critical for the hemolytic activity. Notably, peptide preassembly resulted in an enhanced bactericidal effect.

  12. Advances in the design and higher-order assembly of collagen mimetic peptides for regenerative medicine.

    Science.gov (United States)

    Strauss, Kevin; Chmielewski, Jean

    2017-08-01

    Regenerative medicine makes use of cell-supporting biomaterials to replace lost or damaged tissue. Collagen holds great potential in this regard caused by its biocompatibility and structural versatility. While natural collagen has shown promise for regenerative medicine, collagen mimetic peptides (CMPs) have emerged that allow far higher degrees of customization and ease of preparation. A wide range of two and three-dimensional assemblies have been generated from CMPs, many of which accommodate cellular adhesion and encapsulation, through careful sequence design and the exploitation of electrostatic and hydrophobic forces. But the methodology that has generated the greatest plethora of viable biomaterials is metal-promoted assembly of CMP triple helices-a rapid process that occurs under physiological conditions. Architectures generated in this manner promote cell growth, enable directed attachment of bioactive cargo, and produce living tissue. Copyright © 2017 Elsevier Ltd. All rights reserved.

  13. Proteolysis targeting peptide (PROTAP) strategy for protein ubiquitination and degradation.

    Science.gov (United States)

    Zheng, Jing; Tan, Chunyan; Xue, Pengcheng; Cao, Jiakun; Liu, Feng; Tan, Ying; Jiang, Yuyang

    2016-02-19

    Ubiquitination proteasome pathway (UPP) is the most important and selective way to degrade proteins in vivo. Here, a novel proteolysis targeting peptide (PROTAP) strategy, composed of a target protein binding peptide, a linker and a ubiquitin E3 ligase recognition peptide, was designed to recruit both target protein and E3 ligase and then induce polyubiquitination and degradation of the target protein through UPP. In our study, the PROTAP strategy was proved to be a general method with high specificity using Bcl-xL protein as model target in vitro and in cells, which indicates that the strategy has great potential for in vivo application. Copyright © 2016 Elsevier Inc. All rights reserved.

  14. Orally active-targeted drug delivery systems for proteins and peptides.

    Science.gov (United States)

    Li, Xiuying; Yu, Miaorong; Fan, Weiwei; Gan, Yong; Hovgaard, Lars; Yang, Mingshi

    2014-09-01

    In the past decade, extensive efforts have been devoted to designing 'active targeted' drug delivery systems (ATDDS) to improve oral absorption of proteins and peptides. Such ATDDS enhance cellular internalization and permeability of proteins and peptides via molecular recognition processes such as ligand-receptor or antigen-antibody interaction, and thus enhance drug absorption. This review focuses on recent advances with orally ATDDS, including ligand-protein conjugates, recombinant ligand-protein fusion proteins and ligand-modified carriers. In addition to traditional intestinal active transport systems of substrates and their corresponding receptors, transporters and carriers, new targets such as intercellular adhesion molecule-1 and β-integrin are also discussed. ATDDS can improve oral absorption of proteins and peptides. However, currently, no clinical studies on ATDDS for proteins and peptides are underway, perhaps due to the complexity and limited knowledge of transport mechanisms. Therefore, more research is warranted to optimize ATDDS efficiency.

  15. De Novo Heart Failure After Kidney Transplantation: Trends in Incidence and Outcomes.

    Science.gov (United States)

    Lenihan, Colin R; Liu, Sai; Deswal, Anita; Montez-Rath, Maria E; Winkelmayer, Wolfgang C

    2018-03-29

    Heart failure is an important cause of morbidity and mortality following kidney transplantation. Some studies in the general population have shown that the incidence of heart failure has decreased during the past 20 years. However, it is not currently known whether such a trend exists in the kidney transplantation population. Retrospective observational cohort study. Adult patients included in the US Renal Data System who underwent their first kidney transplantation in the United States between 1998 and 2010 with at least 6 months of continuous Medicare parts A and B coverage before transplantation and no prior evidence for a diagnosis of heart failure before kidney transplantation. Calendar year of transplantation and calendar year of posttransplantation heart failure diagnosis. De novo posttransplantation heart failure defined using International Classification of Diseases, Ninth Revision diagnosis codes and mortality following de novo posttransplantation heart failure diagnosis. Secular trends in de novo post-kidney transplantation heart failure were examined using Cox proportional hazards analysis. Within a study cohort of 48,771 patients, 7,269 developed de novo heart failure within 3 years of kidney transplantation, with a median time to heart failure of 0.76 years. The adjusted HR for heart failure with death as competing risk comparing patients who underwent transplantation in 2010 with those who underwent transplantation in 1998 was 0.69 (95% CI, 0.60-0.79). No temporal trend in mortality following a diagnosis of post-kidney transplantation heart failure was observed. Potential residual confounding from either incorrectly ascertained or unavailable confounders. The cohort was limited to Medicare beneficiaries. Adjusted for demographic and clinical characteristics, the risk for developing de novo post-kidney transplantation heart failure has declined significantly between 1998 and 2010, with no apparent change in subsequent mortality. Copyright © 2018

  16. Solid-Binding Peptides in Biomedicine.

    Science.gov (United States)

    Care, Andrew; Bergquist, Peter L; Sunna, Anwar

    2017-01-01

    Some peptides are able to bind to inorganic materials such as silica and gold. Over the past decade, Solid-binding peptides (SBPs) have been used increasingly as molecular building blocks in nanobiotechnology. These peptides show selectivity and bind with high affinity to a diverse range of inorganic surfaces e.g. metals, metal oxides, metal compounds, magnetic materials, semiconductors, carbon materials, polymers and minerals. They can be used in applications such as protein purification and synthesis, assembly and the functionalization of nanomaterials. They offer simple and versatile bioconjugation methods that can increase biocompatibility and also direct the immobilization and orientation of nanoscale entities onto solid supports without impeding their functionality. SBPs have been employed in numerous nanobiotechnological applications such as the controlled synthesis of nanomaterials and nanostructures, formation of hybrid biomaterials, immobilization of functional proteins and improved nanomaterial biocompatibility. With advances in nanotechnology, a multitude of novel nanomaterials have been designed and synthesized for diagnostic and therapeutic applications. New approaches have been developed recently to exert a greater control over bioconjugation and eventually, over the optimal and functional display of biomolecules on the surfaces of many types of solid materials. In this chapter we describe SBPs and highlight some selected examples of their potential applications in biomedicine.

  17. Flanking signal and mature peptide residues influence signal peptide cleavage

    Directory of Open Access Journals (Sweden)

    Ranganathan Shoba

    2008-12-01

    Full Text Available Abstract Background Signal peptides (SPs mediate the targeting of secretory precursor proteins to the correct subcellular compartments in prokaryotes and eukaryotes. Identifying these transient peptides is crucial to the medical, food and beverage and biotechnology industries yet our understanding of these peptides remains limited. This paper examines the most common type of signal peptides cleavable by the endoprotease signal peptidase I (SPase I, and the residues flanking the cleavage sites of three groups of signal peptide sequences, namely (i eukaryotes (Euk (ii Gram-positive (Gram+ bacteria, and (iii Gram-negative (Gram- bacteria. Results In this study, 2352 secretory peptide sequences from a variety of organisms with amino-terminal SPs are extracted from the manually curated SPdb database for analysis based on physicochemical properties such as pI, aliphatic index, GRAVY score, hydrophobicity, net charge and position-specific residue preferences. Our findings show that the three groups share several similarities in general, but they display distinctive features upon examination in terms of their amino acid compositions and frequencies, and various physico-chemical properties. Thus, analysis or prediction of their sequences should be separated and treated as distinct groups. Conclusion We conclude that the peptide segment recognized by SPase I extends to the start of the mature protein to a limited extent, upon our survey of the amino acid residues surrounding the cleavage processing site. These flanking residues possibly influence the cleavage processing and contribute to non-canonical cleavage sites. Our findings are applicable in defining more accurate prediction tools for recognition and identification of cleavage site of SPs.

  18. Analysis of peptide uptake and location of root hair-promoting peptide accumulation in plant roots.

    Science.gov (United States)

    Matsumiya, Yoshiki; Taniguchi, Rikiya; Kubo, Motoki

    2012-03-01

    Peptide uptake by plant roots from degraded soybean-meal products was analyzed in Brassica rapa and Solanum lycopersicum. B. rapa absorbed about 40% of the initial water volume, whereas peptide concentration was decreased by 75% after 24 h. Analysis by reversed-phase HPLC showed that number of peptides was absorbed by the roots during soaking in degraded soybean-meal products for 24 h. Carboxyfluorescein-labeled root hair-promoting peptide was synthesized, and its localization, movement, and accumulation in roots were investigated. The peptide appeared to be absorbed by root hairs and then moved to trichoblasts. Furthermore, the peptide was moved from trichoblasts to atrichoblasts after 24 h. The peptide was accumulated in epidermal cells, suggesting that the peptide may have a function in both trichoblasts and atrichoblasts. Copyright © 2012 European Peptide Society and John Wiley & Sons, Ltd.

  19. Design and studies of multiple mechanism of anti-Candida activity of a new potent Trp-rich peptide dendrimers.

    Science.gov (United States)

    Zielińska, Paulina; Staniszewska, Monika; Bondaryk, Małgorzata; Koronkiewicz, Mirosława; Urbańczyk-Lipkowska, Zofia

    2015-11-13

    Eight peptide dendrimers were designed as structural mimics of natural cationic amphiphilic peptides with antifungal activity and evaluated for their anti-Candida potential against the wild type strains and mutants. Dendrimer 14 containing four Trp residues and dodecyl tail and a slightly smaller dendrimer 9 decorated with four N-methylated Trp that displayed 100 and 99.7% of growth inhibition at 16 μg/mL respectively, were selected for evaluation against the Candida albicans mutants with disabled biosynthesis of aspartic proteases responsible for host tissue colonization and morphogenesis during biofilm formation (sessile model). Flow cytometry method was employed to detect apoptotic cells with membrane alterations (phosphatidylserine translocation), and differentiation of apoptotic from necrotic cells was also performed. Simultaneous staining of cell surface phosphatidylserine with Annexin-V-Fluorescein and necrotic cells with propidium iodide was conducted. 14 at 16 μg/mL caused C. albicans cells to undergo cellular apoptosis but its increasing concentrations induced necrosis. 14 influenced C. albicans biofilm viability as well as hyphal and cell wall morphology. Confocal microscopy and cell wall staining with calcofluor white revealed that in epithelial model the cell surface structure was perturbed at MIC of peptide dendrimer. It appears that tryptophan or 1-methyltryptophan groups displayed at the surface and positive charges hidden in the dendrimer tree along with hydrocarbon tail located at C-terminus are important for the anti-Candida activity since dendrimers containing tryptamine at C-terminus showed only a moderate activity. Our results suggest that membranolytic dendrimer 14, targeting cellular apoptotic pathway and impairing the cell wall formation in mature biofilm, may be a potential multifunctional antifungal lead compound for the control of C. albicans infections. Copyright © 2015 Elsevier Masson SAS. All rights reserved.

  20. Photoreactivation of conversion and de novo suppressor mutation in Escherichia coli

    Energy Technology Data Exchange (ETDEWEB)

    Bockrath, R C; Plamer, J E [Indiana Univ., Indianapolis (USA). Dept. of Microbiology

    1977-04-01

    Studies of mutagenesis and photoreactivation in various E.coli strains have shown that conversion mutation of a mutant containing an amber suppressor to one containing an ochre suppressor is sensitive to photoreactivation. Direct photoreactivation by photoreactivating light (PRL) after uv mutagenesis reduced mutation frequencies by a factor of about 2 for each minute of exposure during the first 5 to 8 min of exposure for cells with normal repair capacity. Conversion and potential de novo suppressor mutations were about equally sensitive. For conversion, the sensitivities to PRL were identical in the repair-normal and excisions-repair-deficient strains. For de novo suppressor mutation, the rate of mutation frequency reduction by PRL in the repair-deficient strain was about one-half that in the other strains. The results suggest that ultraviolet radiation produces both de novo suppressor mutation and conversion at the sup(E,B) locus by photoreversible pyrimidine dimers in the DNA. The causative dimers could be Thy()Cyt dimers in the transcribed strand or the non-transcribed strand, respectively.

  1. Self-assembly of cationic multidomain peptide hydrogels: supramolecular nanostructure and rheological properties dictate antimicrobial activity

    Science.gov (United States)

    Jiang, Linhai; Xu, Dawei; Sellati, Timothy J.; Dong, He

    2015-11-01

    Hydrogels are an important class of biomaterials that have been widely utilized for a variety of biomedical/medical applications. The biological performance of hydrogels, particularly those used as wound dressing could be greatly advanced if imbued with inherent antimicrobial activity capable of staving off colonization of the wound site by opportunistic bacterial pathogens. Possessing such antimicrobial properties would also protect the hydrogel itself from being adversely affected by microbial attachment to its surface. We have previously demonstrated the broad-spectrum antimicrobial activity of supramolecular assemblies of cationic multi-domain peptides (MDPs) in solution. Here, we extend the 1-D soluble supramolecular assembly to 3-D hydrogels to investigate the effect of the supramolecular nanostructure and its rheological properties on the antimicrobial activity of self-assembled hydrogels. Among designed MDPs, the bactericidal activity of peptide hydrogels was found to follow an opposite trend to that in solution. Improved antimicrobial activity of self-assembled peptide hydrogels is dictated by the combined effect of supramolecular surface chemistry and storage modulus of the bulk materials, rather than the ability of individual peptides/peptide assemblies to penetrate bacterial cell membrane as observed in solution. The structure-property-activity relationship developed through this study will provide important guidelines for designing biocompatible peptide hydrogels with built-in antimicrobial activity for various biomedical applications.Hydrogels are an important class of biomaterials that have been widely utilized for a variety of biomedical/medical applications. The biological performance of hydrogels, particularly those used as wound dressing could be greatly advanced if imbued with inherent antimicrobial activity capable of staving off colonization of the wound site by opportunistic bacterial pathogens. Possessing such antimicrobial properties would

  2. The Key Drivers behind Novo Nordisk’s Growth in the Diabetes Market in China

    Directory of Open Access Journals (Sweden)

    Hind Louiza CHITOUR

    2013-12-01

    Full Text Available To enter the Chinese Pharmaceutical market, “Big Pharma” has adopted different strategies to tackle the challenges specific to the country in terms of size, demographics, specific sales channels and logistics adjustments. While the majority of Global Pharmaceutical players have opted for an aggressive M&A approach to penetrate the Chinese market and gain local insight; the Danish Novo Nordisk has instead chosen a strategy focusing on innovation and developing its R&D structure to capitalize on the local talent pool. To illustrate Novo Nordisk’s growth strategy in the Mainland, we analyzed its competitiveness in the diabetes market by demonstrating the key drivers behind this success. We applied a various set of tools for this research: Novo Nordisk, Dong Bao Pharmaceutical executives’ interviews and personal observations accounting for the primary data, we also reviewed secondary data to perform a PEST analysis in addition to Porter’s competitive advantage model in order to extract the reasons behind Novo Nordisk’s marching success in the Mainland.

  3. Origin of anti-tumor activity of the cysteine-containing GO peptides and further optimization of their cytotoxic properties

    Science.gov (United States)

    Tyuryaeva, Irina I.; Lyublinskaya, Olga G.; Podkorytov, Ivan S.; Skrynnikov, Nikolai R.

    2017-01-01

    Antitumor GO peptides have been designed as dimerization inhibitors of prominent oncoprotein mucin 1. In this study we demonstrate that activity of GO peptides is independent of the level of cellular expression of mucin 1. Furthermore, these peptides prove to be broadly cytotoxic, causing cell death also in normal cells such as dermal fibroblasts and endometrial mesenchymal stem cells. To explore molecular mechanism of their cytotoxicity, we have designed and tested a number of new peptide sequences containing the key CxC or CxxC motifs. Of note, these sequences bear no similarity to mucin 1 except that they also contain a pair of proximal cysteines. Several of the new peptides turned out to be significantly more potent than their GO prototypes. The results suggest that cytotoxicity of these peptides stems from their (moderate) activity as disulfide oxidoreductases. It is expected that such peptides, which we have termed DO peptides, are involved in disulfide-dithiol exchange reaction, resulting in formation of adventitious disulfide bridges in cell proteins. In turn, this leads to a partial loss of protein function and rapid onset of apoptosis. We anticipate that coupling DO sequences with tumor-homing transduction domains can create a potentially valuable new class of tumoricidal peptides.

  4. The self-assembly of redox active peptides: Synthesis and electrochemical capacitive behavior.

    Science.gov (United States)

    Piccoli, Julia P; Santos, Adriano; Santos-Filho, Norival A; Lorenzón, Esteban N; Cilli, Eduardo M; Bueno, Paulo R

    2016-05-01

    The present work reports on the synthesis of a redox-tagged peptide with self-assembling capability aiming applications in electrochemically active capacitive surfaces (associated with the presence of the redox centers) generally useful in electroanalytical applications. Peptide containing ferrocene (fc) molecular (redox) group (Ac-Cys-Ile-Ile-Lys(fc)-Ile-Ile-COOH) was thus synthesized by solid phase peptide synthesis (SPPS). To obtain the electrochemically active capacitive interface, the side chain of the cysteine was covalently bound to the gold electrode (sulfur group) and the side chain of Lys was used to attach the ferrocene in the peptide chain. After obtaining the purified redox-tagged peptide, the self-assembly and redox capability was characterized by cyclic voltammetry (CV) and electrochemical impedance-based capacitance spectroscopy techniques. The obtained results confirmed that the redox-tagged peptide was successfully attached by forming an electroactive self-assembled monolayer onto gold electrode. The design of redox active self-assembly ferrocene-tagged peptide is predictably useful in the development of biosensor devices precisely to detect, in a label-free platform, those biomarkers of clinical relevance. © 2016 Wiley Periodicals, Inc. Biopolymers (Pept Sci) 106: 357-367, 2016. © 2016 Wiley Periodicals, Inc.

  5. Synergistic gene and drug tumor therapy using a chimeric peptide.

    Science.gov (United States)

    Han, Kai; Chen, Si; Chen, Wei-Hai; Lei, Qi; Liu, Yun; Zhuo, Ren-Xi; Zhang, Xian-Zheng

    2013-06-01

    Co-delivery of gene and drug for synergistic therapy has provided a promising strategy to cure devastating diseases. Here, an amphiphilic chimeric peptide (Fmoc)2KH7-TAT with pH-responsibility for gene and drug delivery was designed and fabricated. As a drug carrier, the micelles self-assembled from the peptide exhibited a much faster doxorubicin (DOX) release rate at pH 5.0 than that at pH 7.4. As a non-viral gene vector, (Fmoc)(2)KH(7)-TAT peptide could satisfactorily mediate transfection of pGL-3 reporter plasmid with or without the existence of serum in both 293T and HeLa cell-lines. Besides, the endosome escape capability of peptide/DNA complexes was investigated by confocal laser scanning microscopy (CLSM). To evaluate the co-delivery efficiency and the synergistic anti-tumor effect of gene and drug, p53 plasmid and DOX were simultaneously loaded in the peptide micelles to form micelleplexes during the self-assembly of the peptide. Cellular uptake and intracellular delivery of gene and drug were studied by CLSM and flow cytometry respectively. And p53 protein expression was determined via Western blot analysis. The in vitro cytotoxicity and in vivo tumor inhibition effect were also studied. Results suggest that the co-delivery of gene and drug from peptide micelles resulted in effective cell growth inhibition in vitro and significant tumor growth restraining in vivo. The chimeric peptide-based gene and drug co-delivery system will find great potential for tumor therapy. Copyright © 2013 Elsevier Ltd. All rights reserved.

  6. Bovine and human lactoferricin peptides: chimeras and new cyclic analogs.

    Science.gov (United States)

    Arias, Mauricio; McDonald, Lindsey J; Haney, Evan F; Nazmi, Kamran; Bolscher, Jan G M; Vogel, Hans J

    2014-10-01

    Lactoferrin (LF) is an important antimicrobial and immune regulatory protein present in neutrophils and most exocrine secretions of mammals. The antimicrobial activity of LF has been related to the presence of an antimicrobial peptide sequence, called lactoferricin (LFcin), located in the N-terminal region of the protein. The antimicrobial activity of bovine LFcin is considerably stronger than the human version. In this work, chimera peptides combining segments of bovine and human LFcin were generated in order to study their antimicrobial activity and mechanism of action. In addition, the relevance of the conserved disulfide bridge and the resulting cyclic structure of both LFcins were analyzed by using "click chemistry" and sortase A-catalyzed cyclization of the peptides. The N-terminal region of bovine LFcin (residues 17-25 of bovine LF) proved to be very important for the antimicrobial activity of the chimera peptides against E. coli, when combined with the C-terminal region of human LFcin. Similarly the cyclic bovine LFcin analogs generated by "click chemistry" and sortase A preserved the antimicrobial activity of the original peptide, showing the significance of these two techniques in the design of cyclic antimicrobial peptides. The mechanism of action of bovine LFcin and its active derived peptides was strongly correlated with membrane leakage in E. coli and up to some extent with the ability to induce vesicle aggregation. This mechanism was also preserved under conditions of high ionic strength (150 mM NaCl) illustrating the importance of these peptides in a more physiologically relevant system.

  7. Investigation of the antibiofilm capacity of peptide-modified stainless steel.

    Science.gov (United States)

    Cao, Pan; Li, Wen-Wu; Morris, Andrew R; Horrocks, Paul D; Yuan, Cheng-Qing; Yang, Ying

    2018-03-01

    Biofilm formation on surfaces is an important research topic in ship tribology and medical implants. In this study, dopamine and two types of synthetic peptides were designed and attached to 304 stainless steel surfaces, aiming to inhibit the formation of biofilms. A combinatory surface modification procedure was applied in which dopamine was used as a coupling agent, allowing a strong binding ability with the two peptides. X-ray photoelectron spectroscopy (XPS), elemental analysis, contact angle measurement and surface roughness test were used to evaluate the efficiency of the peptide modification. An antibiofilm assay against Staphylococcus aureus was conducted to validate the antibiofilm capacity of the peptide-modified stainless steel samples. XPS analysis confirmed that the optimal dopamine concentration was 40 µg ml -1 in the coupling reaction. Element analysis showed that dopamine and the peptides had bound to the steel surfaces. The robustness assay of the modified surface demonstrated that most peptide molecules had bound on the surface of the stainless steel firmly. The contact angle of the modified surfaces was significantly changed. Modified steel samples exhibited improved antibiofilm properties in comparison to untreated and dopamine-only counterpart, with the peptide 1 modification displaying the best antibiofilm effect. The modified surfaces showed antibacterial capacity. The antibiofilm capacity of the modified surfaces was also surface topography sensitive. The steel sample surfaces polished with 600# sandpaper exhibited stronger antibiofilm capacity than those polished with other types of sandpapers after peptide modification. These findings present valuable information for future antifouling material research.

  8. Identifying wrong assemblies in de novo short read primary ...

    Indian Academy of Sciences (India)

    2016-08-05

    Aug 5, 2016 ... Most of these assemblies are done using some de novo short read assemblers and other related approaches. .... benchmarking projects like Assemblathon 1, Assemblathon ... from a large insert library (at least 1000 bases).

  9. Insulin C-peptide test

    Science.gov (United States)

    C-peptide ... the test depends on the reason for the C-peptide measurement. Ask your health care provider if ... C-peptide is measured to tell the difference between insulin the body produces and insulin someone injects ...

  10. Designer bFGF-incorporated D-form self-assembly peptide nanofiber scaffolds to promote bone repair

    Energy Technology Data Exchange (ETDEWEB)

    He, Bin, E-mail: binheing@163.com [Department of Orthopedics, The First Affiliated Hospital of Chongqing Medical University, Chongqing 400016 (China); Ou, Yunsheng; Chen, Shuo; Zhao, Weikang; Zhou, Ao [Department of Orthopedics, The First Affiliated Hospital of Chongqing Medical University, Chongqing 400016 (China); Zhao, Jinqiu [Department of Infectious Diseases, The First Affiliated Hospital of Chongqing Medical University, Chongqing 400016 (China); Li, Hong [School of Physical Science and Technology, Sichuan University, Chengdu 610000 (China); Jiang, Dianming [Department of Orthopedics, The First Affiliated Hospital of Chongqing Medical University, Chongqing 400016 (China); Zhu, Yong, E-mail: 568731668@qq.com [Department of Orthopedics, The First Affiliated Hospital of Chongqing Medical University, Chongqing 400016 (China)

    2017-05-01

    D-Form and L-form peptide nanofiber scaffolds can spontaneously form stable β-sheet secondary structures and nanofiber hydrogel scaffolds, and hold some promise in hemostasis and wound healing. We report here on the synthetic self-assembling peptide D-RADA16 and L-RADA16 are both found to produce stable β-sheet secondary structure and nanofiber hydrogel scaffolds based on circular dichroism (CD) spectroscopy, transmission electron microscopy (TEM) and rheology analysis etc. D-RADA16 hydrogel and L-RADA16 hydrogel can enhance obvious bone repair in femoral condyle defects of the Sprague-Dawley (SD) rat model compared to PBS treatment. Based on micro-computed tomography (CT), it was revealed that D-RADA16 hydrogel and L-RADA16 hydrogel were capable to obtain the extensive bone healing. Histological evaluation also found that these two hydrogels facilitate the presence of more mature bone tissue within the femoral condyle defects. Additionally, D-RADA16 hydrogel showed some potential in storing and releasing basic-fibroblast growth factor (bFGF) which was able to further promote bone regeneration based on micro-CT analysis. These results indicate that D-form peptide nanofiber hydrogel have some special capacity for bone repair. - Highlights: • Peptide D-RADA16 and L-RADA16 can form stable hydrogels. • D-RADA16 hydrogel can obtain the comparable and extensive promotion to bone healing compared to L-RADA16 hydrogel. • L-RADA16 hydrogel allows for slow release of bFGF.

  11. Analysis of 60 706 Exomes Questions the Role of De Novo Variants Previously Implicated in Cardiac Disease

    DEFF Research Database (Denmark)

    Paludan-Müller, Christian; Ahlberg, Gustav; Ghouse, Jonas

    2017-01-01

    BACKGROUND: De novo variants in the exome occur at a rate of 1 per individual per generation, and because of the low reproductive fitness for de novo variants causing severe disease, the likelihood of finding these as standing variations in the general population is low. Therefore, this study...... sought to evaluate the pathogenicity of de novo variants previously associated with cardiac disease based on a large population-representative exome database. METHODS AND RESULTS: We performed a literature search for previous publications on de novo variants associated with severe arrhythmias...... trio studies (>1000 subjects). Of the monogenic variants, 11% (23/211) were present in ExAC, whereas 26% (802/3050) variants believed to increase susceptibility of disease were identified in ExAC. Monogenic de novo variants in ExAC had a total allele count of 109 and with ≈844 expected cases in Ex...

  12. CycloPs: generating virtual libraries of cyclized and constrained peptides including nonnatural amino acids.

    Science.gov (United States)

    Duffy, Fergal J; Verniere, Mélanie; Devocelle, Marc; Bernard, Elise; Shields, Denis C; Chubb, Anthony J

    2011-04-25

    We introduce CycloPs, software for the generation of virtual libraries of constrained peptides including natural and nonnatural commercially available amino acids. The software is written in the cross-platform Python programming language, and features include generating virtual libraries in one-dimensional SMILES and three-dimensional SDF formats, suitable for virtual screening. The stand-alone software is capable of filtering the virtual libraries using empirical measurements, including peptide synthesizability by standard peptide synthesis techniques, stability, and the druglike properties of the peptide. The software and accompanying Web interface is designed to enable the rapid generation of large, structurally diverse, synthesizable virtual libraries of constrained peptides quickly and conveniently, for use in virtual screening experiments. The stand-alone software, and the Web interface for evaluating these empirical properties of a single peptide, are available at http://bioware.ucd.ie .

  13. Double-Stranded Peptide Nucleic Acids

    DEFF Research Database (Denmark)

    2001-01-01

    A novel class of compounds, known as peptide nucleic acids, form double-stranded structures with one another and with ssDNA. The peptide nucleic acids generally comprise ligands such as naturally occurring DNA bases attached to a peptide backbone through a suitable linker.......A novel class of compounds, known as peptide nucleic acids, form double-stranded structures with one another and with ssDNA. The peptide nucleic acids generally comprise ligands such as naturally occurring DNA bases attached to a peptide backbone through a suitable linker....

  14. De novo structural modeling and computational sequence analysis ...

    African Journals Online (AJOL)

    Different bioinformatics tools and machine learning techniques were used for protein structural classification. De novo protein modeling was performed by using I-TASSER server. The final model obtained was accessed by PROCHECK and DFIRE2, which confirmed that the final model is reliable. Until complete biochemical ...

  15. Micropattern printing of adhesion, spreading, and migration peptides on poly(tetrafluoroethylene) films to promote endothelialization.

    Science.gov (United States)

    Gauvreau, Virginie; Laroche, Gaétan

    2005-01-01

    We report here the development of an original multistep micropatterning technique for printing peptides on surfaces, based on the ink-jet printer technology. Contrary to most micropatterning methods used nowadays, this technique is advantageous because it allows displaying 2D-arrays of multiple biomolecules. Moreover, this low cost procedure allies the advantages of computer-aided design with high flexibility and reproducibility. A Hewlett-Packard printer was modified to print peptide solutions, and Adobe Illustrator was used as the graphic-editing software to design high-resolution checkerboard-like micropatterns. In a first step, PTFE films were treated with ammonia plasma to introduce amino groups on the surface. These chemical functionalities were reacted with heterobifunctional cross-linker sulfo-succinimidyl 4-(N-maleimidomethyl)cycloexane-1-carboxylate (S-SMCC) to allow the subsequent surface covalent conjugation of various cysteine-modified peptides to the polymer substrate. These peptidic molecules containing RGD and WQPPRARI sequences were selected for their adhesive, spreading, and migrational properties toward endothelial cells. On one hand, our data demonstrated that the initial cell adhesion does not depend on the chemical structure and combination of the peptides covalently bonded either through conventional conjugation or micropatterning. On the other hand, spreading and migration of endothelial cells is clearly enhanced while coconjugating the GRGDS peptide in conjunction with WQPPRARI. This behavior is further improved by micropatterning these peptides on specific areas of the polymer surface.

  16. Similar prognosis of transformed and de novo diffuse large B-cell lymphomas in patients treated with immunochemotherapy.

    Science.gov (United States)

    Sorigue, Marc; Garcia, Olga; Baptista, Maria Joao; Sancho, Juan-Manuel; Tapia, Gustavo; Mate, José Luis; Feliu, Evarist; Navarro, José-Tomás; Ribera, Josep-Maria

    2017-03-22

    The prognosis of diffuse large B-cell lymphomas (DLBCL) transformed from indolent lymphoma (TL) has been considered poorer than that of de novo DLBCL. However, it seems to have improved since the introduction of rituximab. We compared the characteristics (including the cell-of-origin), and the prognosis of 29 patients with TL and 101 with de novo DLBCL treated with immunochemotherapy. Patients with TL and de novo DLBCL had similar characteristics. All TL cases evolving from follicular lymphoma were germinal-center B-cell-like, while those TL from marginal zone lymphoma or chronic lymphocytic leukemia were non-germinal-center B-cell-like. The complete response rate was similar in TL and de novo DLBCL (62 vs. 66%, P=.825). The 5-year overall and progression-free survival probabilities (95% CI) were 59% (40-78) and 41% (22-60) for TL and 63% (53-73) and 60% (50-70) for de novo DLBCL, respectively (P=.732 for overall survival and P=.169 for progression-free survival). In this study, the prognosis of TL and de novo DLBCL treated with immunochemotherapy was similar. The role of intensification with stem cell transplantation in the management of TL may be questionable in the rituximab era. Copyright © 2016 Elsevier España, S.L.U. All rights reserved.

  17. Preliminary screening and identification of the hepatocarcinoma cell-binding peptide

    International Nuclear Information System (INIS)

    Zhu Xiaohua; Wu Hua

    2004-01-01

    Objective: To explore the feasibility of screening and isolating homing peptides that bind specifically, or preferentially, to hepatocarcinoma cells using phage display random peptide library and to develop a new peptide which may be potentially used as targeting delivery carrier in the biological targeted diagnosis or therapy for liver cancer. Methods: A 12-mer peptide phage display library was used to screen and isolate peptides that bind to human hepatocarcinoma cells, and four rounds of subtractive panning were carried out with the human hepatocarcinoma cell line HepG2 as the target. The affinities of selected phage clones for human hepatocarcinoma cells were determined with enzyme-linked immunosorbent assay (ELISA) and compared with that to human liver cell and other tumor cells of different tissue origins, respectively. In addition, the binding site in the tumor cells was observed with immunofluorescence analysis under confocal light microscopy. The amino acid sequences of phages that bind HepG2 specifically were deduced through DNA sequencing. Based on the results of DNA sequence, a 16-mer peptide (WH16) was designed and synthesized. Binding ability of the new peptide, WH16, was determined with competitive inhibition test. Results: After four rounds of panning, the phages that were bound to and internalized in human hepatocarcinoma cells were isolated. ELISA and immunofluorescence analysis confirmed the affinity of these phages for hepatocarcinoma cells. 56.67%(17/30) of the isolated phages displayed repeated sequence FLLEPHLMDTSM, and FLEP was defined as conservative motif . Binding of the selected phage to HepG2 cells was inhibited by synthesized peptide WH16, that strongly support that cellular binding of the phage is mediated through its displayed peptide, and WH16 can also bind to HepG2. Conclusions: It is feasible to screen and isolate homing peptides that bind specifically, or preferentially, to hepatocarcinoma cells using phage display random peptide

  18. Preliminary screening and identification of the hepatocarcinoma cell-binding peptide

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    Xiaohua, Zhu; Hua, Wu [Department of Nuclear Medicine, Tongji Hospital, Tongji Medical College, Huazhong Univ. of Science and Technology, Wuhan (China)

    2004-12-15

    Objective: To explore the feasibility of screening and isolating homing peptides that bind specifically, or preferentially, to hepatocarcinoma cells using phage display random peptide library and to develop a new peptide which may be potentially used as targeting delivery carrier in the biological targeted diagnosis or therapy for liver cancer. Methods: A 12-mer peptide phage display library was used to screen and isolate peptides that bind to human hepatocarcinoma cells, and four rounds of subtractive panning were carried out with the human hepatocarcinoma cell line HepG2 as the target. The affinities of selected phage clones for human hepatocarcinoma cells were determined with enzyme-linked immunosorbent assay (ELISA) and compared with that to human liver cell and other tumor cells of different tissue origins, respectively. In addition, the binding site in the tumor cells was observed with immunofluorescence analysis under confocal light microscopy. The amino acid sequences of phages that bind HepG2 specifically were deduced through DNA sequencing. Based on the results of DNA sequence, a 16-mer peptide (WH16) was designed and synthesized. Binding ability of the new peptide, WH16, was determined with competitive inhibition test. Results: After four rounds of panning, the phages that were bound to and internalized in human hepatocarcinoma cells were isolated. ELISA and immunofluorescence analysis confirmed the affinity of these phages for hepatocarcinoma cells. 56.67%(17/30) of the isolated phages displayed repeated sequence FLLEPHLMDTSM, and FLEP was defined as conservative motif . Binding of the selected phage to HepG2 cells was inhibited by synthesized peptide WH16, that strongly support that cellular binding of the phage is mediated through its displayed peptide, and WH16 can also bind to HepG2. Conclusions: It is feasible to screen and isolate homing peptides that bind specifically, or preferentially, to hepatocarcinoma cells using phage display random peptide

  19. C-terminal substitution of MDM2 interacting peptides modulates binding affinity by distinctive mechanisms.

    Directory of Open Access Journals (Sweden)

    Christopher J Brown

    Full Text Available The complex between the proteins MDM2 and p53 is a promising drug target for cancer therapy. The residues 19-26 of p53 have been biochemically and structurally demonstrated to be a most critical region to maintain the association of MDM2 and p53. Variation of the amino acid sequence in this range obviously alters the binding affinity. Surprisingly, suitable substitutions contiguous to this region of the p53 peptides can yield tightly binding peptides. The peptide variants may differ by a single residue that vary little in their structural conformations and yet are characterized by large differences in their binding affinities. In this study a systematic analysis into the role of single C-terminal mutations of a 12 residue fragment of the p53 transactivation domain (TD and an equivalent phage optimized peptide (12/1 were undertaken to elucidate their mechanistic and thermodynamic differences in interacting with the N-terminal of MDM2. The experimental results together with atomistically detailed dynamics simulations provide insight into the principles that govern peptide design protocols with regard to protein-protein interactions and peptidomimetic design.

  20. Application of synthetic peptides for detection of anti-citrullinated peptide antibodies

    DEFF Research Database (Denmark)

    Trier, Nicole Hartwig; Holm, Bettina Eide; Slot, Ole

    2016-01-01

    Anti-citrullinated protein antibodies (ACPAs) are a hallmark of rheumatoid arthritis (RA) and represent an important tool for the serological diagnosis of RA. In this study, we describe ACPA reactivity to overlapping citrullinated Epstein-Barr virus nuclear antigen-1 (EBNA-1)-derived peptides...... (n=40), systemic lupus erythematosus (n=20), Sjögren's syndrome (n=40)) were screened for antibody reactivity. Antibodies to a panel of five citrullinated EBNA-1 peptides were found in 67% of RA sera, exclusively of the IgG isotype, while 53% of the patient sera reacted with a single peptide......, ARGGSRERARGRGRG-Cit-GEKR, accounting for more than half of the ACPA reactivity alone. Moreover, these antibodies were detected in 10% of CCP2-negative RA sera. In addition, 47% of the RA sera reacted with two or three citrullinated EBNA-1 peptides from the selected peptide panel. Furthermore, a negative...