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Sample records for short peptide sequence

  1. Effect of stereochemistry, chain length and sequence pattern on antimicrobial properties of short synthetic β-sheet forming peptide amphiphiles.

    Science.gov (United States)

    Ong, Zhan Yuin; Cheng, Junchi; Huang, Yuan; Xu, Kaijin; Ji, Zhongkang; Fan, Weimin; Yang, Yi Yan

    2014-01-01

    In the face of mounting global antibiotics resistance, the identification and development of membrane-active antimicrobial peptides (AMPs) as an alternative class of antimicrobial agent have gained significant attention. The physical perturbation and disruption of microbial membranes by the AMPs have been proposed to be an effective means to overcome conventional mechanisms of drug resistance. Recently, we have reported the design of a series of short synthetic β-sheet folding peptide amphiphiles comprised of recurring (X1Y1X2Y2)n-NH2 sequences where X: hydrophobic amino acids, Y: cationic amino acids and n: number of repeat units. In efforts to investigate the effects of key parameters including stereochemistry, chain length and sequence pattern on antimicrobial effects, systematic d-amino acid substitutions of the lead peptides (IRIK)2-NH2 (IK8-all L) and (IRVK)3-NH2 (IK12-all L) were performed. It was found that the corresponding D-enantiomers exhibited stronger antimicrobial activities with minimal or no change in hemolytic activities, hence translating very high selectivity indices of 407.0 and >9.8 for IK8-all D and IK12-all D respectively. IK8-all D was also demonstrated to be stable to degradation by broad spectrum proteases trypsin and proteinase K. The membrane disrupting bactericidal properties of IK8-all D effectively prevented drug resistance development and inhibited the growth of various clinically isolated MRSA, VRE, Acinetobacter baumanni, Pseudomonas aeruginosa, Cryptococcus. neoformans and Mycobacterium tuberculosis. Significant reduction in intracellular bacteria counts was also observed following treatment with IK8-all D in the Staphylococcus. aureus infected mouse macrophage cell line RAW264.7 (P < 0.01). These results suggest that the d-amino acids substituted β-sheet forming peptide IK8-all D with its enhanced antimicrobial activities and improved protease stability, is a promising therapeutic candidate with potential to combat

  2. Short peptides allowing preferential detection of Candida albicans hyphae.

    Science.gov (United States)

    Kaba, Hani E J; Pölderl, Antonia; Bilitewski, Ursula

    2015-09-01

    Whereas the detection of pathogens via recognition of surface structures by specific antibodies and various types of antibody mimics is frequently described, the applicability of short linear peptides as sensor molecules or diagnostic tools is less well-known. We selected peptides which were previously reported to bind to recombinant S. cerevisiae cells, expressing members of the C. albicans Agglutinin-Like-Sequence (ALS) cell wall protein family. We slightly modified amino acid sequences to evaluate peptide sequence properties influencing binding to C. albicans cells. Among the selected peptides, decamer peptides with an "AP"-N-terminus were superior to shorter peptides. The new decamer peptide FBP4 stained viable C. albicans cells more efficiently in their mature hyphal form than in their yeast form. Moreover, it allowed distinction of C. albicans from other related Candida spp. and could thus be the basis for the development of a useful tool for the diagnosis of invasive candidiasis.

  3. Sequencing Cyclic Peptides by Multistage Mass Spectrometry

    Science.gov (United States)

    Mohimani, Hosein; Yang, Yu-Liang; Liu, Wei-Ting; Hsieh, Pei-Wen; Dorrestein, Pieter C.; Pevzner, Pavel A.

    2012-01-01

    Some of the most effective antibiotics (e.g., Vancomycin and Daptomycin) are cyclic peptides produced by non-ribosomal biosynthetic pathways. While hundreds of biomedically important cyclic peptides have been sequenced, the computational techniques for sequencing cyclic peptides are still in their infancy. Previous methods for sequencing peptide antibiotics and other cyclic peptides are based on Nuclear Magnetic Resonance spectroscopy, and require large amount (miligrams) of purified materials that, for most compounds, are not possible to obtain. Recently, development of mass spectrometry based methods has provided some hope for accurate sequencing of cyclic peptides using picograms of materials. In this paper we develop a method for sequencing of cyclic peptides by multistage mass spectrometry, and show its advantages over single stage mass spectrometry. The method is tested on known and new cyclic peptides from Bacillus brevis, Dianthus superbus and Streptomyces griseus, as well as a new family of cyclic peptides produced by marine bacteria. PMID:21751357

  4. Antimalarial Activity of Ultra-Short Peptides

    Directory of Open Access Journals (Sweden)

    María Yolanda Rios

    2009-12-01

    Full Text Available Ultra-short peptides 1-9 were designed and synthesized with phenylalanine, ornithine and proline amino acid residues and their effect on antimalarial activity was analyzed. On the basis of the IC50 data for these compounds, the effects of nature, polarity, and amino acid sequence on Plasmodium berghei schizont cultures were analyzed too. Tetrapeptides Phe-Orn-Phe-Orn (4 and Lys-Phe-Phe-Orn (5 showed a very important activity with IC50 values of 3.31 and 2.57 μM, respectively. These two tetrapeptides are candidates for subsequent in vivo assays and SARS investigations.

  5. Sequence dependent aggregation of peptides and fibril formation

    Science.gov (United States)

    Hung, Nguyen Ba; Le, Duy-Manh; Hoang, Trinh X.

    2017-09-01

    Deciphering the links between amino acid sequence and amyloid fibril formation is key for understanding protein misfolding diseases. Here we use Monte Carlo simulations to study the aggregation of short peptides in a coarse-grained model with hydrophobic-polar (HP) amino acid sequences and correlated side chain orientations for hydrophobic contacts. A significant heterogeneity is observed in the aggregate structures and in the thermodynamics of aggregation for systems of different HP sequences and different numbers of peptides. Fibril-like ordered aggregates are found for several sequences that contain the common HPH pattern, while other sequences may form helix bundles or disordered aggregates. A wide variation of the aggregation transition temperatures among sequences, even among those of the same hydrophobic fraction, indicates that not all sequences undergo aggregation at a presumable physiological temperature. The transition is found to be the most cooperative for sequences forming fibril-like structures. For a fibril-prone sequence, it is shown that fibril formation follows the nucleation and growth mechanism. Interestingly, a binary mixture of peptides of an aggregation-prone and a non-aggregation-prone sequence shows the association and conversion of the latter to the fibrillar structure. Our study highlights the role of a sequence in selecting fibril-like aggregates and also the impact of a structural template on fibril formation by peptides of unrelated sequences.

  6. Polymeric peptide pigments with sequence-encoded properties

    Energy Technology Data Exchange (ETDEWEB)

    Lampel, Ayala; McPhee, Scott A.; Park, Hang-Ah; Scott, Gary G.; Humagain, Sunita; Hekstra, Doeke R.; Yoo, Barney; Frederix, Pim W. J. M.; Li, Tai-De; Abzalimov, Rinat R.; Greenbaum, Steven G.; Tuttle, Tell; Hu, Chunhua; Bettinger, Christopher J.; Ulijn, Rein V.

    2017-06-08

    Melanins are a family of heterogeneous polymeric pigments that provide ultraviolet (UV) light protection, structural support, coloration, and free radical scavenging. Formed by oxidative oligomerization of catecholic small molecules, the physical properties of melanins are influenced by covalent and noncovalent disorder. We report the use of tyrosine-containing tripeptides as tunable precursors for polymeric pigments. In these structures, phenols are presented in a (supra-)molecular context dictated by the positions of the amino acids in the peptide sequence. Oxidative polymerization can be tuned in a sequence-dependent manner, resulting in peptide sequence–encoded properties such as UV absorbance, morphology, coloration, and electrochemical properties over a considerable range. Short peptides have low barriers to application and can be easily scaled, suggesting near-term applications in cosmetics and biomedicine.

  7. Short Anabolic Peptides for Bone Growth.

    Science.gov (United States)

    Amso, Zaid; Cornish, Jillian; Brimble, Margaret A

    2016-07-01

    Loss of bone occurs in the age-related skeletal disorder, osteoporosis, leading to bone fragility and increased incidence of fractures, which are associated with enormous costs and substantial morbidity and mortality. Recent data indicate that osteoporotic fractures are more common than other diseases, which usually attract public attention (e.g., heart attack and breast cancer). The prevention and treatment of this skeletal disorder are therefore of paramount importance. Majority of osteoporosis medications restore skeletal balance by reducing osteoclastic activity, thereby reducing bone resorption. These agents, however, do not regenerate damaged bone tissue, leaving limited options for patients once bone loss has occurred. Recently, attention has turned to bone-anabolic agents. Such agents have the ability to increase bone mass and strength, potentially reversing structural damage. To date, only one bone-anabolic drug is available in the market. The discovery of more novel, cost-effective bone anabolic agents is therefore a priority to treat those suffering from this disabling condition. Short peptides offer an important alternative for the development of novel bone-anabolic agents given their high target binding specificity, which translates into potent activity with limited side effects. This review summarizes attempts in the identification of bone-anabolic peptides, and their development for promoting bone growth. © 2016 Wiley Periodicals, Inc.

  8. Aggregation properties of a short peptide that mediates amyloid fibril ...

    Indian Academy of Sciences (India)

    Short peptides have been identified from amyloidogenic proteins that form amyloid fibrils in isolation. The ... proteins. These peptide fibrils have the conformational features of β-structure that .... water and immediately deposited on freshly cleaved surface of mica .... with the peptide via electrostatic interactions. NaCl would.

  9. 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

  10. Folding and activity of hybrid sequence, disulfide-stabilized peptides

    Energy Technology Data Exchange (ETDEWEB)

    Pease, J.H.B.; Storrs, R.W.; Wemmer, D.E. (Univ. of California, Berkeley (USA))

    1990-08-01

    Peptides have been synthesized that have hybrid sequences, partially derived from the bee venom peptide apamin and partially from the S peptide of ribonuclease A. The hybrid peptides were demonstrated by NMR spectroscopy to fold, forming the same disulfides and basic three-dimensional structure as native apamin, containing a {beta}-turn and an {alpha}-helix. These hybrids were active in complementing S protein, reactivating nuclease activity. In addition, the hybrid peptide was effective in inducing antibodies that cross-react with the RNase, without conjugation to a carrier protein. The stability of the folded structure of this peptide suggests that it should be possible to elicit antibodies that will react not only with a specific sequence, but also with a specific secondary structure. Hybrid sequence peptides also provide opportunities to study separately nucleation and propagation steps in formation of secondary structure. The authors show that in S peptide the {alpha}-helix does not end abruptly but rather terminates gradually over four or five residues. In general, these hybrid sequence peptides, which fold predictably because of disulfide bond formation, can provide opportunities for examining structure - function relationships for many biologically active sequences.

  11. Folding and activity of hybrid sequence, disulfide-stabilized peptides

    International Nuclear Information System (INIS)

    Pease, J.H.B.; Storrs, R.W.; Wemmer, D.E.

    1990-01-01

    Peptides have been synthesized that have hybrid sequences, partially derived from the bee venom peptide apamin and partially from the S peptide of ribonuclease A. The hybrid peptides were demonstrated by NMR spectroscopy to fold, forming the same disulfides and basic three-dimensional structure as native apamin, containing a β-turn and an α-helix. These hybrids were active in complementing S protein, reactivating nuclease activity. In addition, the hybrid peptide was effective in inducing antibodies that cross-react with the RNase, without conjugation to a carrier protein. The stability of the folded structure of this peptide suggests that it should be possible to elicit antibodies that will react not only with a specific sequence, but also with a specific secondary structure. Hybrid sequence peptides also provide opportunities to study separately nucleation and propagation steps in formation of secondary structure. The authors show that in S peptide the α-helix does not end abruptly but rather terminates gradually over four or five residues. In general, these hybrid sequence peptides, which fold predictably because of disulfide bond formation, can provide opportunities for examining structure - function relationships for many biologically active sequences

  12. 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...

  13. Short sequence motifs, overrepresented in mammalian conservednon-coding sequences

    Energy Technology Data Exchange (ETDEWEB)

    Minovitsky, Simon; Stegmaier, Philip; Kel, Alexander; Kondrashov,Alexey S.; Dubchak, Inna

    2007-02-21

    Background: A substantial fraction of non-coding DNAsequences of multicellular eukaryotes is under selective constraint. Inparticular, ~;5 percent of the human genome consists of conservednon-coding sequences (CNSs). CNSs differ from other genomic sequences intheir nucleotide composition and must play important functional roles,which mostly remain obscure.Results: We investigated relative abundancesof short sequence motifs in all human CNSs present in the human/mousewhole-genome alignments vs. three background sets of sequences: (i)weakly conserved or unconserved non-coding sequences (non-CNSs); (ii)near-promoter sequences (located between nucleotides -500 and -1500,relative to a start of transcription); and (iii) random sequences withthe same nucleotide composition as that of CNSs. When compared tonon-CNSs and near-promoter sequences, CNSs possess an excess of AT-richmotifs, often containing runs of identical nucleotides. In contrast, whencompared to random sequences, CNSs contain an excess of GC-rich motifswhich, however, lack CpG dinucleotides. Thus, abundance of short sequencemotifs in human CNSs, taken as a whole, is mostly determined by theiroverall compositional properties and not by overrepresentation of anyspecific short motifs. These properties are: (i) high AT-content of CNSs,(ii) a tendency, probably due to context-dependent mutation, of A's andT's to clump, (iii) presence of short GC-rich regions, and (iv) avoidanceof CpG contexts, due to their hypermutability. Only a small number ofshort motifs, overrepresented in all human CNSs are similar to bindingsites of transcription factors from the FOX family.Conclusion: Human CNSsas a whole appear to be too broad a class of sequences to possess strongfootprints of any short sequence-specific functions. Such footprintsshould be studied at the level of functional subclasses of CNSs, such asthose which flank genes with a particular pattern of expression. Overallproperties of CNSs are affected by

  14. Folding very short peptides using molecular dynamics.

    Directory of Open Access Journals (Sweden)

    Bosco K Ho

    2006-04-01

    Full Text Available Peptides often have conformational preferences. We simulated 133 peptide 8-mer fragments from six different proteins, sampled by replica-exchange molecular dynamics using Amber7 with a GB/SA (generalized-Born/solvent-accessible electrostatic approximation to water implicit solvent. We found that 85 of the peptides have no preferred structure, while 48 of them converge to a preferred structure. In 85% of the converged cases (41 peptides, the structures found by the simulations bear some resemblance to their native structures, based on a coarse-grained backbone description. In particular, all seven of the beta hairpins in the native structures contain a fragment in the turn that is highly structured. In the eight cases where the bioinformatics-based I-sites library picks out native-like structures, the present simulations are largely in agreement. Such physics-based modeling may be useful for identifying early nuclei in folding kinetics and for assisting in protein-structure prediction methods that utilize the assembly of peptide fragments.

  15. 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.

  16. Serum Stabilities of Short Tryptophan-and Arginine-Rich Antimicrobial Peptide Analogs

    NARCIS (Netherlands)

    Nguyen, L.T.; Chau, J.K.; Perry, N.A.; de Boer, L.; Zaat, S.A.J.; Vogel, H.J.

    2010-01-01

    Background: 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

  17. 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.

  18. Targeted assembly of short sequence reads.

    Directory of Open Access Journals (Sweden)

    René L Warren

    Full Text Available As next-generation sequence (NGS production continues to increase, analysis is becoming a significant bottleneck. However, in situations where information is required only for specific sequence variants, it is not necessary to assemble or align whole genome data sets in their entirety. Rather, NGS data sets can be mined for the presence of sequence variants of interest by localized assembly, which is a faster, easier, and more accurate approach. We present TASR, a streamlined assembler that interrogates very large NGS data sets for the presence of specific variants by only considering reads within the sequence space of input target sequences provided by the user. The NGS data set is searched for reads with an exact match to all possible short words within the target sequence, and these reads are then assembled stringently to generate a consensus of the target and flanking sequence. Typically, variants of a particular locus are provided as different target sequences, and the presence of the variant in the data set being interrogated is revealed by a successful assembly outcome. However, TASR can also be used to find unknown sequences that flank a given target. We demonstrate that TASR has utility in finding or confirming genomic mutations, polymorphisms, fusions and integration events. Targeted assembly is a powerful method for interrogating large data sets for the presence of sequence variants of interest. TASR is a fast, flexible and easy to use tool for targeted assembly.

  19. BIOPEP database and other programs for processing bioactive peptide sequences.

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    Minkiewicz, Piotr; Dziuba, Jerzy; Iwaniak, Anna; Dziuba, Marta; Darewicz, Małgorzata

    2008-01-01

    This review presents the potential for application of computational tools in peptide science based on a sample BIOPEP database and program as well as other programs and databases available via the World Wide Web. The BIOPEP application contains a database of biologically active peptide sequences and a program enabling construction of profiles of the potential biological activity of protein fragments, calculation of quantitative descriptors as measures of the value of proteins as potential precursors of bioactive peptides, and prediction of bonds susceptible to hydrolysis by endopeptidases in a protein chain. Other bioactive and allergenic peptide sequence databases are also presented. Programs enabling the construction of binary and multiple alignments between peptide sequences, the construction of sequence motifs attributed to a given type of bioactivity, searching for potential precursors of bioactive peptides, and the prediction of sites susceptible to proteolytic cleavage in protein chains are available via the Internet as are other approaches concerning secondary structure prediction and calculation of physicochemical features based on amino acid sequence. Programs for prediction of allergenic and toxic properties have also been developed. This review explores the possibilities of cooperation between various programs.

  20. Salt-bridging effects on short amphiphilic helical structure and introducing sequence-based short beta-turn motifs.

    Science.gov (United States)

    Guarracino, Danielle A; Gentile, Kayla; Grossman, Alec; Li, Evan; Refai, Nader; Mohnot, Joy; King, Daniel

    2018-02-01

    Determining the minimal sequence necessary to induce protein folding is beneficial in understanding the role of protein-protein interactions in biological systems, as their three-dimensional structures often dictate their activity. Proteins are generally comprised of discrete secondary structures, from α-helices to β-turns and larger β-sheets, each of which is influenced by its primary structure. Manipulating the sequence of short, moderately helical peptides can help elucidate the influences on folding. We created two new scaffolds based on a modestly helical eight-residue peptide, PT3, we previously published. Using circular dichroism (CD) spectroscopy and changing the possible salt-bridging residues to new combinations of Lys, Arg, Glu, and Asp, we found that our most helical improvements came from the Arg-Glu combination, whereas the Lys-Asp was not significantly different from the Lys-Glu of the parent scaffold, PT3. The marked 3 10 -helical contributions in PT3 were lessened in the Arg-Glu-containing peptide with the beginning of cooperative unfolding seen through a thermal denaturation. However, a unique and unexpected signature was seen for the denaturation of the Lys-Asp peptide which could help elucidate the stages of folding between the 3 10 and α-helix. In addition, we developed a short six-residue peptide with β-turn/sheet CD signature, again to help study minimal sequences needed for folding. Overall, the results indicate that improvements made to short peptide scaffolds by fine-tuning the salt-bridging residues can enhance scaffold structure. Likewise, with the results from the new, short β-turn motif, these can help impact future peptidomimetic designs in creating biologically useful, short, structured β-sheet-forming peptides.

  1. Biodegradable copolymers carrying cell-adhesion peptide sequences.

    Science.gov (United States)

    Proks, Vladimír; Machová, Lud'ka; Popelka, Stepán; Rypácek, Frantisek

    2003-01-01

    Amphiphilic block copolymers are used to create bioactive surfaces on biodegradable polymer scaffolds for tissue engineering. Cell-selective biomaterials can be prepared using copolymers containing peptide sequences derived from extracellular-matrix proteins (ECM). Here we discuss alternative ways for preparation of amphiphilic block copolymers composed of hydrophobic polylactide (PLA) and hydrophilic poly(ethylene oxide) (PEO) blocks with cell-adhesion peptide sequences. Copolymers PLA-b-PEO were prepared by a living polymerisation of lactide in dioxane with tin(II)2-ethylhexanoate as a catalyst. The following approaches for incorporation of peptides into copolymers were elaborated. (a) First, a side-chain protected Gly-Arg-Gly-Asp-Ser-Gly (GRGDSG) peptide was prepared by solid-phase peptide synthesis (SPPS) and then coupled with delta-hydroxy-Z-amino-PEO in solution. In the second step, the PLA block was grafted to it via a controlled polymerisation of lactide initiated by the hydroxy end-groups of PEO in the side-chain-protected GRGDSG-PEO. Deprotection of the peptide yielded a GRGDSG-b-PEO-b-PLA copolymer, with the peptide attached through its C-end. (b) A protected GRGDSG peptide was built up on a polymer resin and coupled with Z-carboxy-PEO using a solid-phase approach. After cleavage of the delta-hydroxy-PEO-GRGDSG copolymer from the resin, polymerisation of lactide followed by deprotection of the peptide yielded a PLA-b-PEO-b-GRGDSG block copolymer, in which the peptide is linked through its N-terminus.

  2. Temperature dependence of looping rates in a short peptide.

    Science.gov (United States)

    Roccatano, Danilo; Sahoo, Harekrushna; Zacharias, Martin; Nau, Werner M

    2007-03-15

    Knowledge of the influence of chain length and amino acid sequence on the structural and dynamic properties of small peptides in solution provides essential information on protein folding pathways. The combination of time-resolved optical spectroscopy and molecular dynamics (MD) simulation methods has become a powerful tool to investigate the kinetics of end-to-end collisions (looping rates) in short peptides, which are relevant in early protein folding events. We applied the combination of both techniques to study temperature-dependent (280-340 K) looping rates of the Dbo-AlaGlyGln-Trp-NH2 peptide, where Dbo represents a 2,3-diazabicyclo[2.2.2]oct-2-ene-labeled asparagine, which served as a fluorescent probe in the time-resolved spectroscopic experiments. The experimental looping rates increased from 4.8 x 10(7) s(-1) at 283 K to 2.0 x 10(8) s(-1) at 338 K in H2O. The corresponding Arrhenius plot provided as activation parameters Ea = 21.5 +/- 1.0 kJ mol(-1) and ln(A/s-1) = 26.8 +/- 0.2 in H2O. The results in D2O were consistent with a slight solvent viscosity effect, i.e., the looping rates were 10-20% slower. MD simulations were performed with the GROMOS96 force field in a water solvent model, which required first a parametrization of the synthetic amino acid Dbo. After corrections for solvent viscosity effects, the calculated looping rates varied from 1.5 x 10(8) s(-1) at 280 K to 8.2 x 10(8) s(-1) at 340 K in H2O, which was about four times larger than the experimental data. The calculated activation parameters were Ea = 24.7 +/- 1.5 kJ mol(-1) and ln(A/s(-1)) = 29.4 +/- 0.1 in H2O.

  3. 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

  4. 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

  5. GuiTope: an application for mapping random-sequence peptides to protein sequences.

    Science.gov (United States)

    Halperin, Rebecca F; Stafford, Phillip; Emery, Jack S; Navalkar, Krupa Arun; Johnston, Stephen Albert

    2012-01-03

    Random-sequence peptide libraries are a commonly used tool to identify novel ligands for binding antibodies, other proteins, and small molecules. It is often of interest to compare the selected peptide sequences to the natural protein binding partners to infer the exact binding site or the importance of particular residues. The ability to search a set of sequences for similarity to a set of peptides may sometimes enable the prediction of an antibody epitope or a novel binding partner. We have developed a software application designed specifically for this task. GuiTope provides a graphical user interface for aligning peptide sequences to protein sequences. All alignment parameters are accessible to the user including the ability to specify the amino acid frequency in the peptide library; these frequencies often differ significantly from those assumed by popular alignment programs. It also includes a novel feature to align di-peptide inversions, which we have found improves the accuracy of antibody epitope prediction from peptide microarray data and shows utility in analyzing phage display datasets. Finally, GuiTope can randomly select peptides from a given library to estimate a null distribution of scores and calculate statistical significance. GuiTope provides a convenient method for comparing selected peptide sequences to protein sequences, including flexible alignment parameters, novel alignment features, ability to search a database, and statistical significance of results. The software is available as an executable (for PC) at http://www.immunosignature.com/software and ongoing updates and source code will be available at sourceforge.net.

  6. GuiTope: an application for mapping random-sequence peptides to protein sequences

    Directory of Open Access Journals (Sweden)

    Halperin Rebecca F

    2012-01-01

    Full Text Available Abstract Background Random-sequence peptide libraries are a commonly used tool to identify novel ligands for binding antibodies, other proteins, and small molecules. It is often of interest to compare the selected peptide sequences to the natural protein binding partners to infer the exact binding site or the importance of particular residues. The ability to search a set of sequences for similarity to a set of peptides may sometimes enable the prediction of an antibody epitope or a novel binding partner. We have developed a software application designed specifically for this task. Results GuiTope provides a graphical user interface for aligning peptide sequences to protein sequences. All alignment parameters are accessible to the user including the ability to specify the amino acid frequency in the peptide library; these frequencies often differ significantly from those assumed by popular alignment programs. It also includes a novel feature to align di-peptide inversions, which we have found improves the accuracy of antibody epitope prediction from peptide microarray data and shows utility in analyzing phage display datasets. Finally, GuiTope can randomly select peptides from a given library to estimate a null distribution of scores and calculate statistical significance. Conclusions GuiTope provides a convenient method for comparing selected peptide sequences to protein sequences, including flexible alignment parameters, novel alignment features, ability to search a database, and statistical significance of results. The software is available as an executable (for PC at http://www.immunosignature.com/software and ongoing updates and source code will be available at sourceforge.net.

  7. BLEACHING EUCALYPTUS PULPS WITH SHORT SEQUENCES

    Directory of Open Access Journals (Sweden)

    Flaviana Reis Milagres

    2011-03-01

    Full Text Available Eucalyptus spp kraft pulp, due to its high content of hexenuronic acids, is quite easy to bleach. Therefore, investigations have been made attempting to decrease the number of stages in the bleaching process in order to minimize capital costs. This study focused on the evaluation of short ECF (Elemental Chlorine Free and TCF (Totally Chlorine Free sequences for bleaching oxygen delignified Eucalyptus spp kraft pulp to 90% ISO brightness: PMoDP (Molybdenum catalyzed acid peroxide, chlorine dioxide and hydrogen peroxide, PMoD/P (Molybdenum catalyzed acid peroxide, chlorine dioxide and hydrogen peroxide, without washing PMoD(PO (Molybdenum catalyzed acid peroxide, chlorine dioxide and pressurized peroxide, D(EPODP (chlorine dioxide, extraction oxidative with oxygen and peroxide, chlorine dioxide and hydrogen peroxide, PMoQ(PO (Molybdenum catalyzed acid peroxide, DTPA and pressurized peroxide, and XPMoQ(PO (Enzyme, molybdenum catalyzed acid peroxide, DTPA and pressurized peroxide. Uncommon pulp treatments, such as molybdenum catalyzed acid peroxide (PMo and xylanase (X bleaching stages, were used. Among the ECF alternatives, the two-stage PMoD/P sequence proved highly cost-effective without affecting pulp quality in relation to the traditional D(EPODP sequence and produced better quality effluent in relation to the reference. However, a four stage sequence, XPMoQ(PO, was required to achieve full brightness using the TCF technology. This sequence was highly cost-effective although it only produced pulp of acceptable quality.

  8. A theoretical justification for single molecule peptide sequencing.

    Directory of Open Access Journals (Sweden)

    Jagannath Swaminathan

    2015-02-01

    Full Text Available The proteomes of cells, tissues, and organisms reflect active cellular processes and change continuously in response to intracellular and extracellular cues. Deep, quantitative profiling of the proteome, especially if combined with mRNA and metabolite measurements, should provide an unprecedented view of cell state, better revealing functions and interactions of cell components. Molecular diagnostics and biomarker discovery should benefit particularly from the accurate quantification of proteomes, since complex diseases like cancer change protein abundances and modifications. Currently, shotgun mass spectrometry is the primary technology for high-throughput protein identification and quantification; while powerful, it lacks high sensitivity and coverage. We draw parallels with next-generation DNA sequencing and propose a strategy, termed fluorosequencing, for sequencing peptides in a complex protein sample at the level of single molecules. In the proposed approach, millions of individual fluorescently labeled peptides are visualized in parallel, monitoring changing patterns of fluorescence intensity as N-terminal amino acids are sequentially removed, and using the resulting fluorescence signatures (fluorosequences to uniquely identify individual peptides. We introduce a theoretical foundation for fluorosequencing and, by using Monte Carlo computer simulations, we explore its feasibility, anticipate the most likely experimental errors, quantify their potential impact, and discuss the broad potential utility offered by a high-throughput peptide sequencing technology.

  9. Prediction of Scylla olivacea (Crustacea; Brachyura) peptide hormones using publicly accessible transcriptome shotgun assembly (TSA) sequences.

    Science.gov (United States)

    Christie, Andrew E

    2016-05-01

    The aquaculture of crabs from the genus Scylla is of increasing economic importance for many Southeast Asian countries. Expansion of Scylla farming has led to increased efforts to understand the physiology and behavior of these crabs, and as such, there are growing molecular resources for them. Here, publicly accessible Scylla olivacea transcriptomic data were mined for putative peptide-encoding transcripts; the proteins deduced from the identified sequences were then used to predict the structures of mature peptide hormones. Forty-nine pre/preprohormone-encoding transcripts were identified, allowing for the prediction of 187 distinct mature peptides. The identified peptides included isoforms of adipokinetic hormone-corazonin-like peptide, allatostatin A, allatostatin B, allatostatin C, bursicon β, CCHamide, corazonin, crustacean cardioactive peptide, crustacean hyperglycemic hormone/molt-inhibiting hormone, diuretic hormone 31, eclosion hormone, FMRFamide-like peptide, HIGSLYRamide, insulin-like peptide, intocin, leucokinin, myosuppressin, neuroparsin, neuropeptide F, orcokinin, pigment dispersing hormone, pyrokinin, red pigment concentrating hormone, RYamide, short neuropeptide F, SIFamide and tachykinin-related peptide, all well-known neuropeptide families. Surprisingly, the tissue used to generate the transcriptome mined here is reported to be testis. Whether or not the testis samples had neural contamination is unknown. However, if the peptides are truly produced by this reproductive organ, it could have far reaching consequences for the study of crustacean endocrinology, particularly in the area of reproductive control. Regardless, this peptidome is the largest thus far predicted for any brachyuran (true crab) species, and will serve as a foundation for future studies of peptidergic control in members of the commercially important genus Scylla. Copyright © 2016 Elsevier Inc. All rights reserved.

  10. Optimization of short amino acid sequences classifier

    Science.gov (United States)

    Barcz, Aleksy; Szymański, Zbigniew

    This article describes processing methods used for short amino acid sequences classification. The data processed are 9-symbols string representations of amino acid sequences, divided into 49 data sets - each one containing samples labeled as reacting or not with given enzyme. The goal of the classification is to determine for a single enzyme, whether an amino acid sequence would react with it or not. Each data set is processed separately. Feature selection is performed to reduce the number of dimensions for each data set. The method used for feature selection consists of two phases. During the first phase, significant positions are selected using Classification and Regression Trees. Afterwards, symbols appearing at the selected positions are substituted with numeric values of amino acid properties taken from the AAindex database. In the second phase the new set of features is reduced using a correlation-based ranking formula and Gram-Schmidt orthogonalization. Finally, the preprocessed data is used for training LS-SVM classifiers. SPDE, an evolutionary algorithm, is used to obtain optimal hyperparameters for the LS-SVM classifier, such as error penalty parameter C and kernel-specific hyperparameters. A simple score penalty is used to adapt the SPDE algorithm to the task of selecting classifiers with best performance measures values.

  11. Sequence diversity and evolution of antimicrobial peptides in invertebrates.

    Science.gov (United States)

    Tassanakajon, Anchalee; Somboonwiwat, Kunlaya; Amparyup, Piti

    2015-02-01

    Antimicrobial peptides (AMPs) are evolutionarily ancient molecules that act as the key components in the invertebrate innate immunity against invading pathogens. Several AMPs have been identified and characterized in invertebrates, and found to display considerable diversity in their amino acid sequence, structure and biological activity. AMP genes appear to have rapidly evolved, which might have arisen from the co-evolutionary arms race between host and pathogens, and enabled organisms to survive in different microbial environments. Here, the sequence diversity of invertebrate AMPs (defensins, cecropins, crustins and anti-lipopolysaccharide factors) are presented to provide a better understanding of the evolution pattern of these peptides that play a major role in host defense mechanisms. Copyright © 2014 Elsevier Ltd. All rights reserved.

  12. Enhancement of the Enterocin CRL35 Activity by a Synthetic Peptide Derived from the NH2-Terminal Sequence

    Science.gov (United States)

    Saavedra, Lucila; Minahk, Carlos; de Ruiz Holgado, Aída P.; Sesma, Fernando

    2004-01-01

    The enterocin CRL35 biosynthetic gene cluster was cloned and sequenced. The sequence was revealed to be highly identical to that of the mundticin KS gene cluster (S. Kawamoto, J. Shima, R. Sato, T. Eguchi, S. Ohmomo, J. Shibato, N. Horikoshi, K. Takeshita, and T. Sameshima, Appl. Environ. Microbiol. 68:3830-3840, 2002). Short synthetic peptides were designed based on the bacteriocin sequence and were evaluated in antimicrobial competitive assays. The peptide KYYGNGVSCNKKGCS produced an enhancement of enterocin CRL35 antimicrobial activity in a buffer system. PMID:15215149

  13. Hypothesis driven development of new adjuvants: short peptides as immunomodulators.

    Science.gov (United States)

    Dong, Jessica C; Kobinger, Gary P

    2013-04-01

    To date, vaccinations have been one of the key strategies in the prevention and protection against infectious pathogens. Traditional vaccines have well-known limitations such as safety and efficacy issues, which consequently deems it inappropriate for particular populations and may not be an effective strategy against all pathogens. This evidence highlights the need to develop more efficacious vaccination regiments. Higher levels of protection can be achieved by the addition of immunostimulating adjuvants. Many adjuvants elicit strong, undefined inflammation, which produces increased immunogenicity but may also lead to undesirable effects. Hypothesis driven development of adjuvants is needed to achieve a more specific and directed immune response required for optimal and safe vaccine-induced immune protection. An example of such hypothesis driven development includes the use of short immunomodulating peptides as adjuvants. These peptides have the ability to influence the immune response and can be extrapolated for adjuvant use, but requires further investigation.

  14. Partial sequence determination of metabolically labeled radioactive proteins and peptides

    International Nuclear Information System (INIS)

    Anderson, C.W.

    1982-01-01

    The author has used the sequence analysis of radioactive proteins and peptides to approach several problems during the past few years. They, in collaboration with others, have mapped precisely several adenovirus proteins with respect to the nucleotide sequence of the adenovirus genome; identified hitherto missed proteins encoded by bacteriophage MS2 and by simian virus 40; analyzed the aminoterminal maturation of several virus proteins; determined the cleavage sites for processing of the poliovirus polyprotein; and analyzed the mechanism of frameshifting by excess normal tRNAs during cell-free protein synthesis. This chapter is designed to aid those without prior experience at protein sequence determinations. It is based primarily on the experience gained in the studies cited above, which made use of the Beckman 890 series automated protein sequencers

  15. Short read sequence typing (SRST: multi-locus sequence types from short reads

    Directory of Open Access Journals (Sweden)

    Inouye Michael

    2012-07-01

    Full Text Available Abstract Background Multi-locus sequence typing (MLST has become the gold standard for population analyses of bacterial pathogens. This method focuses on the sequences of a small number of loci (usually seven to divide the population and is simple, robust and facilitates comparison of results between laboratories and over time. Over the last decade, researchers and population health specialists have invested substantial effort in building up public MLST databases for nearly 100 different bacterial species, and these databases contain a wealth of important information linked to MLST sequence types such as time and place of isolation, host or niche, serotype and even clinical or drug resistance profiles. Recent advances in sequencing technology mean it is increasingly feasible to perform bacterial population analysis at the whole genome level. This offers massive gains in resolving power and genetic profiling compared to MLST, and will eventually replace MLST for bacterial typing and population analysis. However given the wealth of data currently available in MLST databases, it is crucial to maintain backwards compatibility with MLST schemes so that new genome analyses can be understood in their proper historical context. Results We present a software tool, SRST, for quick and accurate retrieval of sequence types from short read sets, using inputs easily downloaded from public databases. SRST uses read mapping and an allele assignment score incorporating sequence coverage and variability, to determine the most likely allele at each MLST locus. Analysis of over 3,500 loci in more than 500 publicly accessible Illumina read sets showed SRST to be highly accurate at allele assignment. SRST output is compatible with common analysis tools such as eBURST, Clonal Frame or PhyloViz, allowing easy comparison between novel genome data and MLST data. Alignment, fastq and pileup files can also be generated for novel alleles. Conclusions SRST is a novel

  16. Inhibition of Plant-Pathogenic Bacteria by Short Synthetic Cecropin A-Melittin Hybrid Peptides

    OpenAIRE

    Ferre, Rafael; Badosa, Esther; Feliu, Lidia; Planas, Marta; Montesinos, Emili; Bardají, Eduard

    2006-01-01

    Short peptides of 11 residues were synthesized and tested against the economically important plant pathogenic bacteria Erwinia amylovora, Pseudomonas syringae, and Xanthomonas vesicatoria and compared to the previously described peptide Pep3 (WKLFKKILKVL-NH2). The antimicrobial activity of Pep3 and 22 analogues was evaluated in terms of the MIC and the 50% effective dose (ED50) for growth. Peptide cytotoxicity against human red blood cells and peptide stability toward protease degradation wer...

  17. Error Analysis of Deep Sequencing of Phage Libraries: Peptides Censored in Sequencing

    Directory of Open Access Journals (Sweden)

    Wadim L. Matochko

    2013-01-01

    Full Text Available Next-generation sequencing techniques empower selection of ligands from phage-display libraries because they can detect low abundant clones and quantify changes in the copy numbers of clones without excessive selection rounds. Identification of errors in deep sequencing data is the most critical step in this process because these techniques have error rates >1%. Mechanisms that yield errors in Illumina and other techniques have been proposed, but no reports to date describe error analysis in phage libraries. Our paper focuses on error analysis of 7-mer peptide libraries sequenced by Illumina method. Low theoretical complexity of this phage library, as compared to complexity of long genetic reads and genomes, allowed us to describe this library using convenient linear vector and operator framework. We describe a phage library as N×1 frequency vector n=ni, where ni is the copy number of the ith sequence and N is the theoretical diversity, that is, the total number of all possible sequences. Any manipulation to the library is an operator acting on n. Selection, amplification, or sequencing could be described as a product of a N×N matrix and a stochastic sampling operator (Sa. The latter is a random diagonal matrix that describes sampling of a library. In this paper, we focus on the properties of Sa and use them to define the sequencing operator (Seq. Sequencing without any bias and errors is Seq=Sa IN, where IN is a N×N unity matrix. Any bias in sequencing changes IN to a nonunity matrix. We identified a diagonal censorship matrix (CEN, which describes elimination or statistically significant downsampling, of specific reads during the sequencing process.

  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. 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.

  20. 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...

  1. Peptomics, identification of novel cationic Arabidopsis peptides with conserved sequence motifs

    DEFF Research Database (Denmark)

    Olsen, Addie Nina; Mundy, John; Skriver, Karen

    2002-01-01

    Arabidopsis family of 34 genes. The predicted peptides are characterized by a conserved C-terminal sequence motif and additional primary structure conservation in a core region. The majority of these genes had not previously been annotated. A subset of the predicted peptides show high overall sequence...... similarity to Rapid Alkalinization Factor (RALF), a peptide isolated from tobacco. We therefore refer to this peptide family as RALFL for RALF-Like. RT-PCR analysis confirmed that several of the Arabidopsis genes are expressed and that their expression patterns vary. The identification of a large gene family...

  2. 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.

  3. A short TE gradient-echo sequence using asymmetric sampling

    International Nuclear Information System (INIS)

    Fujita, Norihiko; Harada, Kohshi; Sakurai, Kosuke; Nakanishi, Katsuyuki; Kim, Shyogen; Kozuka, Takahiro

    1990-01-01

    We have developed a gradient-echo pulse sequence with a short TE less than 4 msec using a data set of asymmetric off-center sampling with a broad bandwidth. The use of such a short TE significantly reduces T 2 * dephasing effect even in a two-dimensional mode, and by collecting an off-center echo, motion-induced phase dispersion is also considerably decreased. High immunity of this sequence to these dephasing effects permits clear visualization of anatomical details near the skull base where large local field inhomogeneities and rapid blood flow such as in the internal carotid artery are present. (author)

  4. 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.

  5. Prediction of peptide drift time in ion mobility mass spectrometry from sequence-based features

    KAUST Repository

    Wang, Bing; Zhang, Jun; Chen, Peng; Ji, Zhiwei; Deng, Shuping; Li, Chi

    2013-01-01

    Background: Ion mobility-mass spectrometry (IMMS), an analytical technique which combines the features of ion mobility spectrometry (IMS) and mass spectrometry (MS), can rapidly separates ions on a millisecond time-scale. IMMS becomes a powerful tool to analyzing complex mixtures, especially for the analysis of peptides in proteomics. The high-throughput nature of this technique provides a challenge for the identification of peptides in complex biological samples. As an important parameter, peptide drift time can be used for enhancing downstream data analysis in IMMS-based proteomics.Results: In this paper, a model is presented based on least square support vectors regression (LS-SVR) method to predict peptide ion drift time in IMMS from the sequence-based features of peptide. Four descriptors were extracted from peptide sequence to represent peptide ions by a 34-component vector. The parameters of LS-SVR were selected by a grid searching strategy, and a 10-fold cross-validation approach was employed for the model training and testing. Our proposed method was tested on three datasets with different charge states. The high prediction performance achieve demonstrate the effectiveness and efficiency of the prediction model.Conclusions: Our proposed LS-SVR model can predict peptide drift time from sequence information in relative high prediction accuracy by a test on a dataset of 595 peptides. This work can enhance the confidence of protein identification by combining with current protein searching techniques. 2013 Wang et al.; licensee BioMed Central Ltd.

  6. Prediction of peptide drift time in ion mobility mass spectrometry from sequence-based features

    KAUST Repository

    Wang, Bing

    2013-05-09

    Background: Ion mobility-mass spectrometry (IMMS), an analytical technique which combines the features of ion mobility spectrometry (IMS) and mass spectrometry (MS), can rapidly separates ions on a millisecond time-scale. IMMS becomes a powerful tool to analyzing complex mixtures, especially for the analysis of peptides in proteomics. The high-throughput nature of this technique provides a challenge for the identification of peptides in complex biological samples. As an important parameter, peptide drift time can be used for enhancing downstream data analysis in IMMS-based proteomics.Results: In this paper, a model is presented based on least square support vectors regression (LS-SVR) method to predict peptide ion drift time in IMMS from the sequence-based features of peptide. Four descriptors were extracted from peptide sequence to represent peptide ions by a 34-component vector. The parameters of LS-SVR were selected by a grid searching strategy, and a 10-fold cross-validation approach was employed for the model training and testing. Our proposed method was tested on three datasets with different charge states. The high prediction performance achieve demonstrate the effectiveness and efficiency of the prediction model.Conclusions: Our proposed LS-SVR model can predict peptide drift time from sequence information in relative high prediction accuracy by a test on a dataset of 595 peptides. This work can enhance the confidence of protein identification by combining with current protein searching techniques. 2013 Wang et al.; licensee BioMed Central Ltd.

  7. Peptides derivatized with bicyclic quaternary ammonium ionization tags. Sequencing via tandem mass spectrometry.

    Science.gov (United States)

    Setner, Bartosz; Rudowska, Magdalena; Klem, Ewelina; Cebrat, Marek; Szewczuk, Zbigniew

    2014-10-01

    Improving the sensitivity of detection and fragmentation of peptides to provide reliable sequencing of peptides is an important goal of mass spectrometric analysis. Peptides derivatized by bicyclic quaternary ammonium ionization tags: 1-azabicyclo[2.2.2]octane (ABCO) or 1,4-diazabicyclo[2.2.2]octane (DABCO), are characterized by an increased detection sensitivity in electrospray ionization mass spectrometry (ESI-MS) and longer retention times on the reverse-phase (RP) chromatography columns. The improvement of the detection limit was observed even for peptides dissolved in 10 mM NaCl. Collision-induced dissociation tandem mass spectrometry of quaternary ammonium salts derivatives of peptides showed dominant a- and b-type ions, allowing facile sequencing of peptides. The bicyclic ionization tags are stable in collision-induced dissociation experiments, and the resulted fragmentation pattern is not significantly influenced by either acidic or basic amino acid residues in the peptide sequence. Obtained results indicate the general usefulness of the bicyclic quaternary ammonium ionization tags for ESI-MS/MS sequencing of peptides. Copyright © 2014 John Wiley & Sons, Ltd.

  8. 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.

  9. Blocking the RecA activity and SOS-response in bacteria with a short α-helical peptide.

    Science.gov (United States)

    Yakimov, Alexander; Pobegalov, Georgii; Bakhlanova, Irina; Khodorkovskii, Mikhail; Petukhov, Michael; Baitin, Dmitry

    2017-09-19

    The RecX protein, a very active natural RecA protein inhibitor, can completely disassemble RecA filaments at nanomolar concentrations that are two to three orders of magnitude lower than that of RecA protein. Based on the structure of RecX protein complex with the presynaptic RecA filament, we designed a short first in class α-helical peptide that both inhibits RecA protein activities in vitro and blocks the bacterial SOS-response in vivo. The peptide was designed using SEQOPT, a novel method for global sequence optimization of protein α-helices. SEQOPT produces artificial peptide sequences containing only 20 natural amino acids with the maximum possible conformational stability at a given pH, ionic strength, temperature, peptide solubility. It also accounts for restrictions due to known amino acid residues involved in stabilization of protein complexes under consideration. The results indicate that a few key intermolecular interactions inside the RecA protein presynaptic complex are enough to reproduce the main features of the RecX protein mechanism of action. Since the SOS-response provides a major mechanism of bacterial adaptation to antibiotics, these results open new ways for the development of antibiotic co-therapy that would not cause bacterial resistance. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

  10. Aggregation and toxicity of amyloid-beta peptide in relation to peptide sequence variation

    OpenAIRE

    Vandersteen, A.

    2012-01-01

    Generally, aggregation of the amyloid-ß peptide is considered the cause of neuronal death in Alzheimer disease. The heterogenous Aß peptide occurs in various lengths in vivo: Aß40 and Aß42 are the predominant forms while both shorter and longer peptides exist. Aß40 and shorter isoforms are less aggregation-prone and hence considered less dangerous than Aß42 and longer isoforms, which are more aggregation-prone. Up to now research mainly focussed on the predominant Aß peptides and their indivi...

  11. Practical multipeptide synthesis: dedicated software for the definition of multiple, overlapping peptides covering polypeptide sequences.

    Science.gov (United States)

    Heegaard, P M; Holm, A; Hagerup, M

    1993-01-01

    A personal computer program for the conversion of linear amino acid sequences to multiple, small, overlapping peptide sequences has been developed. Peptide lengths and "jumps" (the distance between two consecutive overlapping peptides) are defined by the user. To facilitate the use of the program for parallel solid-phase chemical peptide syntheses for the synchronous production of multiple peptides, amino acids at each acylation step are laid out by the program in a convenient standard multi-well setup. Also, the total number of equivalents, as well as the derived amount in milligrams (depend-ending on user-defined equivalent weights and molar surplus), of each amino acid are given. The program facilitates the implementation of multipeptide synthesis, e.g., for the elucidation of polypeptide structure-function relationships, and greatly reduces the risk of introducing mistakes at the planning step. It is written in Pascal and runs on any DOS-based personal computer. No special graphic display is needed.

  12. BlockLogo: Visualization of peptide and sequence motif conservation

    DEFF Research Database (Denmark)

    Olsen, Lars Rønn; Kudahl, Ulrich Johan; Simon, Christian

    2013-01-01

    BlockLogo is a web-server application for the visualization of protein and nucleotide fragments, continuous protein sequence motifs, and discontinuous sequence motifs using calculation of block entropy from multiple sequence alignments. The user input consists of a multiple sequence alignment, se...

  13. Short, multiple-stranded β-hairpin peptides have antimicrobial potency with high selectivity and salt resistance.

    Science.gov (United States)

    Chou, Shuli; Shao, Changxuan; Wang, Jiajun; Shan, Anshan; Xu, Lin; Dong, Na; Li, Zhongyu

    2016-01-01

    The β-hairpin structure has been proposed to exhibit potent antimicrobial properties with low cytotoxicity, thus, multiple β-hairpin structures have been proved to be highly stable in structures containing tightly packed hydrophobic cores. The aim of this study was to develop peptide-based synthetic strategies for generating short, but effective AMPs as inexpensive antimicrobial agents. Multiple-stranded β-hairpin peptides with the same β-hairpin unit, (WRXxRW)n where n=1, 2, 3, or 4 and Xx represent the turn sequence, were synthesized, and their potential as antimicrobial agents was evaluated. Owning to the tightly packed hydrophobic core and paired Trp of this multiple-stranded β-hairpin structure, all the 12-residues peptides exhibited high cell selectivity towards bacterial cells over human red blood cells (hRBCs), and the peptide W2 exhibited stronger antimicrobial activities with the MIC values of 2-8μM against various tested bacteria. Not only that, but W2 also showed obvious synergy with streptomycin and chloramphenicol against Escherichia coli, and displayed synergy with ciprofloxacin against Staphylococcus aureus with the FICI values ⩽0.5. Fluorescence spectroscopy and electron microscopy analyses indicated that W2 kills microbial cells by permeabilizing the cell membrane and damaging membrane integrity. Collectively, based on the multiple β-hairpin peptides, the ability to develop libraries of short and effective peptides will be a powerful approach to the discovery of novel antimicrobial agents. We successfully screened a peptide W2 ((WRPGRW)2) from a series of multiple-stranded β-hairpin antimicrobial peptides based on the "S-shaped" motif that induced the formation of a globular structure, and Trp zipper was used to replace the disulfide bonds to reduce the cost of production. This novel structure applied to AMPs improved cell selectivity and salt stability. The findings of this study will promote the development of peptide

  14. 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.

  15. Proteomics shows Hsp70 does not bind peptide sequences indiscriminately in vivo

    International Nuclear Information System (INIS)

    Grossmann, Michael E.; Madden, Benjamin J.; Gao, Fan; Pang, Yuan-Ping; Carpenter, John E.; McCormick, Daniel; Young, Charles Y.F.

    2004-01-01

    Heat shock protein 70 (Hsp70) binds peptide and has several functions that include protein folding, protein trafficking, and involvement with immune function. However, endogenous Hsp70-binding peptides had not previously been identified. Therefore, we eluted and identified several hundred endogenously bound peptides from Hsp70 using liquid chromatography ion trap mass spectrophotometry (LC-ITMS). Our work shows that the peptides are capable of binding Hsp70 as previously described. They are generally 8-26 amino acids in length and correspond to specific regions of many proteins. Through computationally assisted analysis of peptides eluted from Hsp70 we determined variable amino acid sequences, including a 5 amino acid core sequence that Hsp70 favorably binds. We also developed a computer algorithm that predicts Hsp70 binding within proteins. This work helps to define what peptides are bound by Hsp70 in vivo and suggests that Hsp70 facilitates peptide selection by aiding a funneling mechanism that is flexible but allows only a limited number of peptides to be processed

  16. Studying a free fall experiment using short sequences of images

    International Nuclear Information System (INIS)

    Vera, Francisco; Romanque, Cristian

    2008-01-01

    We discuss a new alternative for obtaining position and time coordinates from a video of a free fall experiment. In our approach, after converting the video to a short sequence of images, the images are analyzed using a web page application developed by the author. The main advantage of the setup explained in this work, is that it is simple to use, no software license fees are necessary, and can be scaled-up to be used by a big number of students in introductory physics courses. The steps involved in the full analysis of a falling object are: we grab a short digital video of the experiment and convert it to a sequence of images, then, using a web page that includes all the necessary javascript, the student can easily click on the object of interest to obtain the (x,y,t) coordinates, finally, the student analyze motion using a spreadsheet.

  17. Sequence-Specific Model for Peptide Retention Time Prediction in Strong Cation Exchange Chromatography.

    Science.gov (United States)

    Gussakovsky, Daniel; Neustaeter, Haley; Spicer, Victor; Krokhin, Oleg V

    2017-11-07

    The development of a peptide retention prediction model for strong cation exchange (SCX) separation on a Polysulfoethyl A column is reported. Off-line 2D LC-MS/MS analysis (SCX-RPLC) of S. cerevisiae whole cell lysate was used to generate a retention dataset of ∼30 000 peptides, sufficient for identifying the major sequence-specific features of peptide retention mechanisms in SCX. In contrast to RPLC/hydrophilic interaction liquid chromatography (HILIC) separation modes, where retention is driven by hydrophobic/hydrophilic contributions of all individual residues, SCX interactions depend mainly on peptide charge (number of basic residues at acidic pH) and size. An additive model (incorporating the contributions of all 20 residues into the peptide retention) combined with a peptide length correction produces a 0.976 R 2 value prediction accuracy, significantly higher than the additive models for either HILIC or RPLC. Position-dependent effects on peptide retention for different residues were driven by the spatial orientation of tryptic peptides upon interaction with the negatively charged surface functional groups. The positively charged N-termini serve as a primary point of interaction. For example, basic residues (Arg, His, Lys) increase peptide retention when located closer to the N-terminus. We also found that hydrophobic interactions, which could lead to a mixed-mode separation mechanism, are largely suppressed at 20-30% of acetonitrile in the eluent. The accuracy of the final Sequence-Specific Retention Calculator (SSRCalc) SCX model (∼0.99 R 2 value) exceeds all previously reported predictors for peptide LC separations. This also provides a solid platform for method development in 2D LC-MS protocols in proteomics and peptide retention prediction filtering of false positive identifications.

  18. Short term aerobic exercise training increases postprandial pancreatic polypeptide but not peptide YY concentrations in obese individuals

    OpenAIRE

    Kanaley, Jill A.; Heden, Timothy D.; Liu, Ying; Whaley-Connell, Adam T.; Chockalingam, Anand; Dellsperger, Kevin C.; Fairchild, Timothy J.

    2013-01-01

    Objective Short-term exercise training improves glycemic control, but the effect of short-term training on postprandial satiety peptide responses or perceived satiety remains unknown. We tested the hypothesis that short-term aerobic exercise training (15 days) would alter postprandial pancreatic and gut peptide [pancreatic polypeptide (PP) and peptide YY (PYY)] responses and perceived appetite and satiety in obese individuals. Subjects Thirteen healthy obese men and women (age: 42±2 y; BMI: 3...

  19. SAAS: Short Amino Acid Sequence - A Promising Protein Secondary Structure Prediction Method of Single Sequence

    Directory of Open Access Journals (Sweden)

    Zhou Yuan Wu

    2013-07-01

    Full Text Available In statistical methods of predicting protein secondary structure, many researchers focus on single amino acid frequencies in α-helices, β-sheets, and so on, or the impact near amino acids on an amino acid forming a secondary structure. But the paper considers a short sequence of amino acids (3, 4, 5 or 6 amino acids as integer, and statistics short sequence's probability forming secondary structure. Also, many researchers select low homologous sequences as statistical database. But this paper select whole PDB database. In this paper we propose a strategy to predict protein secondary structure using simple statistical method. Numerical computation shows that, short amino acids sequence as integer to statistics, which can easy see trend of short sequence forming secondary structure, and it will work well to select large statistical database (whole PDB database without considering homologous, and Q3 accuracy is ca. 74% using this paper proposed simple statistical method, but accuracy of others statistical methods is less than 70%.

  20. A fossil protein chimera; difficulties in discriminating dinosaur peptide sequences from modern cross-contamination.

    Science.gov (United States)

    Buckley, Michael; Warwood, Stacey; van Dongen, Bart; Kitchener, Andrew C; Manning, Phillip L

    2017-05-31

    A decade ago, reports that organic-rich soft tissue survived from dinosaur fossils were apparently supported by proteomics-derived sequence information of exceptionally well-preserved bone. This initial claim to the sequencing of endogenous collagen peptides from an approximately 68 Myr Tyrannosaurus rex fossil was highly controversial, largely on the grounds of potential contamination from either bacterial biofilms or from laboratory practice. In a subsequent study, collagen peptide sequences from an approximately 78 Myr Brachylophosaurus canadensis fossil were reported that have remained largely unchallenged. However, the endogeneity of these sequences relies heavily on a single peptide sequence, apparently unique to both dinosaurs. Given the potential for cross-contamination from modern bone analysed by the same team, here we extract collagen from bone samples of three individuals of ostrich, Struthio camelus The resulting LC-MS/MS data were found to match all of the proposed sequences for both the original Tyrannosaurus and Brachylophosaurus studies. Regardless of the true nature of the dinosaur peptides, our finding highlights the difficulty of differentiating such sequences with confidence. Our results not only imply that cross-contamination cannot be ruled out, but that appropriate measures to test for endogeneity should be further evaluated. © 2017 The Authors.

  1. A fossil protein chimera; difficulties in discriminating dinosaur peptide sequences from modern cross-contamination

    Science.gov (United States)

    Warwood, Stacey; van Dongen, Bart; Kitchener, Andrew C.; Manning, Phillip L.

    2017-01-01

    A decade ago, reports that organic-rich soft tissue survived from dinosaur fossils were apparently supported by proteomics-derived sequence information of exceptionally well-preserved bone. This initial claim to the sequencing of endogenous collagen peptides from an approximately 68 Myr Tyrannosaurus rex fossil was highly controversial, largely on the grounds of potential contamination from either bacterial biofilms or from laboratory practice. In a subsequent study, collagen peptide sequences from an approximately 78 Myr Brachylophosaurus canadensis fossil were reported that have remained largely unchallenged. However, the endogeneity of these sequences relies heavily on a single peptide sequence, apparently unique to both dinosaurs. Given the potential for cross-contamination from modern bone analysed by the same team, here we extract collagen from bone samples of three individuals of ostrich, Struthio camelus. The resulting LC–MS/MS data were found to match all of the proposed sequences for both the original Tyrannosaurus and Brachylophosaurus studies. Regardless of the true nature of the dinosaur peptides, our finding highlights the difficulty of differentiating such sequences with confidence. Our results not only imply that cross-contamination cannot be ruled out, but that appropriate measures to test for endogeneity should be further evaluated. PMID:28566488

  2. Complete sequences of glucagon-like peptide-1 from human and pig small intestine

    DEFF Research Database (Denmark)

    Orskov, C; Bersani, M; Johnsen, A H

    1989-01-01

    intestine of the proglucagon precursor were determined by pairs of basic amino acid residues flanking the two peptides. Earlier studies have shown that synthetic glucagon-like peptide-1 (GLP-1) synthesized according to the proposed structure (proglucagon 71-108 or because residue 108 is Gly, 72-107 amide......) had no physiological effects, whereas a truncated from of GLP-1, corresponding to proglucagon 78-107 amide, strongly stimulated insulin secretion and depressed glucagon secretion. To determine the amino acid sequence of the naturally occurring peptide we isolated GLP-1 from human small intestine...

  3. Catalytic center of lecithin:cholesterol acyltransferase: isolation and sequence of diisopropyl fluorophosphate-labeled peptides

    Energy Technology Data Exchange (ETDEWEB)

    Park, Y.B.; Yueksel, U.G.; Gracy, R.W.; Lacko, A.G.

    1987-02-27

    Lecithin:cholesterol acyltransferase (LCAT) was purified from hog plasma and subsequently reacted with (/sup 3/H)-Diisopropyl fluorophosphate (DFP). The labeled enzyme was digested with pepsin and the peptides separated by high performance liquid chromatography (HPLC). Two radioactive peptides were isolated, subjected to automated amino acid sequencing and yielded the following data: A) Ile-Ser-Leu-Gly-Ala-Pro-Trp-Gly-Gly-Ser, and B) Tyr-Ile-Phe-Asp-x-Gly-Phe-Pro-Tyr-x-Asp-Pro-Val. Both of these sequences represent very highly conserved regions of the enzyme when compared to the sequence of human LCAT. Peptide (A) is considered to represent the catalytic center of LCAT based on comparisons with data reported in the literature.

  4. Short Peptides Enhance Single Cell Adhesion and Viability onMicroarrays

    Energy Technology Data Exchange (ETDEWEB)

    Veiseh, Mandana; Veiseh, Omid; Martin, Michael C.; Asphahani,Fareid; Zhang, Miqin

    2007-01-19

    Single cell patterning holds important implications forbiology, biochemistry, biotechnology, medicine, and bioinformatics. Thechallenge for single cell patterning is to produce small islands hostingonly single cells and retaining their viability for a prolonged period oftime. This study demonstrated a surface engineering approach that uses acovalently bound short peptide as a mediator to pattern cells withimproved single cell adhesion and prolonged cellular viabilityon goldpatterned SiO2 substrates. The underlying hypothesis is that celladhesion is regulated bythe type, availability, and stability ofeffective cell adhesion peptides, and thus covalently bound shortpeptides would promote cell spreading and, thus, single cell adhesion andviability. The effectiveness of this approach and the underlyingmechanism for the increased probability of single cell adhesion andprolonged cell viability by short peptides were studied by comparingcellular behavior of human umbilical cord vein endothelial cells on threemodelsurfaces whose gold electrodes were immobilized with fibronectin,physically adsorbed Arg-Glu-Asp-Val-Tyr, and covalently boundLys-Arg-Glu-Asp-Val-Tyr, respectively. The surface chemistry and bindingproperties were characterized by reflectance Fourier transform infraredspectroscopy. Both short peptides were superior to fibronectin inproducing adhesion of only single cells, whereas the covalently boundpeptide also reduced apoptosis and necrosisof adhered cells. Controllingcell spreading by peptide binding domains to regulate apoptosis andviability represents a fundamental mechanism in cell-materialsinteraction and provides an effective strategy in engineering arrays ofsingle cells.

  5. Mapping membrane activity in undiscovered peptide sequence space using machine learning.

    Science.gov (United States)

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

    2016-11-29

    There are some ∼1,100 known antimicrobial peptides (AMPs), which permeabilize microbial membranes but have diverse sequences. Here, we develop a support vector machine (SVM)-based classifier to investigate ⍺-helical AMPs and the interrelated nature of their functional commonality and sequence homology. SVM is used to search the undiscovered peptide sequence space and identify Pareto-optimal candidates that simultaneously maximize the distance σ from the SVM hyperplane (thus maximize its "antimicrobialness") and its ⍺-helicity, but minimize mutational distance to known AMPs. By calibrating SVM machine learning results with killing assays and small-angle X-ray scattering (SAXS), we find that the SVM metric σ correlates not with a peptide's minimum inhibitory concentration (MIC), but rather its ability to generate negative Gaussian membrane curvature. This surprising result provides a topological basis for membrane activity common to AMPs. Moreover, we highlight an important distinction between the maximal recognizability of a sequence to a trained AMP classifier (its ability to generate membrane curvature) and its maximal antimicrobial efficacy. As mutational distances are increased from known AMPs, we find AMP-like sequences that are increasingly difficult for nature to discover via simple mutation. Using the sequence map as a discovery tool, we find a unexpectedly diverse taxonomy of sequences that are just as membrane-active as known AMPs, but with a broad range of primary functions distinct from AMP functions, including endogenous neuropeptides, viral fusion proteins, topogenic peptides, and amyloids. The SVM classifier is useful as a general detector of membrane activity in peptide sequences.

  6. The Cysteine S-Alkylation Reaction as a Synthetic Method to Covalently Modify Peptide Sequences.

    Science.gov (United States)

    Calce, Enrica; De Luca, Stefania

    2017-01-05

    Synthetic methodologies to chemically modify peptide molecules have long been investigated for their impact in the field of chemical biology. They allow the introduction of biochemical probes useful for studying protein functions, for manipulating peptides with therapeutic potential, and for structure-activity relationship investigations. The commonly used approach was the derivatization of an amino acid side chain. In this regard, the cysteine, for its unique reactivity, has been widely employed as the substrate for such modifications. Herein, we report on methodologies developed to modify the cysteine thiol group through the S-alkylation reaction. Some procedures perform the alkylation of cysteine derivatives, in order to prepare building blocks to be used during the peptide synthesis, whilst some others selectively modify peptide sequences containing a cysteine residue with a free thiol group, both in solution and in the solid phase. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  7. 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

  8. Sequence-selective targeting of duplex DNA by peptide nucleic acids

    DEFF Research Database (Denmark)

    Nielsen, Peter E

    2010-01-01

    Sequence-selective gene targeting constitutes an attractive drug-discovery approach for genetic therapy, with the aim of reducing or enhancing the activity of specific genes at the transcriptional level, or as part of a methodology for targeted gene repair. The pseudopeptide DNA mimic peptide...

  9. Identification of Alternative Splice Variants Using Unique Tryptic Peptide Sequences for Database Searches.

    Science.gov (United States)

    Tran, Trung T; Bollineni, Ravi C; Strozynski, Margarita; Koehler, Christian J; Thiede, Bernd

    2017-07-07

    Alternative splicing is a mechanism in eukaryotes by which different forms of mRNAs are generated from the same gene. Identification of alternative splice variants requires the identification of peptides specific for alternative splice forms. For this purpose, we generated a human database that contains only unique tryptic peptides specific for alternative splice forms from Swiss-Prot entries. Using this database allows an easy access to splice variant-specific peptide sequences that match to MS data. Furthermore, we combined this database without alternative splice variant-1-specific peptides with human Swiss-Prot. This combined database can be used as a general database for searching of LC-MS data. LC-MS data derived from in-solution digests of two different cell lines (LNCaP, HeLa) and phosphoproteomics studies were analyzed using these two databases. Several nonalternative splice variant-1-specific peptides were found in both cell lines, and some of them seemed to be cell-line-specific. Control and apoptotic phosphoproteomes from Jurkat T cells revealed several nonalternative splice variant-1-specific peptides, and some of them showed clear quantitative differences between the two states.

  10. Next-generation ELISA diagnostic assay for Chagas Disease based on the combination of short peptidic epitopes.

    Directory of Open Access Journals (Sweden)

    Juan Mucci

    2017-10-01

    Full Text Available Chagas Disease, caused by the protozoan Trypanosoma cruzi, is a major health and economic problem in Latin America for which no vaccine or appropriate drugs for large-scale public health interventions are yet available. Accurate diagnosis is essential for the early identification and follow up of vector-borne cases and to prevent transmission of the disease by way of blood transfusions and organ transplantation. Diagnosis is routinely performed using serological methods, some of which require the production of parasite lysates, parasite antigenic fractions or purified recombinant antigens. Although available serological tests give satisfactory results, the production of reliable reagents remains laborious and expensive. Short peptides spanning linear B-cell epitopes have proven ideal serodiagnostic reagents in a wide range of diseases. Recently, we have conducted a large-scale screening of T. cruzi linear B-cell epitopes using high-density peptide chips, leading to the identification of several hundred novel sequence signatures associated to chronic Chagas Disease. Here, we performed a serological assessment of 27 selected epitopes and of their use in a novel multipeptide-based diagnostic method. A combination of 7 of these peptides were finally evaluated in ELISA format against a panel of 199 sera samples (Chagas-positive and negative, including sera from Leishmaniasis-positive subjects. The multipeptide formulation displayed a high diagnostic performance, with a sensitivity of 96.3% and a specificity of 99.15%. Therefore, the use of synthetic peptides as diagnostic tools are an attractive alternative in Chagas' disease diagnosis.

  11. Toward Structure Prediction for Short Peptides Using the Improved SAAP Force Field Parameters

    Directory of Open Access Journals (Sweden)

    Kenichi Dedachi

    2013-01-01

    Full Text Available Based on the observation that Ramachandran-type potential energy surfaces of single amino acid units in water are in good agreement with statistical structures of the corresponding amino acid residues in proteins, we recently developed a new all-atom force field called SAAP, in which the total energy function for a polypeptide is expressed basically as a sum of single amino acid potentials and electrostatic and Lennard-Jones potentials between the amino acid units. In this study, the SAAP force field (SAAPFF parameters were improved, and classical canonical Monte Carlo (MC simulation was carried out for short peptide models, that is, Met-enkephalin and chignolin, at 300 K in an implicit water model. Diverse structures were reasonably obtained for Met-enkephalin, while three folded structures, one of which corresponds to a native-like structure with three native hydrogen bonds, were obtained for chignolin. The results suggested that the SAAP-MC method is useful for conformational sampling for the short peptides. A protocol of SAAP-MC simulation followed by structural clustering and examination of the obtained structures by ab initio calculation or simply by the number of the hydrogen bonds (or the hardness was demonstrated to be an effective strategy toward structure prediction for short peptide molecules.

  12. Negative Ion In-Source Decay Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry for Sequencing Acidic Peptides

    Science.gov (United States)

    McMillen, Chelsea L.; Wright, Patience M.; Cassady, Carolyn J.

    2016-05-01

    Matrix-assisted laser desorption/ionization (MALDI) in-source decay was studied in the negative ion mode on deprotonated peptides to determine its usefulness for obtaining extensive sequence information for acidic peptides. Eight biological acidic peptides, ranging in size from 11 to 33 residues, were studied by negative ion mode ISD (nISD). The matrices 2,5-dihydroxybenzoic acid, 2-aminobenzoic acid, 2-aminobenzamide, 1,5-diaminonaphthalene, 5-amino-1-naphthol, 3-aminoquinoline, and 9-aminoacridine were used with each peptide. Optimal fragmentation was produced with 1,5-diaminonphthalene (DAN), and extensive sequence informative fragmentation was observed for every peptide except hirudin(54-65). Cleavage at the N-Cα bond of the peptide backbone, producing c' and z' ions, was dominant for all peptides. Cleavage of the N-Cα bond N-terminal to proline residues was not observed. The formation of c and z ions is also found in electron transfer dissociation (ETD), electron capture dissociation (ECD), and positive ion mode ISD, which are considered to be radical-driven techniques. Oxidized insulin chain A, which has four highly acidic oxidized cysteine residues, had less extensive fragmentation. This peptide also exhibited the only charged localized fragmentation, with more pronounced product ion formation adjacent to the highly acidic residues. In addition, spectra were obtained by positive ion mode ISD for each protonated peptide; more sequence informative fragmentation was observed via nISD for all peptides. Three of the peptides studied had no product ion formation in ISD, but extensive sequence informative fragmentation was found in their nISD spectra. The results of this study indicate that nISD can be used to readily obtain sequence information for acidic peptides.

  13. A short synthetic peptide fragment of human C2ORF40 has therapeutic potential in breast cancer

    Energy Technology Data Exchange (ETDEWEB)

    Lin, Chaoyang [Shandong Univ., Jinan (China); Zhang, Pengju [Shandong Univ., Jinan (China); Jiang, Anli [Shandong Univ., Jinan (China); Mao, Jian-Hua [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States); Wei, Guangwei [Shandong Univ. School of Medicine, Jinan (China)

    2017-03-30

    C2ORF40 encodes a secreted protein which is cleaved to generate soluble peptides by proteolytic processing and this process is believed to be necessary for C2ORF40 to exert cell type specific biological activity. Here, we reported a short mimic peptide of human C2ORF40 acts potential therapeutic efficacy in human cancer cells in vitro and in vivo. We synthesized a short peptide of human C2ORF40, named C2ORF40 mimic peptide fragment and assessed its biological function on cancer cell growth, migration and tumorigenesis. Cell growth assay showed that C2ORF40 mimic peptide fragment significantly suppressed cell proliferation of breast and lung cancer cells. Moreover, C2ORF40 mimic peptide fragment significantly inhibited the migration and invasion of breast cancer cells. Furthermore, we showed that this peptide suppressed tumorigenesis in breast tumor xenograft model. Cell cycle assay indicated that the C2ORF40 mimic peptide fragment suppressed the growth of tumor cells through inducing mitotic phase arrest. In conclusion, our results firstly suggested that this short synthetic peptide of human C2ORF40 may be a candidate tumor therapeutic agent.

  14. Modeling of the Ebola Virus Delta Peptide Reveals a Potential Lytic Sequence Motif

    Directory of Open Access Journals (Sweden)

    William R. Gallaher

    2015-01-01

    Full Text Available Filoviruses, such as Ebola and Marburg viruses, cause severe outbreaks of human infection, including the extensive epidemic of Ebola virus disease (EVD in West Africa in 2014. In the course of examining mutations in the glycoprotein gene associated with 2014 Ebola virus (EBOV sequences, a differential level of conservation was noted between the soluble form of glycoprotein (sGP and the full length glycoprotein (GP, which are both encoded by the GP gene via RNA editing. In the region of the proteins encoded after the RNA editing site sGP was more conserved than the overlapping region of GP when compared to a distant outlier species, Tai Forest ebolavirus. Half of the amino acids comprising the “delta peptide”, a 40 amino acid carboxy-terminal fragment of sGP, were identical between otherwise widely divergent species. A lysine-rich amphipathic peptide motif was noted at the carboxyl terminus of delta peptide with high structural relatedness to the cytolytic peptide of the non-structural protein 4 (NSP4 of rotavirus. EBOV delta peptide is a candidate viroporin, a cationic pore-forming peptide, and may contribute to EBOV pathogenesis.

  15. 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

  16. Dynamic stability of nano-fibers self-assembled from short amphiphilic A6D peptides.

    Science.gov (United States)

    Nikoofard, Narges; Maghsoodi, Fahimeh

    2018-04-07

    Self-assembly of A 6 D amphiphilic peptides in explicit water is studied by using coarse-grained molecular dynamics simulations. It is observed that the self-assembly of randomly distributed A 6 D peptides leads to the formation of a network of nano-fibers. Two other simulations with cylindrical nano-fibers as the initial configuration show the dynamic stability of the self-assembled nano-fibers. As a striking feature, notable fluctuations occur along the axes of the nano-fibers. Depending on the number of peptides per unit length of the nano-fiber, flat-shaped bulges or spiral shapes along the nano-fiber axis are observed at the fluctuations. Analysis of the particle distribution around the nano-fiber indicates that the hydrophobic core and the hydrophilic shell of the nano-structure are preserved in both simulations. The size of the deformations and their correlation times are different in the two simulations. This study gives new insights into the dynamics of the self-assembled nano-structures of short amphiphilic peptides.

  17. Dynamic stability of nano-fibers self-assembled from short amphiphilic A6D peptides

    Science.gov (United States)

    Nikoofard, Narges; Maghsoodi, Fahimeh

    2018-04-01

    Self-assembly of A6D amphiphilic peptides in explicit water is studied by using coarse-grained molecular dynamics simulations. It is observed that the self-assembly of randomly distributed A6D peptides leads to the formation of a network of nano-fibers. Two other simulations with cylindrical nano-fibers as the initial configuration show the dynamic stability of the self-assembled nano-fibers. As a striking feature, notable fluctuations occur along the axes of the nano-fibers. Depending on the number of peptides per unit length of the nano-fiber, flat-shaped bulges or spiral shapes along the nano-fiber axis are observed at the fluctuations. Analysis of the particle distribution around the nano-fiber indicates that the hydrophobic core and the hydrophilic shell of the nano-structure are preserved in both simulations. The size of the deformations and their correlation times are different in the two simulations. This study gives new insights into the dynamics of the self-assembled nano-structures of short amphiphilic peptides.

  18. PuLSE: Quality control and quantification of peptide sequences explored by phage display libraries.

    Science.gov (United States)

    Shave, Steven; Mann, Stefan; Koszela, Joanna; Kerr, Alastair; Auer, Manfred

    2018-01-01

    The design of highly diverse phage display libraries is based on assumption that DNA bases are incorporated at similar rates within the randomized sequence. As library complexity increases and expected copy numbers of unique sequences decrease, the exploration of library space becomes sparser and the presence of truly random sequences becomes critical. We present the program PuLSE (Phage Library Sequence Evaluation) as a tool for assessing randomness and therefore diversity of phage display libraries. PuLSE runs on a collection of sequence reads in the fastq file format and generates tables profiling the library in terms of unique DNA sequence counts and positions, translated peptide sequences, and normalized 'expected' occurrences from base to residue codon frequencies. The output allows at-a-glance quantitative quality control of a phage library in terms of sequence coverage both at the DNA base and translated protein residue level, which has been missing from toolsets and literature. The open source program PuLSE is available in two formats, a C++ source code package for compilation and integration into existing bioinformatics pipelines and precompiled binaries for ease of use.

  19. The two capsid proteins of maize rayado fino virus contain common peptide sequences.

    Science.gov (United States)

    Falk, B W; Tsai, J H

    1986-01-01

    Virions of maize rayado fino virus (MRFV) were purified and two major capsid proteins (ca. Mr 29,000 and 22,000) were resolved by SDS-PAGE. When the two major capsid proteins were isolated from gels and compared by one-dimensional peptide mapping after digestion with Staphylococcus aureus V-8 protease, indistinguishable peptide maps were obtained, suggesting that these two proteins contain common peptide sequences. Some preparations also showed minor protein components that were intermediate between the Mr 22,000 and Mr 29,000 capsid proteins. One of the minor proteins, ca. Mr 27,000, gave a peptide map indistinguishable from the major capsid proteins. In vitro ageing of partially purified preparations or virion treatment with proteolytic enzymes failed to show conversion of the Mr 29,000 protein to a Mr 22,000. Protease inhibitors added to the buffers used for virion purification did not affect the apparent 1:3 ratio of 29,000 to 22,000 proteins in the purified preparations.

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

    Directory of Open Access Journals (Sweden)

    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.

  1. A Novel MS-Cleavable Azo Cross-Linker for Peptide Structure Analysis by Free Radical Initiated Peptide Sequencing (FRIPS)

    Science.gov (United States)

    Iacobucci, Claudio; Hage, Christoph; Schäfer, Mathias; Sinz, Andrea

    2017-10-01

    The chemical cross-linking/mass spectrometry (MS) approach is a growing research field in structural proteomics that allows gaining insights into protein conformations. It relies on creating distance constraints between cross-linked amino acid side chains that can further be used to derive protein structures. Currently, the most urgent task for designing novel cross-linking principles is an unambiguous and automated assignment of the created cross-linked products. Here, we introduce the homobifunctional, amine-reactive, and water soluble cross-linker azobisimidoester (ABI) as a prototype of a novel class of cross-linkers. The ABI-linker possesses an innovative modular scaffold combining the benefits of collisional activation lability with open shell chemistry. This MS-cleavable cross-linker can be efficiently operated via free radical initiated peptide sequencing (FRIPS) in positive ionization mode. Our proof-of-principle study challenges the gas phase behavior of the ABI-linker for the three amino acids, lysine, leucine, and isoleucine, as well as the model peptide thymopentin. The isomeric amino acids leucine and isoleucine could be discriminated by their characteristic side chain fragments. Collisional activation experiments were conducted via positive electrospray ionization (ESI) on two Orbitrap mass spectrometers. The ABI-mediated formation of odd electron product ions in MS/MS and MS3 experiments was evaluated and compared with a previously described azo-based cross-linker. All cross-linked products were amenable to automated analysis by the MeroX software, underlining the future potential of the ABI-linker for structural proteomics studies. [Figure not available: see fulltext.

  2. 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.

  3. De novo assembly of human genomes with massively parallel short read sequencing

    DEFF Research Database (Denmark)

    Li, Ruiqiang; Zhu, Hongmei; Ruan, Jue

    2010-01-01

    genomes from short read sequences. We successfully assembled both the Asian and African human genome sequences, achieving an N50 contig size of 7.4 and 5.9 kilobases (kb) and scaffold of 446.3 and 61.9 kb, respectively. The development of this de novo short read assembly method creates new opportunities...... for building reference sequences and carrying out accurate analyses of unexplored genomes in a cost-effective way....

  4. Structural studies of polypeptides: Mechanism of immunoglobin catalysis and helix propagation in hybrid sequence, disulfide containing peptides

    Energy Technology Data Exchange (ETDEWEB)

    Storrs, Richard Wood [Univ. of California, Berkeley, CA (United States)

    1992-08-01

    Catalytic immunoglobin fragments were studied Nuclear Magnetic Resonance spectroscopy to identify amino acid residues responsible for the catalytic activity. Small, hybrid sequence peptides were analyzed for helix propagation following covalent initiation and for activity related to the protein from which the helical sequence was derived. Hydrolysis of p-nitrophenyl carbonates and esters by specific immunoglobins is thought to involve charge complementarity. The pK of the transition state analog P-nitrophenyl phosphate bound to the immunoglobin fragment was determined by 31P-NMR to verify the juxtaposition of a positively charged amino acid to the binding/catalytic site. Optical studies of immunoglobin mediated photoreversal of cis, syn cyclobutane thymine dimers implicated tryptophan as the photosensitizing chromophore. Research shows the chemical environment of a single tryptophan residue is altered upon binding of the thymine dimer. This tryptophan residue was localized to within 20 Å of the binding site through the use of a nitroxide paramagnetic species covalently attached to the thymine dimer. A hybrid sequence peptide was synthesized based on the bee venom peptide apamin in which the helical residues of apamin were replaced with those from the recognition helix of the bacteriophage 434 repressor protein. Oxidation of the disufide bonds occured uniformly in the proper 1-11, 3-15 orientation, stabilizing the 434 sequence in an α-helix. The glycine residue stopped helix propagation. Helix propagation in 2,2,2-trifluoroethanol mixtures was investigated in a second hybrid sequence peptide using the apamin-derived disulfide scaffold and the S-peptide sequence. The helix-stop signal previously observed was not observed in the NMR NOESY spectrum. Helical connectivities were seen throughout the S-peptide sequence. The apamin/S-peptide hybrid binded to the S-protein (residues 21-166 of ribonuclease A) and reconstituted enzymatic activity.

  5. Structural studies of polypeptides: Mechanism of immunoglobin catalysis and helix propagation in hybrid sequence, disulfide containing peptides

    Energy Technology Data Exchange (ETDEWEB)

    Storrs, R.W.

    1992-08-01

    Catalytic immunoglobin fragments were studied Nuclear Magnetic Resonance spectroscopy to identify amino acid residues responsible for the catalytic activity. Small, hybrid sequence peptides were analyzed for helix propagation following covalent initiation and for activity related to the protein from which the helical sequence was derived. Hydrolysis of p-nitrophenyl carbonates and esters by specific immunoglobins is thought to involve charge complementarity. The pK of the transition state analog P-nitrophenyl phosphate bound to the immunoglobin fragment was determined by [sup 31]P-NMR to verify the juxtaposition of a positively charged amino acid to the binding/catalytic site. Optical studies of immunoglobin mediated photoreversal of cis, syn cyclobutane thymine dimers implicated tryptophan as the photosensitizing chromophore. Research shows the chemical environment of a single tryptophan residue is altered upon binding of the thymine dimer. This tryptophan residue was localized to within 20 [Angstrom] of the binding site through the use of a nitroxide paramagnetic species covalently attached to the thymine dimer. A hybrid sequence peptide was synthesized based on the bee venom peptide apamin in which the helical residues of apamin were replaced with those from the recognition helix of the bacteriophage 434 repressor protein. Oxidation of the disufide bonds occured uniformly in the proper 1-11, 3-15 orientation, stabilizing the 434 sequence in an [alpha]-helix. The glycine residue stopped helix propagation. Helix propagation in 2,2,2-trifluoroethanol mixtures was investigated in a second hybrid sequence peptide using the apamin-derived disulfide scaffold and the S-peptide sequence. The helix-stop signal previously observed was not observed in the NMR NOESY spectrum. Helical connectivities were seen throughout the S-peptide sequence. The apamin/S-peptide hybrid binded to the S-protein (residues 21-166 of ribonuclease A) and reconstituted enzymatic activity.

  6. Interference by clustered regularly interspaced short palindromic repeat (CRISPR) RNA is governed by a seed sequence

    NARCIS (Netherlands)

    Semenova, E.V.; Jore, M.M.; Westra, E.R.; Oost, van der J.; Brouns, S.J.J.

    2011-01-01

    Prokaryotic clustered regularly interspaced short palindromic repeat (CRISPR)/Cas (CRISPR-associated sequences) systems provide adaptive immunity against viruses when a spacer sequence of small CRISPR RNA (crRNA) matches a protospacer sequence in the viral genome. Viruses that escape CRISPR/Cas

  7. Prediction of Xaa-Pro peptide bond conformation from sequence and chemical shifts

    Energy Technology Data Exchange (ETDEWEB)

    Shen Yang; Bax, Ad, E-mail: bax@nih.go [National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Laboratory of Chemical Physics (United States)

    2010-03-15

    We present a program, named Promega, to predict the Xaa-Pro peptide bond conformation on the basis of backbone chemical shifts and the amino acid sequence. Using a chemical shift database of proteins of known structure together with the PDB-extracted amino acid preference of cis Xaa-Pro peptide bonds, a cis/trans probability score is calculated from the backbone and {sup 13}C{sup {beta}} chemical shifts of the proline and its neighboring residues. For an arbitrary number of input chemical shifts, which may include Pro-{sup 13}C{sup {gamma}}, Promega calculates the statistical probability that a Xaa-Pro peptide bond is cis. Besides its potential as a validation tool, Promega is particularly useful for studies of larger proteins where Pro-{sup 13}C{sup {gamma}} assignments can be challenging, and for on-going efforts to determine protein structures exclusively on the basis of backbone and {sup 13}C{sup {beta}} chemical shifts.

  8. Crystal structure of importin-{alpha} complexed with a classic nuclear localization sequence obtained by oriented peptide library screening

    Energy Technology Data Exchange (ETDEWEB)

    Takeda, A.A.S.; Fontes, M.R.M. [UNESP, Universidade Estadual Paulista, Botucatu, SP (Brazil); Yang, S.N.Y. [University of Melbourne, Melbourne (Australia); Harris, J.M. [Queensland University of Technology, Brisbane (Australia); Jans, D.A. [Monash University, Clayton (Australia); Kobe, B. [University of Queensland, Brisbane, QU (Australia)

    2012-07-01

    Full text: Importin-{alpha} (Imp{alpha}) plays a role in the classical nuclear import pathway, binding to cargo proteins with activities in the nucleus. Different Imp{alpha} paralogs responsible for specific cargos can be found in a single organism. The cargos contain nuclear localization sequences (NLSs), which are characterized by one or two clusters of basic amino acids (monopartite and bipartite NLSs, respectively). In this work we present the crystal structure of Imp{alpha} from M. musculus (residues 70-529, lacking the auto inhibitory domain) bound to a NLS peptide (pepTM). The peptide corresponds to the optimal sequence obtained by an oriented peptide library experiment designed to probe the specificity of the major NLS binding site. The peptide library used five degenerate positions and identified the sequence KKKRR as the optimal sequence for binding to this site for mouse Imp{alpha} (70-529). The protein was obtained using an E. coli expression system and purified by affinity chromatography followed by an ion exchange chromatography. A single crystal of Imp{alpha} -pepTM complex was grown by the hanging drop method. The data were collected using the Synchrotron Radiation Source LNLS, Brazil and processed to 2.3. Molecular replacement techniques were used to determine the crystal structure. Electron density corresponding to the peptide was present in both major and minor binding sites The peptide is bound to Imp{alpha} similar as the simian virus 40 (SV40) large tumour (T)-antigen NLS. Binding assays confirmed that the peptide bound to Imp{alpha} with low nM affinities. This is the first time that structural information has been linked to an oriented peptide library screening approach for importin-{alpha}; the results will contribute to understanding of the sequence determinants of classical NLSs, and may help identify as yet unidentified classical NLSs in novel proteins. (author)

  9. ISRNA: an integrative online toolkit for short reads from high-throughput sequencing data.

    Science.gov (United States)

    Luo, Guan-Zheng; Yang, Wei; Ma, Ying-Ke; Wang, Xiu-Jie

    2014-02-01

    Integrative Short Reads NAvigator (ISRNA) is an online toolkit for analyzing high-throughput small RNA sequencing data. Besides the high-speed genome mapping function, ISRNA provides statistics for genomic location, length distribution and nucleotide composition bias analysis of sequence reads. Number of reads mapped to known microRNAs and other classes of short non-coding RNAs, coverage of short reads on genes, expression abundance of sequence reads as well as some other analysis functions are also supported. The versatile search functions enable users to select sequence reads according to their sub-sequences, expression abundance, genomic location, relationship to genes, etc. A specialized genome browser is integrated to visualize the genomic distribution of short reads. ISRNA also supports management and comparison among multiple datasets. ISRNA is implemented in Java/C++/Perl/MySQL and can be freely accessed at http://omicslab.genetics.ac.cn/ISRNA/.

  10. Inhibition of the myostatin/Smad signaling pathway by short decorin-derived peptides.

    Science.gov (United States)

    El Shafey, Nelly; Guesnon, Mickaël; Simon, Françoise; Deprez, Eric; Cosette, Jérémie; Stockholm, Daniel; Scherman, Daniel; Bigey, Pascal; Kichler, Antoine

    2016-02-15

    Myostatin, also known as growth differentiation factor 8, is a member of the transforming growth factor-beta superfamily that has been shown to play a key role in the regulation of the skeletal muscle mass. Indeed, while myostatin deletion or loss of function induces muscle hypertrophy, its overexpression or systemic administration causes muscle atrophy. Since myostatin blockade is effective in increasing skeletal muscle mass, myostatin inhibitors have been actively sought after. Decorin, a member of the small leucine-rich proteoglycan family is a metalloprotein that was previously shown to bind and inactivate myostatin in a zinc-dependent manner. Furthermore, the myostatin-binding site has been shown to be located in the decorin N-terminal domain. In the present study, we investigated the anti-myostatin activity of short and soluble fragments of decorin. Our results indicate that the murine decorin peptides DCN48-71 and 42-65 are sufficient for inactivating myostatin in vitro. Moreover, we show that the interaction of mDCN48-71 to myostatin is strictly zinc-dependent. Binding of myostatin to activin type II receptor results in the phosphorylation of Smad2/3. Addition of the decorin peptide 48-71 decreased in a dose-dependent manner the myostatin-induced phosphorylation of Smad2 demonstrating thereby that the peptide inhibits the activation of the Smad signaling pathway. Finally, we found that mDCN48-71 displays a specificity towards myostatin, since it does not inhibit other members of the transforming growth factor-beta family. Copyright © 2016 Elsevier Inc. All rights reserved.

  11. Car sequencing is NP-hard: a short proof

    OpenAIRE

    B Estellon; F Gardi

    2013-01-01

    In this note, a new proof is given that the car sequencing (CS) problem is NP-hard. Established from the Hamiltonian Path problem, the reduction is direct while closing some gaps remaining in the previous NP-hardness results. Since CS is studied in many operational research courses, this result and its proof are particularly interesting for teaching purposes.

  12. Short Note DNA sequences from the Little Brown Bustard Eupodotis ...

    African Journals Online (AJOL)

    Taxonomic classification of birds based exclusively on morphology and plumage traits has often been found to be inconsistent with true evolutionary history when tested with molecular phylogenies based on neutrally evolving markers. Here we present cytochrome-b gene sequences for the poorly known Little Brown ...

  13. Transition metal ions mediated tyrosine based short peptide amphiphile nanostructures inhibit bacterial growth.

    Science.gov (United States)

    Joshi, Khashti Ballabh; Singh, Ramesh; Mishra, Narendra Kumar; Kumar, Vikas; Vinayak, Vandana

    2018-05-17

    We report the design and synthesis of biocompatible small peptide based molecule for the controlled and targeted delivery of the encapsulated bioactive metal ions via transforming their internal nanostructures. Tyrosine based short peptide amphiphile (sPA) was synthesized which self-assembled into β-sheet like secondary structures. The self assembly of the designed sPA was modulated by using different bioactive transition metal ions which is confirmed by spectroscopic and microscopic techniques. These bioactive metal ions conjugated sPA hybrid structures are further used to develop antibacterial materials. It is due to the excellent antibacterial activity of zinc ions that the growth of clinically relevant bacteria such as E. Coli was inhibited in the presence of zinc-sPA conjugate. The bacterial test demonstrated that owing to high biocompatibility with bacterial cell, the designed sPA worked as metal ions delivery agent and therefore it can show great potential in locally addressing bacterial infections. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  14. Short-term administration of glucagon-like peptide-2. Effects on bone mineral density and markers of bone turnover in short-bowel patients with no colon

    DEFF Research Database (Denmark)

    Haderslev, K V; Jeppesen, P B; Hartmann, B

    2002-01-01

    Glucagon-like peptide 2 (GLP-2) is a newly discovered intestinotrophic hormone. We have recently reported that a 5-week GLP-2 treatment improved the intestinal absorptive capacity of short-bowel patients with no colon. Additionally, GLP-2 treatment was associated with changes in body composition ...

  15. Phosphorylation-induced conformational changes in short peptides probed by fluorescence resonance energy transfer in the 10A domain.

    Science.gov (United States)

    Sahoo, Harekrushna; Nau, Werner M

    2007-03-26

    Phosphorylation-induced conformational changes in short polypeptides were probed by a fluorescence resonance energy transfer (FRET) method by employing a short-distance FRET pair (R(0) approximately 10 A) based on tryptophan as natural donor and a 2,3-diazabicyclo[2.2.2]oct-2-ene-labeled asparagine (Dbo) as synthetic acceptor. Two substrates for kinases, LeuArgArgTrpSerLeuGly-Dbo (peptide I) and TrpLysArgThrLeuArgArg-Dbo (peptide II), were investigated, with serine and threonine, respectively, as phosphorylation sites. Steady-state and time-resolved fluorescence experiments in H(2)O revealed a decrease in FRET efficiency for peptide I and an increase for peptide II; this suggested that the effective distances between donor and acceptor increased and decreased, respectively. The same trends and similar absolute variations in effective donor-acceptor distances were observed in propylene glycol, a less polar and highly viscous solvent; this suggested that the variations are due to intrinsic structural preferences. Fitting of the time-resolved decay traces according to a distribution function model (Gaussian distribution) provided the mean donor-acceptor distances, which showed an increase upon phosphorylation for peptide I (from 9.7 to 10.5 A) and a decrease for peptide II (from 10.9 to 9.3 A) in H(2)O. The broadness (half-width) of the distributions, which provides a measure of the rigidity of the peptides, remained similar upon phosphorylation of peptide I (3.0 versus 3.1 A), but decreased for peptide II (from 3.1 to 0.73 A in H(2)O); this suggests a more compact, structured conformation upon phosphorylation of the latter peptide. The elongation of the peptide backbone (by ca. 0.7 A) for peptide I is attributed to an increase in steric demand upon phosphorylation, which favors an extended conformation. The contraction (by ca. 1.4 A) and structural rigidification of peptide II is attributed to attractive Coulombic interactions and hydrogen bonding between the

  16. Effects on in vitro and in vivo angiogenesis induced by small peptides carrying adhesion sequences.

    Science.gov (United States)

    Conconi, Maria Teresa; Ghezzo, Francesca; Dettin, Monica; Urbani, Luca; Grandi, Claudio; Guidolin, Diego; Nico, Beatrice; Di Bello, Carlo; Ribatti, Domenico; Parnigotto, Pier Paolo

    2010-07-01

    It is well known that tumor growth is strictly dependent on neo-vessel formation inside the tumor mass and that cell adhesion is required to allow EC proliferation and migration inside the tumor. In this work, we have evaluated the in vitro and in vivo effects on angiogenesis of some peptides, originally designed to promote cell adhesion on biomaterials, containing RGD motif mediating cell adhesion via integrin receptors [RGD, GRGDSPK, and (GRGDSP)(4)K] or the heparin-binding sequence of human vitronectin that interacts with HSPGs [HVP(351-359)]. Cell adhesion, proliferation, migration, and capillary-like tube formation in Matrigel were determined on HUVECs, whereas the effects on in vivo angiogenesis were evaluated using the CAM assay. (GRGDSP)(4)K linear sequence inhibited cell adhesion, decreased cell proliferation, migration and morphogenesis in Matrigel, and induced anti-angiogenic responses on CAM at higher degree than that determined after incubation with RGD or GRGDSPK. Moreover, it counteracted both in vitro and in vivo the pro-angiogenic effects induced by the Fibroblast growth factor (FGF-2). On the other hand, HVP was not able to affect cell adhesion and appeared less effective than (GRGDSP)(4)K. Our data indicate that the activity of RGD-containing peptides is related to their adhesive properties, and their effects are modulated by the number of cell adhesion motifs and the aminoacidic residues next to these sequences. The anti-angiogenic properties of (GRGDSP)(4)K seem to depend on its interaction with integrins, whereas the effects of HVP may be partially due to an impairment of HSPGs/FGF-2.

  17. Short peptide based nanotubes capable of effective curcumin delivery for treating drug resistant malaria.

    Science.gov (United States)

    Alam, Shadab; Panda, Jiban Jyoti; Mukherjee, Tapan Kumar; Chauhan, Virander Singh

    2016-04-05

    Curcumin (Ccm) has shown immense potential as an antimalarial agent; however its low solubility and less bioavailability attenuate the in vivo efficacy of this potent compound. In order to increase Ccm's bioavailability, a number of organic/inorganic polymer based nanoparticles have been investigated. However, most of the present day nano based delivery systems pose a conundrum with respect to their complex synthesis procedures, poor in vivo stability and toxicity issues. Peptides due to their high biocompatibility could act as excellent materials for the synthesis of nanoparticulate drug delivery systems. Here, we have investigated dehydrophenylalanine (ΔPhe) di-peptide based self-assembled nanoparticles for the efficient delivery of Ccm as an antimalarial agent. The self-assembly and curcumin loading capacity of different ΔPhe dipeptides, phenylalanine-α,β-dehydrophenylalanine (FΔF), arginine-α,β-dehydrophenylalanine (RΔF), valine-α,β-dehydrophenylalanine (VΔF) and methonine-α,β-dehydrophenylalanine (MΔF) were investigated for achieving enhanced and effective delivery of the compound for potential anti-malarial therapy. FΔF, RΔF, VΔF and MΔF peptides formed different types of nanoparticles like nanotubes and nanovesicles under similar assembling conditions. Out of these, F∆F nanotubes showed maximum curcumin loading capacity of almost 68 % W/W. Ccm loaded F∆F nanotubes (Ccm-F∆F) showed comparatively higher (IC50, 3.0 µM) inhibition of Plasmodium falciparum (Indo strain) as compared to free Ccm (IC50, 13 µM). Ccm-F∆F nano formulation further demonstrated higher inhibition of parasite growth in malaria infected mice as compared to free Ccm. The dipeptide nanoparticles were highly biocompatible and didn't show any toxic effect on mammalian cell lines and normal blood cells. This work provides a proof of principle of using highly biocompatible short peptide based nanoparticles for entrapment and in vivo delivery of Ccm leading to an

  18. A Combinational Strategy upon RNA Sequencing and Peptidomics Unravels a Set of Novel Toxin Peptides in Scorpion Mesobuthus martensii

    Directory of Open Access Journals (Sweden)

    Ning Luan

    2016-10-01

    Full Text Available Scorpion venom is deemed to contain many toxic peptides as an important source of natural compounds. Out of the two hundred proteins identified in Mesobuthus martensii (M. martensii, only a few peptide toxins have been found so far. Herein, a combinational approach based upon RNA sequencing and Liquid chromatography-mass spectrometry/mass spectrometry (LC MS/MS was employed to explore the venom peptides in M. martensii. A total of 153 proteins were identified from the scorpion venom, 26 previously known and 127 newly identified. Of the novel toxins, 97 proteins exhibited sequence similarities to known toxins, and 30 were never reported. Combining peptidomic and transcriptomic analyses, the peptide sequence of BmKKx1 was reannotated and four disulfide bridges were confirmed within it. In light of the comparison of conservation and variety of toxin amino acid sequences, highly conserved and variable regions were perceived in 24 toxins that were parts of two sodium channel and two potassium channel toxins families. Taking all of this evidences together, the peptidomic analysis on M. martensii indeed identified numerous novel scorpion peptides, expanded our knowledge towards the venom diversity, and afforded a set of pharmaceutical candidates.

  19. 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.

  20. Tools for analyzing genetic variants from sequencing data Case study: short tandem repeats

    OpenAIRE

    Gymrek, Melissa

    2016-01-01

    This was presented as a BitesizeBio Webinar entitled "Tools for analyzing genetic variants from sequencing data Case study: short tandem repeats"Accompanying scripts can be accessed on github:https://github.com/mgymrek/mgymrek-bitesizebio-webinar 

  1. Characterization of Campylobacter jejuni applying flaA short variable region sequencing, multilocus sequencing and Fourier transform infrared spectroscopy

    DEFF Research Database (Denmark)

    Josefsen, Mathilde Hartmann; Bonnichsen, Lise; Larsson, Jonas

    flaA short variable region sequencing and phenetic Fourier transform infrared (FTIR) spectroscopy was applied on a collection of 102 Campylobacter jejuni isolated from continuous sampling of organic, free range geese and chickens. FTIR has been shown to serve as a valuable tool in typing...

  2. Lanthanide-binding peptides with two pendant aminodiacetate arms: impact of the sequence on chelation.

    Science.gov (United States)

    Niedźwiecka, Agnieszka; Cisnetti, Federico; Lebrun, Colette; Gateau, Christelle; Delangle, Pascale

    2012-03-21

    Lanthanide complexes with a series of hexapeptides-incorporating two unnatural chelating amino acids with aminodiacetate groups, Ada(1) and Ada(2)-have been examined in terms of their speciation, structure, stability and luminescence properties. Whereas Ada(2) acts as a tridentate donor in all cases, Ada(1) may act as a tetradentate donor thanks to the coordination of the amide carbonyl function assisted by the formation of a six-membered chelate ring. The position of the Ada(1) residue in the sequence is demonstrated to be critical for the lanthanide complex speciation and structure. Ada(1) promotes the coordination of the backbone amide function to afford a highly dehydrated Ln complex and an S-shape structure of the peptide backbone, only when found in position 2.

  3. Effect of positively charged short peptides on stability of cubic phases of monoolein/dioleoylphosphatidic acid mixtures.

    Science.gov (United States)

    Masum, Shah Md; Li, Shu Jie; Awad, Tarek S; Yamazaki, Masahito

    2005-06-07

    To elucidate the stability and phase transition of cubic phases of biomembranes with infinite periodic minimal surface is indispensable from biological and physicochemical aspects. In this report, we investigated the effect of positively charged peptide-3K (LLKKK) and poly(L-lysine) on the phase stability of monoolein (MO) membranes containing negatively charged dioleoylphosphatidic acid (DOPA) (i.e., DOPA/MO membranes) using small-angle X-ray scattering. At first, the effect of peptide-3K on 10% DOPA/90% MO membrane in excess water, which is in the Q229 phase, was investigated. At 3.4 mM peptide-3K, a Q229 to Q230 phase transition occurred, and at >3.4 mM peptide-3K, the membrane was in the Q230 phase. Poly(L-lysine) (M(w) 1K-4K) also induced the Q230 phase, but peptide-2K (LLKK) could not induce it in the same membrane. We also investigated the effect of peptide-3K on the multilamellar vesicle (MLV) of 25% DOPA/75% MO membrane, which is in L(alpha) phase. In the absence of peptide, the spacing of MLV was very large (11.3 nm), but at > or = 8 mM peptide-3K, it greatly decreased to a constant value (5.2 nm), irrespective of the peptide concentration, indicating that peptide-3K and the membranes form an electrostatically stabilized aggregation with low water content. Poly(L-lysine) also decreased greatly the spacing of the 25% DOPA/75% MO MLV, indicating the formation of a similar aggregation. To compare the effects of peptide-3K and poly(L-lysine) with that of osmotic stress on stability of the cubic phase, we investigated the effect of poly(ethylene glycol) with molecular weight 7500 (PEG-6K) on the phase stability of 10% DOPA/90% MO membrane. With an increase in PEG-6K concentration, i.e., with an increase in osmotic stress, the most stable phase changed as follows; Q229 (Schwartz's P surface) --> Q224 (D) --> Q230 (G). On the basis of these results, we discuss the mechanism of the effects of the positively charged short peptides (peptide-3K) and poly

  4. Massively parallel amplicon sequencing reveals isotype-specific variability of antimicrobial peptide transcripts in Mytilus galloprovincialis.

    Directory of Open Access Journals (Sweden)

    Umberto Rosani

    Full Text Available BACKGROUND: Effective innate responses against potential pathogens are essential in the living world and possibly contributed to the evolutionary success of invertebrates. Taken together, antimicrobial peptide (AMP precursors of defensin, mytilin, myticin and mytimycin can represent about 40% of the hemocyte transcriptome in mussels injected with viral-like and bacterial preparations, and unique profiles of myticin C variants are expressed in single mussels. Based on amplicon pyrosequencing, we have ascertained and compared the natural and Vibrio-induced diversity of AMP transcripts in mussel hemocytes from three European regions. METHODOLOGY/PRINCIPAL FINDINGS: Hemolymph was collected from mussels farmed in the coastal regions of Palavas (France, Vigo (Spain and Venice (Italy. To represent the AMP families known in M. galloprovincialis, nine transcript sequences have been selected, amplified from hemocyte RNA and subjected to pyrosequencing. Hemolymph from farmed (offshore and wild (lagoon Venice mussels, both injected with 10(7 Vibrio cells, were similarly processed. Amplicon pyrosequencing emphasized the AMP transcript diversity, with Single Nucleotide Changes (SNC minimal for mytilin B/C and maximal for arthropod-like defensin and myticin C. Ratio of non-synonymous vs. synonymous changes also greatly differed between AMP isotypes. Overall, each amplicon revealed similar levels of nucleotidic variation across geographical regions, with two main sequence patterns confirmed for mytimycin and no substantial changes after immunostimulation. CONCLUSIONS/SIGNIFICANCE: Barcoding and bidirectional pyrosequencing allowed us to map and compare the transcript diversity of known mussel AMPs. Though most of the genuine cds variation was common to the analyzed samples we could estimate from 9 to 106 peptide variants in hemolymph pools representing 100 mussels, depending on the AMP isoform and sampling site. In this study, no prevailing SNC patterns related

  5. Identification of novel bacteriophage peptides using a combination of gene sequence LC-MS-MS analysis and BLASTP

    Science.gov (United States)

    Introduction: In an effort to characterize novel bacteriophage with lytic activity against pathogenic E.coli associated with foodborne illness, gene sequencing and mass spectrometry have been used to identify expressed peptides which differentiate isolated bacteriophage from other known phage. Here,...

  6. Investigating the microstructure of keratin extracted from wool: peptide sequence (MALDI-TOF/TOF) and protein conformation (FTIR)

    Science.gov (United States)

    Keratin was extracted from wool by reduction with 2-mercaptoethanol. It was isolated as intact keratin and characterized by its similar molecular weight, protein composition, and secondary structure to native keratin. Gel electrophoresis patterns and MALDI-TOF/TOF peptide sequences provided the ide...

  7. Human acid β-glucosidase: isolation and amino acid sequence of a peptide containing the catalytic site

    International Nuclear Information System (INIS)

    Dinur, T.; Osiecki, K.M.; Legler, G.; Gatt, S.; Desnick, R.J.; Grabowski, G.A.

    1986-01-01

    Human acid β-glucosidase (D-glucosyl-N-acylsphingosine glucohydrolase, EC 3.2.1.45) cleaves the glucosidic bonds of glucosylceramide and synthetic β-glucosides. The deficient activity of this hydrolase is the enzymatic defect in the subtypes and variants of Gaucher disease, the most prevalent lysosomal storage disease. To isolate and characterize the catalytic site of the normal enzyme, brominated 3 H-labeled conduritol B epoxide ( 3 H-Br-CBE), which inhibits the enzyme by binding covalently to this site, was used as an affinity label. Under optimal conditions 1 mol of 3 H-Br-CBE bound to 1 mol of pure enzyme protein, indicating the presence of a single catalytic site per enzyme subunit. After V 8 protease digestion of the 3 H-Br-CBE-labeled homogeneous enzyme, three radiolabeled peptides, designated peptide A, B, or C, were resolved by reverse-phase HPLC. The partial amino acid sequence (37 residues) of peptide A (M/sub r/, 5000) was determined. The sequence of this peptide, which contained the catalytic site, had exact homology to the sequence near the carboxyl terminus of the protein, as predicted from the nucleotide sequence of the full-length cDNA encoding acid β-glucosidase

  8. 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

  9. Modulation of intra- and inter-sheet interactions in short peptide self-assembly by acetonitrile in aqueous solution

    International Nuclear Information System (INIS)

    Deng Li; Zhao Yurong; Zhou Peng; Xu Hai; Wang Yanting

    2016-01-01

    Besides our previous experimental discovery (Zhao Y R, et al . 2015 Langmuir , 31, 12975) that acetonitrile (ACN) can tune the morphological features of nanostructures self-assembled by short peptides KIIIIK (KI4K) in aqueous solution, further experiments reported in this work demonstrate that ACN can also tune the mass of the self-assembled nanostructures. To understand the microscopic mechanism how ACN molecules interfere peptide self-assembly process, we conducted a series of molecular dynamics simulations on a monomer, a cross- β sheet structure, and a proto-fibril of KI4K in pure water, pure ACN, and ACN-water mixtures, respectively. The simulation results indicate that ACN enhances the intra-sheet interaction dominated by the hydrogen bonding (H-bonding) interactions between peptide backbones, but weakens the inter-sheet interaction dominated by the interactions between hydrophobic side chains. Through analyzing the correlations between different groups of solvent and peptides and the solvent behaviors around the proto-fibril, we have found that both the polar and nonpolar groups of ACN play significant roles in causing the opposite effects on intermolecular interactions among peptides. The weaker correlation of the polar group of ACN than water molecule with the peptide backbone enhances H-bonding interactions between peptides in the proto-fibril. The stronger correlation of the nonpolar group of ACN than water molecule with the peptide side chain leads to the accumulation of ACN molecules around the proto-fibril with their hydrophilic groups exposed to water, which in turn allows more water molecules close to the proto-fibril surface and weakens the inter-sheet interactions. The two opposite effects caused by ACN form a microscopic mechanism clearly explaining our experimental observations. (paper)

  10. Dissociation Behavior of a TEMPO-Active Ester Cross-Linker for Peptide Structure Analysis by Free Radical Initiated Peptide Sequencing (FRIPS) in Negative ESI-MS.

    Science.gov (United States)

    Hage, Christoph; Ihling, Christian H; Götze, Michael; Schäfer, Mathias; Sinz, Andrea

    2017-01-01

    We have synthesized a homobifunctional amine-reactive cross-linking reagent, containing a TEMPO (2,2,6,6-tetramethylpiperidine-1-oxy) and a benzyl group (Bz), termed TEMPO-Bz-linker, to derive three-dimensional structural information of proteins. The aim for designing this novel cross-linker was to facilitate the mass spectrometric analysis of cross-linked products by free radical initiated peptide sequencing (FRIPS). In an initial study, we had investigated the fragmentation behavior of TEMPO-Bz-derivatized peptides upon collision activation in (+)-electrospray ionization collision-induced dissociation tandem mass spectrometry (ESI-CID-MS/MS) experiments. In addition to the homolytic NO-C bond cleavage FRIPS pathway delivering the desired odd-electron product ions, an alternative heterolytic NO-C bond cleavage, resulting in even-electron product ions mechanism was found to be relevant. The latter fragmentation route clearly depends on the protonation of the TEMPO-Bz-moiety itself, which motivated us to conduct (-)-ESI-MS, CID-MS/MS, and MS 3 experiments of TEMPO-Bz-cross-linked peptides to further clarify the fragmentation behavior of TEMPO-Bz-peptide molecular ions. We show that the TEMPO-Bz-linker is highly beneficial for conducting FRIPS in negative ionization mode as the desired homolytic cleavage of the NO-C bond is the major fragmentation pathway. Based on characteristic fragments, the isomeric amino acids leucine and isoleucine could be discriminated. Interestingly, we observed pronounced amino acid side chain losses in cross-linked peptides if the cross-linked peptides contain a high number of acidic amino acids. Graphical Abstract ᅟ.

  11. Precursors of vertebrate peptide antibiotics dermaseptin b and adenoregulin have extensive sequence identities with precursors of opioid peptides dermorphin, dermenkephalin, and deltorphins.

    Science.gov (United States)

    Amiche, M; Ducancel, F; Mor, A; Boulain, J C; Menez, A; Nicolas, P

    1994-07-08

    The dermaseptins are a family of broad spectrum antimicrobial peptides, 27-34 amino acids long, involved in the defense of the naked skin of frogs against microbial invasion. They are the first vertebrate peptides to show lethal effects against the filamentous fungi responsible for severe opportunistic infections accompanying immunodeficiency syndrome and the use of immunosuppressive agents. A cDNA library was constructed from skin poly(A+) RNA of the arboreal frog Phyllomedusa bicolor and screened with an oligonucleotide probe complementary to the COOH terminus of dermaseptin b. Several clones contained a full-length DNA copy of a 443-nucleotide mRNA that encoded a 78-residue dermaseptin b precursor protein. The deduced precursor contained a putative signal sequence at the NH2 terminus, a 20-residue spacer sequence extremely rich (60%) in glutamic and aspartic acids, and a single copy of a dermaseptin b progenitor sequence at the COOH terminus. One clone contained a complete copy of adenoregulin, a 33-residue peptide reported to enhance the binding of agonists to the A1 adenosine receptor. The mRNAs encoding adenoregulin and dermaseptin b were very similar: 70 and 75% nucleotide identities between the 5'- and 3'-untranslated regions, respectively; 91% amino acid identity between the signal peptides; 82% identity between the acidic spacer sequences; and 38% identity between adenoregulin and dermaseptin b. Because adenoregulin and dermaseptin b have similar precursor designs and antimicrobial spectra, adenoregulin should be considered as a new member of the dermaseptin family and alternatively named dermaseptin b II. Preprodermaseptin b and preproadenoregulin have considerable sequence identities to the precursors encoding the opioid heptapeptides dermorphin, dermenkephalin, and deltorphins. This similarity extended into the 5'-untranslated regions of the mRNAs. These findings suggest that the genes encoding the four preproproteins are all members of the same family

  12. SRComp: short read sequence compression using burstsort and Elias omega coding.

    Directory of Open Access Journals (Sweden)

    Jeremy John Selva

    Full Text Available Next-generation sequencing (NGS technologies permit the rapid production of vast amounts of data at low cost. Economical data storage and transmission hence becomes an increasingly important challenge for NGS experiments. In this paper, we introduce a new non-reference based read sequence compression tool called SRComp. It works by first employing a fast string-sorting algorithm called burstsort to sort read sequences in lexicographical order and then Elias omega-based integer coding to encode the sorted read sequences. SRComp has been benchmarked on four large NGS datasets, where experimental results show that it can run 5-35 times faster than current state-of-the-art read sequence compression tools such as BEETL and SCALCE, while retaining comparable compression efficiency for large collections of short read sequences. SRComp is a read sequence compression tool that is particularly valuable in certain applications where compression time is of major concern.

  13. Homologies between the amino acid sequences of some vertebrate peptide hormones and peptides isolated from invertebrate sources.

    Science.gov (United States)

    De Loof, A; Schoofs, L

    1990-01-01

    1. The 4K-prothoracicotropic hormone (PTTH) or bombyxin and the melanization-reddish coloration hormone of the silkworm Bombyx mori resemble insulin and insulin-like growth factors. 2. The family of adipokinetic/red pigment concentrating hormones has some similarity with glucagon. 3. Members of the FMRFamide family are found in vertebrates as well as in invertebrates. 4. In Locusta, a molecule immunologically and biologically related to amphibian melanophore stimulating hormone has been partially characterized. 5. Enkephalins and enkephalin-related peptides occur in insects and other invertebrates. 6. Peptides belonging to the tachykinin family have been isolated from molluscan (Octopus) salivary glands and from insect nervous tissue (Locusta migratoria). 7. Invertebrate arginine-vasotocin homologs have been isolated from an insect (Locusta migratoria) and from a mollusc (Conus). 8. In Leucophaea, Locusta and Drosophila, peptides resembling those of the vertebrate gastrin/cholecystokinin family have been identified. 9. As the number of different neuro-/gut peptides with possible function(s) as hormone, neurotransmitter or neuromodulator is now estimated to be of the order of a few hundred, more similarities will probably show up in the near future.

  14. Hybridization Capture Using Short PCR Products Enriches Small Genomes by Capturing Flanking Sequences (CapFlank)

    DEFF Research Database (Denmark)

    Tsangaras, Kyriakos; Wales, Nathan; Sicheritz-Pontén, Thomas

    2014-01-01

    , a non-negligible fraction of the resulting sequence reads are not homologous to the bait. We demonstrate that during capture, the bait-hybridized library molecules add additional flanking library sequences iteratively, such that baits limited to targeting relatively short regions (e.g. few hundred...... nucleotides) can result in enrichment across entire mitochondrial and bacterial genomes. Our findings suggest that some of the off-target sequences derived in capture experiments are non-randomly enriched, and that CapFlank will facilitate targeted enrichment of large contiguous sequences with minimal prior...

  15. Characterizing novel endogenous retroviruses from genetic variation inferred from short sequence reads

    DEFF Research Database (Denmark)

    Mourier, Tobias; Mollerup, Sarah; Vinner, Lasse

    2015-01-01

    From Illumina sequencing of DNA from brain and liver tissue from the lion, Panthera leo, and tumor samples from the pike-perch, Sander lucioperca, we obtained two assembled sequence contigs with similarity to known retroviruses. Phylogenetic analyses suggest that the pike-perch retrovirus belongs...... to the epsilonretroviruses, and the lion retrovirus to the gammaretroviruses. To determine if these novel retroviral sequences originate from an endogenous retrovirus or from a recently integrated exogenous retrovirus, we assessed the genetic diversity of the parental sequences from which the short Illumina reads...

  16. Expanding the peptide beta-turn in alphagamma hybrid sequences: 12 atom hydrogen bonded helical and hairpin turns.

    Science.gov (United States)

    Chatterjee, Sunanda; Vasudev, Prema G; Raghothama, Srinivasarao; Ramakrishnan, Chandrasekharan; Shamala, Narayanaswamy; Balaram, Padmanabhan

    2009-04-29

    Hybrid peptide segments containing contiguous alpha and gamma amino acid residues can form C(12) hydrogen bonded turns which may be considered as backbone expanded analogues of C(10) (beta-turns) found in alphaalpha segments. Exploration of the regular hydrogen bonded conformations accessible for hybrid alphagamma sequences is facilitated by the use of a stereochemically constrained gamma amino acid residue gabapentin (1-aminomethylcyclohexaneacetic acid, Gpn), in which the two torsion angles about C(gamma)-C(beta) (theta(1)) and C(beta)-C(alpha) (theta(2)) are predominantly restricted to gauche conformations. The crystal structures of the octapeptides Boc-Gpn-Aib-Gpn-Aib-Gpn-Aib-Gpn-Aib-OMe (1) and Boc-Leu-Phe-Val-Aib-Gpn-Leu-Phe-Val-OMe (2) reveal two distinct conformations for the Aib-Gpn segment. Peptide 1 forms a continuous helix over the Aib(2)-Aib(6) segment, while the peptide 2 forms a beta-hairpin structure stabilized by four cross-strand hydrogen bonds with the Aib-Gpn segment forming a nonhelical C(12) turn. The robustness of the helix in peptide 1 in solution is demonstrated by NMR methods. Peptide 2 is conformationally fragile in solution with evidence of beta-hairpin conformations being obtained in methanol. Theoretical calculations permit delineation of the various C(12) hydrogen bonded structures which are energetically feasible in alphagamma and gammaalpha sequences.

  17. Multi-species sequence comparison reveals conservation of ghrelin gene-derived splice variants encoding a truncated ghrelin peptide.

    Science.gov (United States)

    Seim, Inge; Jeffery, Penny L; Thomas, Patrick B; Walpole, Carina M; Maugham, Michelle; Fung, Jenny N T; Yap, Pei-Yi; O'Keeffe, Angela J; Lai, John; Whiteside, Eliza J; Herington, Adrian C; Chopin, Lisa K

    2016-06-01

    The peptide hormone ghrelin is a potent orexigen produced predominantly in the stomach. It has a number of other biological actions, including roles in appetite stimulation, energy balance, the stimulation of growth hormone release and the regulation of cell proliferation. Recently, several ghrelin gene splice variants have been described. Here, we attempted to identify conserved alternative splicing of the ghrelin gene by cross-species sequence comparisons. We identified a novel human exon 2-deleted variant and provide preliminary evidence that this splice variant and in1-ghrelin encode a C-terminally truncated form of the ghrelin peptide, termed minighrelin. These variants are expressed in humans and mice, demonstrating conservation of alternative splicing spanning 90 million years. Minighrelin appears to have similar actions to full-length ghrelin, as treatment with exogenous minighrelin peptide stimulates appetite and feeding in mice. Forced expression of the exon 2-deleted preproghrelin variant mirrors the effect of the canonical preproghrelin, stimulating cell proliferation and migration in the PC3 prostate cancer cell line. This is the first study to characterise an exon 2-deleted preproghrelin variant and to demonstrate sequence conservation of ghrelin gene-derived splice variants that encode a truncated ghrelin peptide. This adds further impetus for studies into the alternative splicing of the ghrelin gene and the function of novel ghrelin peptides in vertebrates.

  18. Accurate typing of short tandem repeats from genome-wide sequencing data and its applications.

    Science.gov (United States)

    Fungtammasan, Arkarachai; Ananda, Guruprasad; Hile, Suzanne E; Su, Marcia Shu-Wei; Sun, Chen; Harris, Robert; Medvedev, Paul; Eckert, Kristin; Makova, Kateryna D

    2015-05-01

    Short tandem repeats (STRs) are implicated in dozens of human genetic diseases and contribute significantly to genome variation and instability. Yet profiling STRs from short-read sequencing data is challenging because of their high sequencing error rates. Here, we developed STR-FM, short tandem repeat profiling using flank-based mapping, a computational pipeline that can detect the full spectrum of STR alleles from short-read data, can adapt to emerging read-mapping algorithms, and can be applied to heterogeneous genetic samples (e.g., tumors, viruses, and genomes of organelles). We used STR-FM to study STR error rates and patterns in publicly available human and in-house generated ultradeep plasmid sequencing data sets. We discovered that STRs sequenced with a PCR-free protocol have up to ninefold fewer errors than those sequenced with a PCR-containing protocol. We constructed an error correction model for genotyping STRs that can distinguish heterozygous alleles containing STRs with consecutive repeat numbers. Applying our model and pipeline to Illumina sequencing data with 100-bp reads, we could confidently genotype several disease-related long trinucleotide STRs. Utilizing this pipeline, for the first time we determined the genome-wide STR germline mutation rate from a deeply sequenced human pedigree. Additionally, we built a tool that recommends minimal sequencing depth for accurate STR genotyping, depending on repeat length and sequencing read length. The required read depth increases with STR length and is lower for a PCR-free protocol. This suite of tools addresses the pressing challenges surrounding STR genotyping, and thus is of wide interest to researchers investigating disease-related STRs and STR evolution. © 2015 Fungtammasan et al.; Published by Cold Spring Harbor Laboratory Press.

  19. Large-scale analysis of peptide sequence variants: the case for high-field asymmetric waveform ion mobility spectrometry.

    Science.gov (United States)

    Creese, Andrew J; Smart, Jade; Cooper, Helen J

    2013-05-21

    Large scale analysis of proteins by mass spectrometry is becoming increasingly routine; however, the presence of peptide isomers remains a significant challenge for both identification and quantitation in proteomics. Classes of isomers include sequence inversions, structural isomers, and localization variants. In many cases, liquid chromatography is inadequate for separation of peptide isomers. The resulting tandem mass spectra are composite, containing fragments from multiple precursor ions. The benefits of high-field asymmetric waveform ion mobility spectrometry (FAIMS) for proteomics have been demonstrated by a number of groups, but previously work has focused on extending proteome coverage generally. Here, we present a systematic study of the benefits of FAIMS for a key challenge in proteomics, that of peptide isomers. We have applied FAIMS to the analysis of a phosphopeptide library comprising the sequences GPSGXVpSXAQLX(K/R) and SXPFKXpSPLXFG(K/R), where X = ADEFGLSTVY. The library has defined limits enabling us to make valid conclusions regarding FAIMS performance. The library contains numerous sequence inversions and structural isomers. In addition, there are large numbers of theoretical localization variants, allowing false localization rates to be determined. The FAIMS approach is compared with reversed-phase liquid chromatography and strong cation exchange chromatography. The FAIMS approach identified 35% of the peptide library, whereas LC-MS/MS alone identified 8% and LC-MS/MS with strong cation exchange chromatography prefractionation identified 17.3% of the library.

  20. Short-term effects of beta-amyloid25-35 peptide aggregates on transmitter release in neuromuscular synapses.

    Science.gov (United States)

    Garcia, Neus; Santafé, Manel M; Tomàs, Marta; Lanuza, Maria A; Tomàs, Josep

    2008-03-01

    The beta-amyloid (AB) peptide25-35 contains the functional domain of the AB precursor protein that is both required for neurotrophic effects in normal neural tissues and is involved in the neurotoxic effects in Alzheimer disease. We demonstrated the presence of the amyloid precursor protein/AB peptide in intramuscular axons, presynaptic motor nerve terminals, terminal and myelinating Schwann cells, and the postsynaptic and subsarcolemmal region in the Levator auris longus muscle of adult rats by immunocytochemistry. Using intracellular recording, we investigated possible short-term functional effects of the AB fragment (0.1-10 micromol/L) on acetylcholine release in adult and newborn motor end plates. We found no change in evoked, spontaneous transmitter release or resting membrane potential of the muscle cells. A previous block of the presynaptic muscarinic receptor subtypes and a previous block or stimulation of protein kinase C revealed no masked effect of the peptide on the regulation of transmitter release. The aggregated form of AB peptide25-35, however, interfered acutely with acetylcholine release (quantal content reduction) when synaptic activity was maintained by electric stimulation. The possible relevance of this inhibition of neurotransmission by AB peptide25-35 to the pathogenesis of Alzheimer remains to be determined.

  1. Ranked solutions to a class of combinatorial optimizations - with applications in mass spectrometry based peptide sequencing

    Science.gov (United States)

    Doerr, Timothy; Alves, Gelio; Yu, Yi-Kuo

    2006-03-01

    Typical combinatorial optimizations are NP-hard; however, for a particular class of cost functions the corresponding combinatorial optimizations can be solved in polynomial time. This suggests a way to efficiently find approximate solutions - - find a transformation that makes the cost function as similar as possible to that of the solvable class. After keeping many high-ranking solutions using the approximate cost function, one may then re-assess these solutions with the full cost function to find the best approximate solution. Under this approach, it is important to be able to assess the quality of the solutions obtained, e.g., by finding the true ranking of kth best approximate solution when all possible solutions are considered exhaustively. To tackle this statistical issue, we provide a systematic method starting with a scaling function generated from the fininte number of high- ranking solutions followed by a convergent iterative mapping. This method, useful in a variant of the directed paths in random media problem proposed here, can also provide a statistical significance assessment for one of the most important proteomic tasks - - peptide sequencing using tandem mass spectrometry data.

  2. Sequence-specific unusual (1-->2)-type helical turns in alpha/beta-hybrid peptides.

    Science.gov (United States)

    Prabhakaran, Panchami; Kale, Sangram S; Puranik, Vedavati G; Rajamohanan, P R; Chetina, Olga; Howard, Judith A K; Hofmann, Hans-Jörg; Sanjayan, Gangadhar J

    2008-12-31

    This article describes novel conformationally ordered alpha/beta-hybrid peptides consisting of repeating l-proline-anthranilic acid building blocks. These oligomers adopt a compact, right-handed helical architecture determined by the intrinsic conformational preferences of the individual amino acid residues. The striking feature of these oligomers is their ability to display an unusual periodic pseudo beta-turn network of nine-membered hydrogen-bonded rings formed in the forward direction of the sequence by 1-->2 amino acid interactions both in solid-state and in solution. Conformational investigations of several of these oligomers by single-crystal X-ray diffraction, solution-state NMR, and ab initio MO theory suggest that the characteristic steric and dihedral angle restraints exerted by proline are essential for stabilizing the unusual pseudo beta-turn network found in these oligomers. Replacing proline by the conformationally flexible analogue alanine (Ala) or by the conformationally more constrained alpha-amino isobutyric acid (Aib) had an adverse effect on the stabilization of this structural architecture. These findings increase the potential to design novel secondary structure elements profiting from the steric and dihedral angle constraints of the amino acid constituents and help to augment the conformational space available for synthetic oligomer design with diverse backbone structures.

  3. Systematic discovery of new recognition peptides mediating protein interaction networks

    DEFF Research Database (Denmark)

    Neduva, Victor; Linding, Rune; Su-Angrand, Isabelle

    2005-01-01

    Many aspects of cell signalling, trafficking, and targeting are governed by interactions between globular protein domains and short peptide segments. These domains often bind multiple peptides that share a common sequence pattern, or "linear motif" (e.g., SH3 binding to PxxP). Many domains...... by interactions between globular protein domains and short peptide segments. These domains often bind multiple peptides that share a common sequence pattern, or "linear motif" (e.g., SH3 binding to PxxP). Many domains are known, though comparatively few linear motifs have been discovered. Their short length...

  4. High-throughput sequencing enhanced phage display identifies peptides that bind mycobacteria

    CSIR Research Space (South Africa)

    Ngubane, NAC

    2013-11-01

    Full Text Available . The displayed peptides are flanked by two cysteine residues, which are oxidized during phage assembly to a disulfide bond, resulting in a loop constrained peptide. We initially used the traditional clone picking method to identify the enriched clones... of the library, 1.236109 heptapeptides, it represented sufficient depth to measure the quantitative enrich- ment of relevant peptides. To confirm successful enrichment during selection, we characterized the reduction in diversity of the pool in the consecutive...

  5. Neurospora tryptophan synthase: N-terminal analysis and the sequence of the pyridoxal phosphate active site peptide

    International Nuclear Information System (INIS)

    Pratt, M.L.; Hsu, P.Y.; DeMoss, J.A.

    1986-01-01

    Tryptophan synthase (TS), which catalyzes the final step of tryptophan biosynthesis, is a multifunctional protein requiring pyridoxal phosphate (B6P) for two of its three distinct enzyme activities. TS from Neurospora has a blocked N-terminal, is a homodimer of 150 KDa and binds one mole of B6P per mole of subunit. The authors shown the N-terminal residue to be acyl-serine. The B6P-active site of holoenzyme was labelled by reduction of the B6P-Schiff base with [ 3 H]-NaBH 4 , and resulted in a proportionate loss of activity in the two B6P-requiring reactions. SDS-polyacrylamide gel electrophoresis of CNBr-generated peptides showed the labelled, active site peptide to be 6 KDa. The sequence of this peptide, purified to apparent homogeneity by a combination of C-18 reversed phase and TSK gel filtration HPLC is: gly-arg-pro-gly-gln-leu-his-lys-ala-glu-arg-leu-thr-glu-tyr-ala-gly-gly-ala-gln-ile-xxx-leu-lys-arg-glu-asp-leu-asn-his-xxx-gly-xxx-his-/sub ***/-ile-asn-asn-ala-leu. Although four residues (xxx, /sub ***/) are unidentified, this peptide is minimally 78% homologous with the corresponding peptide from yeast TS, in which residue (/sub ***/) is the lysine that binds B6P

  6. TMS Over the Cerebellum Interferes with Short-term Memory of Visual Sequences.

    Science.gov (United States)

    Ferrari, C; Cattaneo, Z; Oldrati, V; Casiraghi, L; Castelli, F; D'Angelo, E; Vecchi, T

    2018-04-30

    Growing evidence suggests that the cerebellum is not only involved in motor functions, but it significantly contributes to sensory and cognitive processing as well. In particular, it has been hypothesized that the cerebellum identifies recurrent serial events and recognizes their violations. Here we used transcranial magnetic stimulation (TMS) to shed light on the role of the cerebellum in short-term memory of visual sequences. In two experiments, we found that TMS over the right cerebellar hemisphere impaired participants' ability to recognize the correct order of appearance of geometrical stimuli varying in shape and/or size. In turn, cerebellar TMS did not affect recognition of highly familiar short sequences of letters or numbers. Overall, our data suggest that the cerebellum is involved in memorizing the order in which (concatenated) stimuli appear, this process being important for sequence learning.

  7. Sequence-independent control of peptide conformation in liposomal vaccines for targeting protein misfolding diseases

    NARCIS (Netherlands)

    Hickman, D.T.; Nand, D.; Baldus, M.; Muhs, A.

    2011-01-01

    Synthetic peptide immunogens which mimic the conformation of a target epitope of pathological relevance offer the possibility to precisely control the immune response specificity. Here, we performed conformational analyses using a panel of peptides in order to investigate the key parameters

  8. A sensitive short read homology search tool for paired-end read sequencing data.

    Science.gov (United States)

    Techa-Angkoon, Prapaporn; Sun, Yanni; Lei, Jikai

    2017-10-16

    Homology search is still a significant step in functional analysis for genomic data. Profile Hidden Markov Model-based homology search has been widely used in protein domain analysis in many different species. In particular, with the fast accumulation of transcriptomic data of non-model species and metagenomic data, profile homology search is widely adopted in integrated pipelines for functional analysis. While the state-of-the-art tool HMMER has achieved high sensitivity and accuracy in domain annotation, the sensitivity of HMMER on short reads declines rapidly. The low sensitivity on short read homology search can lead to inaccurate domain composition and abundance computation. Our experimental results showed that half of the reads were missed by HMMER for a RNA-Seq dataset. Thus, there is a need for better methods to improve the homology search performance for short reads. We introduce a profile homology search tool named Short-Pair that is designed for short paired-end reads. By using an approximate Bayesian approach employing distribution of fragment lengths and alignment scores, Short-Pair can retrieve the missing end and determine true domains. In particular, Short-Pair increases the accuracy in aligning short reads that are part of remote homologs. We applied Short-Pair to a RNA-Seq dataset and a metagenomic dataset and quantified its sensitivity and accuracy on homology search. The experimental results show that Short-Pair can achieve better overall performance than the state-of-the-art methodology of profile homology search. Short-Pair is best used for next-generation sequencing (NGS) data that lack reference genomes. It provides a complementary paired-end read homology search tool to HMMER. The source code is freely available at https://sourceforge.net/projects/short-pair/ .

  9. Prediction of Antimicrobial Peptides Based on Sequence Alignment and Support Vector Machine-Pairwise Algorithm Utilizing LZ-Complexity

    Directory of Open Access Journals (Sweden)

    Xin Yi Ng

    2015-01-01

    Full Text Available This study concerns an attempt to establish a new method for predicting antimicrobial peptides (AMPs which are important to the immune system. Recently, researchers are interested in designing alternative drugs based on AMPs because they have found that a large number of bacterial strains have become resistant to available antibiotics. However, researchers have encountered obstacles in the AMPs designing process as experiments to extract AMPs from protein sequences are costly and require a long set-up time. Therefore, a computational tool for AMPs prediction is needed to resolve this problem. In this study, an integrated algorithm is newly introduced to predict AMPs by integrating sequence alignment and support vector machine- (SVM- LZ complexity pairwise algorithm. It was observed that, when all sequences in the training set are used, the sensitivity of the proposed algorithm is 95.28% in jackknife test and 87.59% in independent test, while the sensitivity obtained for jackknife test and independent test is 88.74% and 78.70%, respectively, when only the sequences that has less than 70% similarity are used. Applying the proposed algorithm may allow researchers to effectively predict AMPs from unknown protein peptide sequences with higher sensitivity.

  10. Short-peptide-based molecular hydrogels: novel gelation strategies and applications for tissue engineering and drug delivery

    Science.gov (United States)

    Wang, Huaimin; Yang, Zhimou

    2012-08-01

    Molecular hydrogels hold big potential for tissue engineering and controlled drug delivery. Our lab focuses on short-peptide-based molecular hydrogels formed by biocompatible methods and their applications in tissue engineering (especially, 3D cell culture) and controlled drug delivery. This feature article firstly describes our recent progresses of the development of novel methods to form hydrogels, including the strategy of disulfide bond reduction and assistance with specific protein-peptide interactions. We then introduce the applications of our hydrogels in fields of controlled stem cell differentiation, cell culture, surface modifications of polyester materials by molecular self-assembly, and anti-degradation of recombinant complex proteins. A novel molecular hydrogel system of hydrophobic compounds that are only formed by hydrolysis processes was also included in this article. The hydrogels of hydrophobic compounds, especially those of hydrophobic therapeutic agents, may be developed into a carrier-free delivery system for long term delivery of therapeutic agents. With the efforts in this field, we believe that molecular hydrogels formed by short peptides and hydrophobic therapeutic agents can be practically applied for 3D cell culture and long term drug delivery in near future, respectively.

  11. Structural Insights into the Mechanisms of Action of Short-Peptide HIV-1 Fusion Inhibitors Targeting the Gp41 Pocket

    Directory of Open Access Journals (Sweden)

    Xiujuan Zhang

    2018-02-01

    Full Text Available The deep hydrophobic pocket of HIV-1 gp41 has been considered a drug target, but short-peptides targeting this site usually lack potent antiviral activity. By applying the M-T hook structure, we previously generated highly potent short-peptide fusion inhibitors that specifically targeted the pocket site, such as MT-SC22EK, HP23L, and LP-11. Here, the crystal structures of HP23L and LP-11 bound to the target mimic peptide N36 demonstrated the critical intrahelical and interhelical interactions, especially verifying that the hook-like conformation was finely adopted while the methionine residue was replaced by the oxidation-less prone residue leucine, and that addition of an extra glutamic acid significantly enhanced the binding and inhibitory activities. The structure of HP23L bound to N36 with two mutations (E49K and L57R revealed the critical residues and motifs mediating drug resistance and provided new insights into the mechanism of action of inhibitors. Therefore, the present data help our understanding for the structure-activity relationship (SAR of HIV-1 fusion inhibitors and facilitate the development of novel antiviral drugs.

  12. Statistically significant dependence of the Xaa-Pro peptide bond conformation on secondary structure and amino acid sequence

    Directory of Open Access Journals (Sweden)

    Leitner Dietmar

    2005-04-01

    Full Text Available Abstract Background A reliable prediction of the Xaa-Pro peptide bond conformation would be a useful tool for many protein structure calculation methods. We have analyzed the Protein Data Bank and show that the combined use of sequential and structural information has a predictive value for the assessment of the cis versus trans peptide bond conformation of Xaa-Pro within proteins. For the analysis of the data sets different statistical methods such as the calculation of the Chou-Fasman parameters and occurrence matrices were used. Furthermore we analyzed the relationship between the relative solvent accessibility and the relative occurrence of prolines in the cis and in the trans conformation. Results One of the main results of the statistical investigations is the ranking of the secondary structure and sequence information with respect to the prediction of the Xaa-Pro peptide bond conformation. We observed a significant impact of secondary structure information on the occurrence of the Xaa-Pro peptide bond conformation, while the sequence information of amino acids neighboring proline is of little predictive value for the conformation of this bond. Conclusion In this work, we present an extensive analysis of the occurrence of the cis and trans proline conformation in proteins. Based on the data set, we derived patterns and rules for a possible prediction of the proline conformation. Upon adoption of the Chou-Fasman parameters, we are able to derive statistically relevant correlations between the secondary structure of amino acid fragments and the Xaa-Pro peptide bond conformation.

  13. Next-generation ELISA diagnostic assay for Chagas Disease based on the combination of short peptidic epitopes

    DEFF Research Database (Denmark)

    Mucci, Juan; Carmona, Santiago J.; Volcovich, Romina

    2017-01-01

    Chagas Disease, caused by the protozoan Trypanosoma cruzi, is a major health and economic problem in Latin America for which no vaccine or appropriate drugs for large-scale public health interventions are yet available. Accurate diagnosis is essential for the early identification and follow up....... cruzi linear B-cell epitopes using high-density peptide chips, leading to the identification of several hundred novel sequence signatures associated to chronic Chagas Disease. Here, we performed a serological assessment of 27 selected epitopes and of their use in a novel multipeptide-based diagnostic...... method. A combination of 7 of these peptides were finally evaluated in ELISA format against a panel of 199 sera samples (Chagas-positive and negative, including sera from Leishmaniasis-positive subjects). The multipeptide formulation displayed a high diagnostic performance, with a sensitivity of 96...

  14. Mechanism of action and in vitro activity of short hybrid antimicrobial peptide PV3 against Pseudomonas aeruginosa

    International Nuclear Information System (INIS)

    Memariani, Hamed; Shahbazzadeh, Delavar; Sabatier, Jean-Marc; Memariani, Mojtaba; Karbalaeimahdi, Ali; Bagheri, Kamran Pooshang

    2016-01-01

    Antimicrobial peptides are attractive candidates for developing novel therapeutic agents, since they are lethal to a broad spectrum of pathogens and have a unique low tendency for resistance development. In this study, mechanism of action and in vitro anti-pseudomonal activity of previously designed short hybrid antimicrobial peptide PV3 were investigated. Compared to ceftazidime, PV3 had not only higher antibacterial activity but also faster bactericidal activity. PV3 reduced biofilm biomass and viability of biofilm embedded bacteria in a concentration-dependent manner. Although the antimicrobial activity of PV3 was reduced in Mueller-Hinton broth (MHB) containing human serum, it was still active enough to eradication of bacteria at low concentrations. Compared with standard condition (MHB only), there was no significant decrease in antibacterial activity of PV3 against P. aeruginosa strains under 150 mM NaCl (p = 0.615) and 1 mM MgCl 2 (p = 0.3466). Fluorescence microscopy and field emission scanning electron microscopy further indicated that PV3 killed bacteria by disrupting the cell membrane. Since PV3 has potent anti-pseudomonal activity and has little cytotoxicity in vitro, it seems plausible that the peptide should be further investigated with animal studies to support future pharmacological formulations and potential topical applications. - Highlights: • PV3 killed Pseudomonas aeruginosa by membrane-disrupting mechanism. • PV3 reduced biofilm biomass and viability of biofilm embedded bacteria in a concentration-dependent manner. • Short hybrid antimicrobial peptide PV3 exhibited higher and faster bactericidal activity comparing to ceftazidime.

  15. Antimicrobial Activity of Truncated and Polyvalent Peptides Derived from the FKCRRQWQWRMKKGLA Sequence against Escherichia coli ATCC 25922 and Staphylococcus aureus ATCC 25923

    Directory of Open Access Journals (Sweden)

    Nataly de Jesús Huertas

    2017-06-01

    Full Text Available Peptides derived from LfcinB were designed and synthesized, and their antibacterial activity was tested against Escherichia coli ATCC 25922 and Staphylococcus aureus ATCC 25923. Specifically, a peptide library was constructed by systemically removing the flanking residues (N or C-terminal of Lfcin 17–31 (17FKCRRWQWRMKKLGA31, maintaining in all peptides the 20RRWQWR25 sequence that corresponds to the minimal antimicrobial motif. For this research, also included were (i a peptide containing an Ala instead of Cys ([Ala19]-LfcinB 17–31 and (ii polyvalent peptides containing the RRWQWR sequence and a non-natural amino acid (aminocaproic acid. We established that the lineal peptides LfcinB 17–25 and LfcinB 17–26 exhibited the greatest activity against E. coli ATCC 25922 and S. aureus ATCC 25923, respectively. On the other hand, polyvalent peptides, a dimer and a tetramer, exhibited the greatest antibacterial activity, indicating that multiple copies of the sequence increase the activity. Our results suggest that the dimeric and tetrameric sequence forms potentiate the antibacterial activity of lineal sequences that have exhibited moderate antibacterial activity.

  16. Sequence composition and gene content of the short arm of rye (Secale cereale chromosome 1.

    Directory of Open Access Journals (Sweden)

    Silvia Fluch

    Full Text Available BACKGROUND: The purpose of the study is to elucidate the sequence composition of the short arm of rye chromosome 1 (Secale cereale with special focus on its gene content, because this portion of the rye genome is an integrated part of several hundreds of bread wheat varieties worldwide. METHODOLOGY/PRINCIPAL FINDINGS: Multiple Displacement Amplification of 1RS DNA, obtained from flow sorted 1RS chromosomes, using 1RS ditelosomic wheat-rye addition line, and subsequent Roche 454FLX sequencing of this DNA yielded 195,313,589 bp sequence information. This quantity of sequence information resulted in 0.43× sequence coverage of the 1RS chromosome arm, permitting the identification of genes with estimated probability of 95%. A detailed analysis revealed that more than 5% of the 1RS sequence consisted of gene space, identifying at least 3,121 gene loci representing 1,882 different gene functions. Repetitive elements comprised about 72% of the 1RS sequence, Gypsy/Sabrina (13.3% being the most abundant. More than four thousand simple sequence repeat (SSR sites mostly located in gene related sequence reads were identified for possible marker development. The existence of chloroplast insertions in 1RS has been verified by identifying chimeric chloroplast-genomic sequence reads. Synteny analysis of 1RS to the full genomes of Oryza sativa and Brachypodium distachyon revealed that about half of the genes of 1RS correspond to the distal end of the short arm of rice chromosome 5 and the proximal region of the long arm of Brachypodium distachyon chromosome 2. Comparison of the gene content of 1RS to 1HS barley chromosome arm revealed high conservation of genes related to chromosome 5 of rice. CONCLUSIONS: The present study revealed the gene content and potential gene functions on this chromosome arm and demonstrated numerous sequence elements like SSRs and gene-related sequences, which can be utilised for future research as well as in breeding of wheat and rye.

  17. Folding Topology of a Short Coiled-Coil Peptide Structure Templated by an Oligonucleotide Triplex

    DEFF Research Database (Denmark)

    Lou, Chenguang; Christensen, Niels Johan; Martos Maldonado, Manuel Cristo

    2017-01-01

    by oligonucleotide duplex and triplex formation. POC synthesis was achieved by copper-free alkyne-azide cycloaddition between three oligonucleotides and a 23-mer peptide, which by itself exhibited multiple oligomeric states in solution. The oligonucleotide domain was designed to furnish a stable parallel triplex......, small-angle X-ray scattering (SAXS), and molecular modeling. Stabilizing cooperativity was observed between the trimeric peptide and the oligonucleotide triplex domains, and the overall molecular size (ca. 12nm) in solution was revealed to be independent of concentration. The topological folding...

  18. The genome of flax (Linum usitatissimum) assembled de novo from short shotgun sequence reads.

    Science.gov (United States)

    Wang, Zhiwen; Hobson, Neil; Galindo, Leonardo; Zhu, Shilin; Shi, Daihu; McDill, Joshua; Yang, Linfeng; Hawkins, Simon; Neutelings, Godfrey; Datla, Raju; Lambert, Georgina; Galbraith, David W; Grassa, Christopher J; Geraldes, Armando; Cronk, Quentin C; Cullis, Christopher; Dash, Prasanta K; Kumar, Polumetla A; Cloutier, Sylvie; Sharpe, Andrew G; Wong, Gane K-S; Wang, Jun; Deyholos, Michael K

    2012-11-01

    Flax (Linum usitatissimum) is an ancient crop that is widely cultivated as a source of fiber, oil and medicinally relevant compounds. To accelerate crop improvement, we performed whole-genome shotgun sequencing of the nuclear genome of flax. Seven paired-end libraries ranging in size from 300 bp to 10 kb were sequenced using an Illumina genome analyzer. A de novo assembly, comprised exclusively of deep-coverage (approximately 94× raw, approximately 69× filtered) short-sequence reads (44-100 bp), produced a set of scaffolds with N(50) =694 kb, including contigs with N(50)=20.1 kb. The contig assembly contained 302 Mb of non-redundant sequence representing an estimated 81% genome coverage. Up to 96% of published flax ESTs aligned to the whole-genome shotgun scaffolds. However, comparisons with independently sequenced BACs and fosmids showed some mis-assembly of regions at the genome scale. A total of 43384 protein-coding genes were predicted in the whole-genome shotgun assembly, and up to 93% of published flax ESTs, and 86% of A. thaliana genes aligned to these predicted genes, indicating excellent coverage and accuracy at the gene level. Analysis of the synonymous substitution rates (K(s) ) observed within duplicate gene pairs was consistent with a recent (5-9 MYA) whole-genome duplication in flax. Within the predicted proteome, we observed enrichment of many conserved domains (Pfam-A) that may contribute to the unique properties of this crop, including agglutinin proteins. Together these results show that de novo assembly, based solely on whole-genome shotgun short-sequence reads, is an efficient means of obtaining nearly complete genome sequence information for some plant species. © 2012 The Authors. The Plant Journal © 2012 Blackwell Publishing Ltd.

  19. Correction of echo shift in reconstruction processing for ultra-short TE pulse sequence

    International Nuclear Information System (INIS)

    Takizawa, Masahiro; Ootsuka, Takehiro; Abe, Takayuki; Takahashi, Tetsuhiko

    2010-01-01

    An ultra-short echo time (TE) pulse sequence is composed of a radial sampling that acquires echo signals radially in the K-space and a half-echo acquisition that acquires only half of the echo signal. The shift in the position of the echo signal (echo shift) caused by the timing errors in the gradient magnetic field pulses affects the image quality in the radial sampling with the half-echo acquisition. To improve image quality, we have developed a signal correction algorithm that detects and eliminates this echo shift during reconstruction by performing a pre-scan within 10 seconds. The results showed that image quality is improved under oblique and/or off-centering conditions that frequently cause image distortion due to hardware error. In conclusion, we have developed a robust ultra-short TE pulse sequence that allows wide latitude in the scan parameters, including oblique and off-centering conditions. (author)

  20. SeqEntropy: genome-wide assessment of repeats for short read sequencing.

    Directory of Open Access Journals (Sweden)

    Hsueh-Ting Chu

    Full Text Available BACKGROUND: Recent studies on genome assembly from short-read sequencing data reported the limitation of this technology to reconstruct the entire genome even at very high depth coverage. We investigated the limitation from the perspective of information theory to evaluate the effect of repeats on short-read genome assembly using idealized (error-free reads at different lengths. METHODOLOGY/PRINCIPAL FINDINGS: We define a metric H(k to be the entropy of sequencing reads at a read length k and use the relative loss of entropy ΔH(k to measure the impact of repeats for the reconstruction of whole-genome from sequences of length k. In our experiments, we found that entropy loss correlates well with de-novo assembly coverage of a genome, and a score of ΔH(k>1% indicates a severe loss in genome reconstruction fidelity. The minimal read lengths to achieve ΔH(k<1% are different for various organisms and are independent of the genome size. For example, in order to meet the threshold of ΔH(k<1%, a read length of 60 bp is needed for the sequencing of human genome (3.2 10(9 bp and 320 bp for the sequencing of fruit fly (1.8×10(8 bp. We also calculated the ΔH(k scores for 2725 prokaryotic chromosomes and plasmids at several read lengths. Our results indicate that the levels of repeats in different genomes are diverse and the entropy of sequencing reads provides a measurement for the repeat structures. CONCLUSIONS/SIGNIFICANCE: The proposed entropy-based measurement, which can be calculated in seconds to minutes in most cases, provides a rapid quantitative evaluation on the limitation of idealized short-read genome sequencing. Moreover, the calculation can be parallelized to scale up to large euakryotic genomes. This approach may be useful to tune the sequencing parameters to achieve better genome assemblies when a closely related genome is already available.

  1. Detection of discriminative sequence patterns in the neighborhood of proline cis peptide bonds and their functional annotation

    Directory of Open Access Journals (Sweden)

    Papaloukas Costas

    2009-04-01

    Full Text Available Abstract Background Polypeptides are composed of amino acids covalently bonded via a peptide bond. The majority of peptide bonds in proteins is found to occur in the trans conformation. In spite of their infrequent occurrence, cis peptide bonds play a key role in the protein structure and function, as well as in many significant biological processes. Results We perform a systematic analysis of regions in protein sequences that contain a proline cis peptide bond in order to discover non-random associations between the primary sequence and the nature of proline cis/trans isomerization. For this purpose an efficient pattern discovery algorithm is employed which discovers regular expression-type patterns that are overrepresented (i.e. appear frequently repeated in a set of sequences. Four types of pattern discovery are performed: i exact pattern discovery, ii pattern discovery using a chemical equivalency set, iii pattern discovery using a structural equivalency set and iv pattern discovery using certain amino acids' physicochemical properties. The extracted patterns are carefully validated using a specially implemented scoring function and a significance measure (i.e. log-probability estimate indicative of their specificity. The score threshold for the first three types of pattern discovery is 0.90 while for the last type of pattern discovery 0.80. Regarding the significance measure, all patterns yielded values in the range [-9, -31] which ensure that the derived patterns are highly unlikely to have emerged by chance. Among the highest scoring patterns, most of them are consistent with previous investigations concerning the neighborhood of cis proline peptide bonds, and many new ones are identified. Finally, the extracted patterns are systematically compared against the PROSITE database, in order to gain insight into the functional implications of cis prolyl bonds. Conclusion Cis patterns with matches in the PROSITE database fell mostly into two

  2. Design of Tail-Clamp Peptide Nucleic Acid Tethered with Azobenzene Linker for Sequence-Specific Detection of Homopurine DNA

    Directory of Open Access Journals (Sweden)

    Shinjiro Sawada

    2017-10-01

    Full Text Available DNA carries genetic information in its sequence of bases. Synthetic oligonucleotides that can sequence-specifically recognize a target gene sequence are a useful tool for regulating gene expression or detecting target genes. Among the many synthetic oligonucleotides, tail-clamp peptide nucleic acid (TC-PNA offers advantages since it has two homopyrimidine PNA strands connected via a flexible ethylene glycol-type linker that can recognize complementary homopurine sequences via Watson-Crick and Hoogsteen base pairings and form thermally-stable PNA/PNA/DNA triplex structures. Here, we synthesized a series of TC-PNAs that can possess different lengths of azobenzene-containing linkers and studied their binding behaviours to homopurine single-stranded DNA. Introduction of azobenzene at the N-terminus amine of PNA increased the thermal stability of PNA-DNA duplexes. Further extension of the homopyrimidine PNA strand at the N-terminus of PNA-AZO further increased the binding stability of the PNA/DNA/PNA triplex to the target homopurine sequence; however, it induced TC-PNA/DNA/TC-PNA complex formation. Among these TC-PNAs, 9W5H-C4-AZO consisting of nine Watson-Crick bases and five Hoogsteen bases tethered with a beta-alanine conjugated azobenzene linker gave a stable 1:1 TC-PNA/ssDNA complex and exhibited good mismatch recognition. Our design for TC-PNA-AZO can be utilized for detecting homopurine sequences in various genes.

  3. Acute effects of the Glucagon-Like Peptide 2 analogue, teduglutide, on intestinal adaptation in newborn pigs with short bowel syndrome

    DEFF Research Database (Denmark)

    Thymann, Thomas; Stoll, Barbara; Mecklenburg, Lars

    2014-01-01

    Neonatal short bowel syndrome following massive gut resection associates with malabsorption of nutrients. The intestinotrophic factor glucagon-like peptide 2 (GLP-2) improves gut function in adult short bowel patients, but its effect in pediatric patients remains unknown. Our objective was to test...

  4. 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.

  5. How proteases from Enterococcus faecalis contribute to its resistance to short alpha-helical antimicrobial peptides

    Czech Academy of Sciences Publication Activity Database

    Nešuta, Ondřej; Buděšínský, Miloš; Hadravová, Romana; Monincová, Lenka; Humpolíčková, Jana; Čeřovský, Václav

    2017-01-01

    Roč. 75, č. 7 (2017), č. článku ftx091. ISSN 2049-632X R&D Projects: GA TA ČR(CZ) TA04010638; GA MZd(CZ) NV16-27726A Institutional support: RVO:61388963 Keywords : antimicrobial peptides * biofilm * C-terminal deamidation * gelatinase * protease Subject RIV: EE - Microbiology, Virology OBOR OECD: Microbiology Impact factor: 2.335, year: 2016

  6. Potential clinical usefulness of new glandular and circulating parathyroid peptides illuminated by sequence specific radioimmunoassay

    International Nuclear Information System (INIS)

    Lindall, A.W.; Cecchettin, M.

    1981-01-01

    It is now well known that human PTH peptides constitute a heterogeneous population of fragments of the eighty-four amino acid molecules. Over the years, considerable confusion has resulted from measurements made in both clinical and experimental settings with radioimmunoassays of either undefined or partially-defined specificity for a particular region of the intact molecule. The approach of the authors has been to make use of modern technology to synthesize peptide fragments that mimic portions of native molecules, and to use these fragments in RIA. A more detailed description of a portion of this work will appear in the Journal of Clinical Endocrinology and Metabolism. (Auth.)

  7. Structure-Related Roles for the Conservation of the HIV-1 Fusion Peptide Sequence Revealed by Nuclear Magnetic Resonance.

    Science.gov (United States)

    Serrano, Soraya; Huarte, Nerea; Rujas, Edurne; Andreu, David; Nieva, José L; Jiménez, María Angeles

    2017-10-17

    Despite extensive characterization of the human immunodeficiency virus type 1 (HIV-1) hydrophobic fusion peptide (FP), the structure-function relationships underlying its extraordinary degree of conservation remain poorly understood. Specifically, the fact that the tandem repeat of the FLGFLG tripeptide is absolutely conserved suggests that high hydrophobicity may not suffice to unleash FP function. Here, we have compared the nuclear magnetic resonance (NMR) structures adopted in nonpolar media by two FP surrogates, wtFP-tag and scrFP-tag, which had equal hydrophobicity but contained wild-type and scrambled core sequences LFLGFLG and FGLLGFL, respectively. In addition, these peptides were tagged at their C-termini with an epitope sequence that folded independently, thereby allowing Western blot detection without interfering with FP structure. We observed similar α-helical FP conformations for both specimens dissolved in the low-polarity medium 25% (v/v) 1,1,1,3,3,3-hexafluoro-2-propanol (HFIP), but important differences in contact with micelles of the membrane mimetic dodecylphosphocholine (DPC). Thus, whereas wtFP-tag preserved a helix displaying a Gly-rich ridge, the scrambled sequence lost in great part the helical structure upon being solubilized in DPC. Western blot analyses further revealed the capacity of wtFP-tag to assemble trimers in membranes, whereas membrane oligomers were not observed in the case of the scrFP-tag sequence. We conclude that, beyond hydrophobicity, preserving sequence order is an important feature for defining the secondary structures and oligomeric states adopted by the HIV FP in membranes.

  8. Screening of SHOX gene sequence variants in Saudi Arabian children with idiopathic short stature.

    Science.gov (United States)

    Alharthi, Abdulla A; El-Hallous, Ehab I; Talaat, Iman M; Alghamdi, Hamed A; Almalki, Matar I; Gaber, Ahmed

    2017-10-01

    Short stature affects approximately 2%-3% of children, representing one of the most frequent disorders for which clinical attention is sought during childhood. Despite assumed genetic heterogeneity, mutations or deletions in the short stature homeobox-containing gene ( SHOX ) are frequently detected in subjects with short stature. Idiopathic short stature (ISS) refers to patients with short stature for various unknown reasons. The goal of this study was to screen all the exons of SHOX to identify related mutations. We screened all the exons of SHOX for mutations analysis in 105 ISS children patients (57 girls and 48 boys) living in Taif governorate, KSA using a direct DNA sequencing method. Height, arm span, and sitting height were recorded, and subischial leg length was calculated. A total of 30 of 105 ISS patients (28%) contained six polymorphic variants in exons 1, 2, 4, and 6. One mutation was found in the DNA domain binding region of exon 4. Three of these polymorphic variants were novel, while the others were reported previously. There were no significant differences in anthropometric measures in ISS patients with and without identifiable polymorphic variants in SHOX . In Saudi Arabia ISS patients, rather than SHOX , it is possible that new genes are involved in longitudinal growth. Additional molecular analysis is required to diagnose and understand the etiology of this disease.

  9. Technetium-99m somatostatin analogues: effect of labelling methods and peptide sequence

    International Nuclear Information System (INIS)

    Decristoforo, C.; Mather, S.J.

    1999-01-01

    In this paper the preclinical evaluation of the somatostatin analogue RC160 labelled with technetium-99m using bifunctional chelators (BFCs) based on the hydrazinonicotinamide (HYNIC) and N 3 S system is described and a comparison made with [Tyr 3 ]-octreotide (TOC). Conjugates of both peptides with HYNIC, and of RC160 with benzoyl-MAG 3 and an N 3 S-adipate derivative were prepared and radiolabelling performed at high specific activities using tricine, tricine/nicotinic acid and ethylenediamine-N,N'-diacetic adic (EDDA) as co-ligands for HYNIC conjugates. All conjugates and 99m Tc-labelled peptides showed preserved binding affinity for the somatostatin receptor (IC50, Kd 99m Tc-RC160 derivatives compared with 99m Tc-EDDA/HYNIC-[Tyr 3 ]-octreotide (0.2%-3.5%ID/g vs 9.7%ID/g) and correlated well with the reduced internalisation rate for RC160 derivatives. Our results show that the selection of the labelling approach as well as the right choice of the peptide structure are crucial for labelling peptides with 99m Tc to achieve complexes with favourable biodistribution. Despite the relatively low tumour uptake compared with 99m Tc-EDDA/HYNIC-[Tyr 3 ]-octreotide, 99m Tc-RC160 could play a role in imaging tumours that do not bind octreotide derivatives. (orig.)

  10. Membrane fusion is induced by a distinct peptide sequence of the sea urchin fertilization protein bindin

    NARCIS (Netherlands)

    Ulrich, AS; Glabe, CG; Hoekstra, D

    1998-01-01

    Fertilization in the sea urchin is mediated by the membrane-associated acrosomal protein bindin, which plays a key role in the adhesion and fusion between sperm and egg. We have investigated the structure/function relationship of an 18-amino acid peptide fragment "B18," which represents the minimal

  11. Easy and accurate reconstruction of whole HIV genomes from short-read sequence data with shiver

    Science.gov (United States)

    Blanquart, François; Golubchik, Tanya; Gall, Astrid; Bakker, Margreet; Bezemer, Daniela; Croucher, Nicholas J; Hall, Matthew; Hillebregt, Mariska; Ratmann, Oliver; Albert, Jan; Bannert, Norbert; Fellay, Jacques; Fransen, Katrien; Gourlay, Annabelle; Grabowski, M Kate; Gunsenheimer-Bartmeyer, Barbara; Günthard, Huldrych F; Kivelä, Pia; Kouyos, Roger; Laeyendecker, Oliver; Liitsola, Kirsi; Meyer, Laurence; Porter, Kholoud; Ristola, Matti; van Sighem, Ard; Cornelissen, Marion; Kellam, Paul; Reiss, Peter

    2018-01-01

    Abstract Studying the evolution of viruses and their molecular epidemiology relies on accurate viral sequence data, so that small differences between similar viruses can be meaningfully interpreted. Despite its higher throughput and more detailed minority variant data, next-generation sequencing has yet to be widely adopted for HIV. The difficulty of accurately reconstructing the consensus sequence of a quasispecies from reads (short fragments of DNA) in the presence of large between- and within-host diversity, including frequent indels, may have presented a barrier. In particular, mapping (aligning) reads to a reference sequence leads to biased loss of information; this bias can distort epidemiological and evolutionary conclusions. De novo assembly avoids this bias by aligning the reads to themselves, producing a set of sequences called contigs. However contigs provide only a partial summary of the reads, misassembly may result in their having an incorrect structure, and no information is available at parts of the genome where contigs could not be assembled. To address these problems we developed the tool shiver to pre-process reads for quality and contamination, then map them to a reference tailored to the sample using corrected contigs supplemented with the user’s choice of existing reference sequences. Run with two commands per sample, it can easily be used for large heterogeneous data sets. We used shiver to reconstruct the consensus sequence and minority variant information from paired-end short-read whole-genome data produced with the Illumina platform, for sixty-five existing publicly available samples and fifty new samples. We show the systematic superiority of mapping to shiver’s constructed reference compared with mapping the same reads to the closest of 3,249 real references: median values of 13 bases called differently and more accurately, 0 bases called differently and less accurately, and 205 bases of missing sequence recovered. We also

  12. Serodiagnosis of Leishmania donovani infections: assessment of enzyme-linked immunosorbent assays using recombinant L. donovani gene B protein (GBP) and a peptide sequence of L. donovani GBP

    DEFF Research Database (Denmark)

    Jensen, A T; Gasim, S; Moller, T

    1999-01-01

    The repetitive sequence of Leishmania major gene B protein (GBP) has previously been shown to be a useful tool in the diagnosis of cutaneous leishmaniasis (CL). Here, we have assessed enzyme-linked immunosorbent assays (ELISAs) using recombinant L. donovani GBP (rGBP) and a peptide sequence of L...... for malaria but free of leishmaniasis was negative in both assays....

  13. Syntheses and structures of technetium(V) and rhenium(V) oxo complexes of peptide having KYC-sequence

    International Nuclear Information System (INIS)

    Takayama, T.; Suzuki, K.; Sekine, T.; Kudo, H.

    2000-01-01

    Technetium(V) and rhenium(V) oxo complexes of a peptide having a KYC-sequence such as KYCAR (H 3 L 5 ) and KYCAREPPTRTNAYQGQG-NH 2 (H 3 L 18 ) were synthesized, and structures of the complexes were characterized by spectroscopic techniques. All of the complexes were synthesized by the ligand exchange reaction of [(n-C 4 H 9 ) 4 N][MOCl 4 ] (M = 99 Tc, Re) with peptide in methanol or dimethylformamide solution. These complexes have a square pyramidal structure with an oxo ligand at the apical position. The peptide is coordinated to a metal atom through N amine of lysine. S thiol of cysteine, and N amide of tyrosine and cysteine in the equatorial plane. A lysine (CH 2 ) 4 NH 2 group of the L 5 ligand has the syn conformation with respect to metal-oxo bonding in the complex. The syn isomer was selectively formed in the ligand exchange reaction. The conversion of the syn isomer to the anti isomer was observed only for syn-[ReO(L 5 )], in which the coordination of water to the trans position of the oxo ligand was involved. (orig.)

  14. Syntheses and structures of technetium(V) and rhenium(V) oxo complexes of peptide having KYC-sequence

    Energy Technology Data Exchange (ETDEWEB)

    Takayama, T.; Suzuki, K.; Sekine, T.; Kudo, H. [Dept. of Chemistry, Tohoku Univ., Sendai (Japan)

    2000-07-01

    Technetium(V) and rhenium(V) oxo complexes of a peptide having a KYC-sequence such as KYCAR (H{sub 3}L{sup 5}) and KYCAREPPTRTNAYQGQG-NH{sub 2} (H{sub 3}L{sup 18}) were synthesized, and structures of the complexes were characterized by spectroscopic techniques. All of the complexes were synthesized by the ligand exchange reaction of [(n-C{sub 4}H{sub 9}){sub 4}N][MOCl{sub 4}] (M = {sup 99}Tc, Re) with peptide in methanol or dimethylformamide solution. These complexes have a square pyramidal structure with an oxo ligand at the apical position. The peptide is coordinated to a metal atom through N{sub amine} of lysine. S{sub thiol} of cysteine, and N{sub amide} of tyrosine and cysteine in the equatorial plane. A lysine (CH{sub 2}){sub 4}NH{sub 2} group of the L{sup 5} ligand has the syn conformation with respect to metal-oxo bonding in the complex. The syn isomer was selectively formed in the ligand exchange reaction. The conversion of the syn isomer to the anti isomer was observed only for syn-[ReO(L{sup 5})], in which the coordination of water to the trans position of the oxo ligand was involved. (orig.)

  15. Keeping it together: Semantic coherence stabilizes phonological sequences in short-term memory.

    Science.gov (United States)

    Savill, Nicola; Ellis, Rachel; Brooke, Emma; Koa, Tiffany; Ferguson, Suzie; Rojas-Rodriguez, Elena; Arnold, Dominic; Smallwood, Jonathan; Jefferies, Elizabeth

    2018-04-01

    Our ability to hold a sequence of speech sounds in mind, in the correct configuration, supports many aspects of communication, but the contribution of conceptual information to this basic phonological capacity remains controversial. Previous research has shown modest and inconsistent benefits of meaning on phonological stability in short-term memory, but these studies were based on sets of unrelated words. Using a novel design, we examined the immediate recall of sentence-like sequences with coherent meaning, alongside both standard word lists and mixed lists containing words and nonwords. We found, and replicated, substantial effects of coherent meaning on phoneme-level accuracy: The phonemes of both words and nonwords within conceptually coherent sequences were more likely to be produced together and in the correct order. Since nonwords do not exist as items in long-term memory, the semantic enhancement of phoneme-level recall for both item types cannot be explained by a lexically based item reconstruction process employed at the point of retrieval ("redintegration"). Instead, our data show, for naturalistic input, that when meaning emerges from the combination of words, the phonological traces that support language are reinforced by a semantic-binding process that has been largely overlooked by past short-term memory research.

  16. MATAM: reconstruction of phylogenetic marker genes from short sequencing reads in metagenomes.

    Science.gov (United States)

    Pericard, Pierre; Dufresne, Yoann; Couderc, Loïc; Blanquart, Samuel; Touzet, Hélène

    2018-02-15

    Advances in the sequencing of uncultured environmental samples, dubbed metagenomics, raise a growing need for accurate taxonomic assignment. Accurate identification of organisms present within a community is essential to understanding even the most elementary ecosystems. However, current high-throughput sequencing technologies generate short reads which partially cover full-length marker genes and this poses difficult bioinformatic challenges for taxonomy identification at high resolution. We designed MATAM, a software dedicated to the fast and accurate targeted assembly of short reads sequenced from a genomic marker of interest. The method implements a stepwise process based on construction and analysis of a read overlap graph. It is applied to the assembly of 16S rRNA markers and is validated on simulated, synthetic and genuine metagenomes. We show that MATAM outperforms other available methods in terms of low error rates and recovered fractions and is suitable to provide improved assemblies for precise taxonomic assignments. https://github.com/bonsai-team/matam. pierre.pericard@gmail.com or helene.touzet@univ-lille1.fr. Supplementary data are available at Bioinformatics online. © The Author (2017). Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com

  17. 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...

  18. 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.

  19. 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.

  20. Genome dynamics of short oligonucleotides: the example of bacterial DNA uptake enhancing sequences.

    Directory of Open Access Journals (Sweden)

    Mohammed Bakkali

    Full Text Available Among the many bacteria naturally competent for transformation by DNA uptake-a phenomenon with significant clinical and financial implications- Pasteurellaceae and Neisseriaceae species preferentially take up DNA containing specific short sequences. The genomic overrepresentation of these DNA uptake enhancing sequences (DUES causes preferential uptake of conspecific DNA, but the function(s behind this overrepresentation and its evolution are still a matter for discovery. Here I analyze DUES genome dynamics and evolution and test the validity of the results to other selectively constrained oligonucleotides. I use statistical methods and computer simulations to examine DUESs accumulation in Haemophilus influenzae and Neisseria gonorrhoeae genomes. I analyze DUESs sequence and nucleotide frequencies, as well as those of all their mismatched forms, and prove the dependence of DUESs genomic overrepresentation on their preferential uptake by quantifying and correlating both characteristics. I then argue that mutation, uptake bias, and weak selection against DUESs in less constrained parts of the genome combined are sufficient enough to cause DUESs accumulation in susceptible parts of the genome with no need for other DUES function. The distribution of overrepresentation values across sequences with different mismatch loads compared to the DUES suggests a gradual yet not linear molecular drive of DNA sequences depending on their similarity to the DUES. Other genomically overrepresented sequences, both pro- and eukaryotic, show similar distribution of frequencies suggesting that the molecular drive reported above applies to other frequent oligonucleotides. Rare oligonucleotides, however, seem to be gradually drawn to genomic underrepresentation, thus, suggesting a molecular drag. To my knowledge this work provides the first clear evidence of the gradual evolution of selectively constrained oligonucleotides, including repeated, palindromic and protein

  1. Accurate estimation of short read mapping quality for next-generation genome sequencing

    Science.gov (United States)

    Ruffalo, Matthew; Koyutürk, Mehmet; Ray, Soumya; LaFramboise, Thomas

    2012-01-01

    Motivation: Several software tools specialize in the alignment of short next-generation sequencing reads to a reference sequence. Some of these tools report a mapping quality score for each alignment—in principle, this quality score tells researchers the likelihood that the alignment is correct. However, the reported mapping quality often correlates weakly with actual accuracy and the qualities of many mappings are underestimated, encouraging the researchers to discard correct mappings. Further, these low-quality mappings tend to correlate with variations in the genome (both single nucleotide and structural), and such mappings are important in accurately identifying genomic variants. Approach: We develop a machine learning tool, LoQuM (LOgistic regression tool for calibrating the Quality of short read mappings, to assign reliable mapping quality scores to mappings of Illumina reads returned by any alignment tool. LoQuM uses statistics on the read (base quality scores reported by the sequencer) and the alignment (number of matches, mismatches and deletions, mapping quality score returned by the alignment tool, if available, and number of mappings) as features for classification and uses simulated reads to learn a logistic regression model that relates these features to actual mapping quality. Results: We test the predictions of LoQuM on an independent dataset generated by the ART short read simulation software and observe that LoQuM can ‘resurrect’ many mappings that are assigned zero quality scores by the alignment tools and are therefore likely to be discarded by researchers. We also observe that the recalibration of mapping quality scores greatly enhances the precision of called single nucleotide polymorphisms. Availability: LoQuM is available as open source at http://compbio.case.edu/loqum/. Contact: matthew.ruffalo@case.edu. PMID:22962451

  2. Solvent polarity controls the helical conformation of short peptides rich in Calpha-tetrasubstituted amino acids.

    Science.gov (United States)

    Bellanda, Massimo; Mammi, Stefano; Geremia, Silvano; Demitri, Nicola; Randaccio, Lucio; Broxterman, Quirinus B; Kaptein, Bernard; Pengo, Paolo; Pasquato, Lucia; Scrimin, Paolo

    2007-01-01

    The two peptides, rich in C(alpha)-tetrasubstituted amino acids, Ac-[Aib-L-(alphaMe)Val-Aib](2)-L-His-NH(2) (1) and Ac-[Aib-L-(alphaMe)Val-Aib](2)-O-tBu (2 a) are prevalently helical. They present the unique property of changing their conformation from the alpha- to the 3(10)-helix as a function of the polarity of the solvent: alpha in more polar solvents, 3(10) in less polar ones. Conclusive evidence of this reversible change of conformation is reported on the basis of the circular dichroism (CD) spectra and a detailed two-dimensional NMR analysis in two solvents (trifluoroethanol and methanol) refined with molecular dynamics calculations. The X-ray diffractometric analysis of the crystals of both peptides reveals that they assume a prevalent 3(10)-helix conformation in the solid state. This conformation is practically superimposable on that obtained from the NMR analysis of 1 in methanol. The NMR results further validate the reported CD signature of the 3(10)-helix and the use of the CD technique for its assessment.

  3. Efficient Cargo Delivery into Adult Brain Tissue Using Short Cell-Penetrating Peptides.

    Directory of Open Access Journals (Sweden)

    Caghan Kizil

    Full Text Available Zebrafish brains can regenerate lost neurons upon neurogenic activity of the radial glial progenitor cells (RGCs that reside at the ventricular region. Understanding the molecular events underlying this ability is of great interest for translational studies of regenerative medicine. Therefore, functional analyses of gene function in RGCs and neurons are essential. Using cerebroventricular microinjection (CVMI, RGCs can be targeted efficiently but the penetration capacity of the injected molecules reduces dramatically in deeper parts of the brain tissue, such as the parenchymal regions that contain the neurons. In this report, we tested the penetration efficiency of five known cell-penetrating peptides (CPPs and identified two- polyR and Trans - that efficiently penetrate the brain tissue without overt toxicity in a dose-dependent manner as determined by TUNEL staining and L-Plastin immunohistochemistry. We also found that polyR peptide can help carry plasmid DNA several cell diameters into the brain tissue after a series of coupling reactions using DBCO-PEG4-maleimide-based Michael's addition and azide-mediated copper-free click reaction. Combined with the advantages of CVMI, such as rapidness, reproducibility, and ability to be used in adult animals, CPPs improve the applicability of the CVMI technique to deeper parts of the central nervous system tissues.

  4. CORE-SINEs: eukaryotic short interspersed retroposing elements with common sequence motifs.

    Science.gov (United States)

    Gilbert, N; Labuda, D

    1999-03-16

    A 65-bp "core" sequence is dispersed in hundreds of thousands copies in the human genome. This sequence was found to constitute the central segment of a group of short interspersed elements (SINEs), referred to as mammalian-wide interspersed repeats, that proliferated before the radiation of placental mammals. Here, we propose that the core identifies an ancient tRNA-like SINE element, which survived in different lineages such as mammals, reptiles, birds, and fish, as well as mollusks, presumably for >550 million years. This element gave rise to a number of sequence families (CORE-SINEs), including mammalian-wide interspersed repeats, whose distinct 3' ends are shared with different families of long interspersed elements (LINEs). The evolutionary success of the generic CORE-SINE element can be related to the recruitment of the internal promoter from highly transcribed host RNA as well as to its capacity to adapt to changing retropositional opportunities by sequence exchange with actively amplifying LINEs. It reinforces the notion that the very existence of SINEs depends on the cohabitation with both LINEs and the host genome.

  5. Sequence, taste and umami-enhancing effect of the peptides separated from soy sauce.

    Science.gov (United States)

    Zhuang, Mingzhu; Lin, Lianzhu; Zhao, Mouming; Dong, Yi; Sun-Waterhouse, Dongxiao; Chen, Huiping; Qiu, Chaoying; Su, Guowan

    2016-09-01

    Five tasty peptides were separated from soy sauce, by sensory-guided fractionation, using macroporous resin, medium-pressure liquid chromatography and reverse phase-high performance liquid chromatography, and identified by ultra-performance liquid chromatography tandem mass-spectrometry as ALPEEV, LPEEV, AQALQAQA, EQQQQ and EAGIQ (which originated from glycinin A1bB2-445, glycinin A1bB2-445, cobyric acid synthase, leucine-tRNA ligase and glycoprotein glucosyltransferase, respectively). LPEEV, AQALQAQA and EQQQQ tasted umami with threshold values of 0.43, 1.25 and 0.76mmol/l, respectively. ALPEEV and EAGIQ had minimal umami taste, but ALPEEV, EAGIQ and LPEEV showed umami-enhancement with a threshold estimated at 1.52, 1.94 and 3.41mmol/l, respectively. In addition, the synthetic peptides showed much better sensory taste than mixtures of their constitutive amino acids. It indicated that peptides might play an important role in the umami taste of soy sauce. Copyright © 2016 Elsevier Ltd. All rights reserved.

  6. Effects of High Intensity White Noise on Short-Term Memory for Position in a List and Sequence

    Science.gov (United States)

    Daee, Safar; Wilding, J. M.

    1977-01-01

    Seven experiments are described investigating the effecy of high intensity white noise during the visual presentation of words on a number of short-term memory tasks. Examines results relative to position learning and sequence learning. (Editor/RK)

  7. Comparison Review of Short-Acting and Long-Acting Glucagon-like Peptide-1 Receptor Agonists.

    Science.gov (United States)

    Uccellatore, Annachiara; Genovese, Stefano; Dicembrini, Ilaria; Mannucci, Edoardo; Ceriello, Antonio

    2015-09-01

    Glucagon-like peptide-1 (GLP-1) receptor agonists (GLP-1 RAs) are useful tools for treating type 2 diabetes mellitus. In their recent position statement, the American Diabetes Association and European Association for the Study of Diabetes recommend GLP1-RAs as add-on to metformin when therapeutic goals are not achieved with monotherapy, particularly for patients who wish to avoid weight gain or hypoglycemia. GLP1-RAs differ substantially in their duration of action, frequency of administration and clinical profile. Members of this class approved for clinical use include exenatide twice-daily, exenatide once-weekly, liraglutide and lixisenatide once-daily. Recently, two new once-weekly GLP1-RAs have been approved: dulaglutide and albiglutide. This article summarizes properties of short- and long-acting GLP-1 analogs, and provides useful information to help choose the most appropriate compound for individual patients.

  8. Efficient and controllable thermal ablation induced by short-pulsed HIFU sequence assisted with perfluorohexane nanodroplets.

    Science.gov (United States)

    Chang, Nan; Lu, Shukuan; Qin, Dui; Xu, Tianqi; Han, Meng; Wang, Supin; Wan, Mingxi

    2018-07-01

    A HIFU sequence with extremely short pulse duration and high pulse repetition frequency can achieve thermal ablation at a low acoustic power using inertial cavitation. Because of its cavitation-dependent property, the therapeutic outcome is unreliable when the treatment zone lacks cavitation nuclei. To overcome this intrinsic limitation, we introduced perfluorocarbon nanodroplets as extra cavitation nuclei into short-pulsed HIFU-mediated thermal ablation. Two types of nanodroplets were used with perfluorohexane (PFH) as the core material coated with bovine serum albumin (BSA) or an anionic fluorosurfactant (FS) to demonstrate the feasibility of this study. The thermal ablation process was recorded by high-speed photography. The inertial cavitation activity during the ablation was revealed by sonoluminescence (SL). The high-speed photography results show that the thermal ablation volume increased by ∼643% and 596% with BSA-PFH and FS-PFH, respectively, than the short-pulsed HIFU alone at an acoustic power of 19.5 W. Using nanodroplets, much larger ablation volumes were created even at a much lower acoustic power. Meanwhile, the treatment time for ablating a desired volume significantly reduced in the presence of nanodroplets. Moreover, by adjusting the treatment time, lesion migration towards the HIFU transducer could also be avoided. The SL results show that the thermal lesion shape was significantly dependent on the inertial cavitation in this short-pulsed HIFU-mediated thermal ablation. The inertial cavitation activity became more predictable by using nanodroplets. Therefore, the introduction of PFH nanodroplets as extra cavitation nuclei made the short-pulsed HIFU thermal ablation more efficient by increasing the ablation volume and speed, and more controllable by reducing the acoustic power and preventing lesion migration. Copyright © 2018. Published by Elsevier B.V.

  9. Fine-mapping of immunodominant linear B-cell epitopes of the Staphylococcus aureus SEB antigen using short overlapping peptides.

    Directory of Open Access Journals (Sweden)

    Zhuo Zhao

    Full Text Available Staphylococcal enterotoxin B (SEB is one of the most potent Staphylococcus aureus exotoxins (SEs. Due to its conserved sequence and stable structure, SEB might be a good candidate antigen for MRSA vaccines. Although cellular immune responses to SEB are well-characterized, much less is known regarding SEB-specific humoral immune responses, particularly regarding detailed epitope mapping. In this study, we utilized a recombinant nontoxic mutant of SEB (rSEB and an AlPO4 adjuvant to immunize BALB/c mice and confirmed that rSEB can induce a high antibody level and effective immune protection against MRSA infection. Next, the antisera of immunized mice were collected, and linear B cell epitopes within SEB were finely mapped using a series of overlapping synthetic peptides. Three immunodominant B cell epitopes of SEB were screened by ELISA, including a novel epitope, SEB205-222, and two known epitopes, SEB97-114 and SEB247-261. Using truncated peptides, an ELISA was performed with peptide-KLH antisera, and the core sequence of the three immunodominant B cell epitopes were verified as SEB97-112, SEB207-222, and SEB247-257. In vitro, all of the immunodominant epitope-specific antisera (anti-SEB97-112, anti-SEB207-222 and anti-SEB247-257 were observed to inhibit SEB-induced T cell mitogenesis and cytokine production from splenic lymphocytes of BALB/c mice. The homology analysis indicated that SEB97-112 and SEB207-222 were well-conserved among different Staphylococcus aureus strains. The 3D crystal structure of SEB indicated that SEB97-112 was in the loop region inside SEB, whereas SEB207-222 and SEB247-257 were in the β-slice region outside SEB. In summary, the fine-mapping of linear B-cell epitopes of the SEB antigen in this study will be useful to understand anti-SEB immunity against MRSA infection further and will be helpful to optimize MRSA vaccine designs that are based on the SEB antigen.

  10. Sequence-specific inhibition of duck hepatitis B virus reverse transcription by peptide nucleic acids (PNA)

    DEFF Research Database (Denmark)

    Robaczewska, Magdalena; Narayan, Ramamurthy; Seigneres, Beatrice

    2005-01-01

    BACKGROUND/AIMS: Peptide nucleic acids (PNAs) appear as promising new antisense agents, that have not yet been examined as hepatitis B virus (HBV) inhibitors. Our aim was to study the ability of PNAs targeting the duck HBV (DHBV) encapsidation signal epsilon to inhibit reverse transcription (RT...... in primary duck hepatocytes (PDH). RESULTS: Both PNAs reproducibly inhibited DHBV RT in a dose-dependent manner with IC(50) of 10nM, whereas up to 600-fold higher concentration of S-ODNs was required for similar inhibition. The PNA targeting the bulge and upper stem of epsilon appeared as more efficient RT...

  11. Computational complexity of algorithms for sequence comparison, short-read assembly and genome alignment.

    Science.gov (United States)

    Baichoo, Shakuntala; Ouzounis, Christos A

    A multitude of algorithms for sequence comparison, short-read assembly and whole-genome alignment have been developed in the general context of molecular biology, to support technology development for high-throughput sequencing, numerous applications in genome biology and fundamental research on comparative genomics. The computational complexity of these algorithms has been previously reported in original research papers, yet this often neglected property has not been reviewed previously in a systematic manner and for a wider audience. We provide a review of space and time complexity of key sequence analysis algorithms and highlight their properties in a comprehensive manner, in order to identify potential opportunities for further research in algorithm or data structure optimization. The complexity aspect is poised to become pivotal as we will be facing challenges related to the continuous increase of genomic data on unprecedented scales and complexity in the foreseeable future, when robust biological simulation at the cell level and above becomes a reality. Copyright © 2017 Elsevier B.V. All rights reserved.

  12. Sequence specificity for uridylylation of the viral peptide linked to the genome (VPg) of enteroviruses.

    Science.gov (United States)

    Schein, Catherine H; Ye, Mengyi; Paul, Aniko V; Oberste, M Steven; Chapman, Nora; van der Heden van Noort, Gerbrand J; Filippov, Dmitri V; Choi, Kyung H

    2015-10-01

    Enteroviruses (EV) uridylylate a peptide, VPg, as the first step in their replication. VPgpUpU, found free in infected cells, serves as the primer for RNA elongation. The abilities of four polymerases (3D(pol)), from EV-species A-C, to uridylylate VPgs that varied by up to 60% of their residues were compared. Each 3D(pol) was able to uridylylate all five VPgs using polyA RNA as template, while showing specificity for its own genome encoded peptide. All 3D(pol) uridylylated a consensus VPg representing the physical chemical properties of 31 different VPgs. Thus the residues required for uridylylation and the enzymatic mechanism must be similar in diverse EV. As VPg-binding sites differ in co-crystal structures, the reaction is probably done by a second 3D(pol) molecule. The conservation of polymerase residues whose mutation reduces uridylylation but not RNA elongation is compared. Copyright © 2015 Elsevier Inc. All rights reserved.

  13. Novel Kunitz-like Peptides Discovered in the Zoanthid Palythoa caribaeorum through Transcriptome Sequencing.

    Science.gov (United States)

    Liao, Qiwen; Li, Shengnan; Siu, Shirley Weng In; Yang, Binrui; Huang, Chen; Chan, Judy Yuet-Wa; Morlighem, Jean-Étienne R L; Wong, Clarence Tsun Ting; Rádis-Baptista, Gandhi; Lee, Simon Ming-Yuen

    2018-02-02

    Palythoa caribaeorum (class Anthozoa) is a zoanthid that together jellyfishes, hydra, and sea anemones, which are venomous and predatory, belongs to the Phyllum Cnidaria. The distinguished feature in these marine animals is the cnidocytes in the body tissues, responsible for toxin production and injection that are used majorly for prey capture and defense. With exception for other anthozoans, the toxin cocktails of zoanthids have been scarcely studied and are poorly known. Here, on the basis of the analysis of P. caribaeorum transcriptome, numerous predicted venom-featured polypeptides were identified including allergens, neurotoxins, membrane-active, and Kunitz-like peptides (PcKuz). The three predicted PcKuz isotoxins (1-3) were selected for functional studies. Through computational processing comprising structural phylogenetic analysis, molecular docking, and dynamics simulation, PcKuz3 was shown to be a potential voltage gated potassium-channel inhibitor. PcKuz3 fitted well as new functional Kunitz-type toxins with strong antilocomotor activity as in vivo assessed in zebrafish larvae, with weak inhibitory effect toward proteases, as evaluated in vitro. Notably, PcKuz3 can suppress, at low concentration, the 6-OHDA-induced neurotoxicity on the locomotive behavior of zebrafish, which indicated PcKuz3 may have a neuroprotective effect. Taken together, PcKuz3 figures as a novel neurotoxin structure, which differs from known homologous peptides expressed in sea anemone. Moreover, the novel PcKuz3 provides an insightful hint for biodrug development for prospective neurodegenerative disease treatment.

  14. Identification of Cyclin A Binders with a Fluorescent Peptide Sensor.

    Science.gov (United States)

    Pazos, Elena; Mascareñas, José L; Vázquez, M Eugenio

    2016-01-01

    A peptide sensor that integrates the 4-dimethylaminophthalimide (4-DMAP) fluorophore in a short cyclin A binding sequence displays a large fluorescence emission increase upon interacting with the cyclin A Binding Groove (CBG). Competitive displacement assays of this probe allow the straightforward identification of peptides that interact with the CBG, which could potentially block the recognition of CDK/cyclin A kinase substrates.

  15. The genome of flax (Linum usitatissimum) assembled de novo from short shotgun sequence reads

    DEFF Research Database (Denmark)

    Wang, Zhiwen; Hobson, Neil; Galindo, Leonardo

    2012-01-01

    Flax (Linum usitatissimum) is an ancient crop that is widely cultivated as a source of fiber, oil and medicinally relevant compounds. To accelerate crop improvement, we performed whole-genome shotgun sequencing of the nuclear genome of flax. Seven paired-end libraries ranging in size from 300 bp...... these results show that de novo assembly, based solely on whole-genome shotgun short-sequence reads, is an efficient means of obtaining nearly complete genome sequence information for some plant species....

  16. Short Stat5-interacting peptide derived from phospholipase C-β3 inhibits hematopoietic cell proliferation and myeloid differentiation.

    Directory of Open Access Journals (Sweden)

    Hiroki Yasudo

    Full Text Available Constitutive activation of the transcription factor Stat5 in hematopoietic stem/progenitor cells leads to various hematopoietic malignancies including myeloproliferative neoplasm (MPN. Our recent study found that phospholipase C (PLC-β3 is a novel tumor suppressor involved in MPN, lymphoma and other tumors. Stat5 activity is negatively regulated by the SH2 domain-containing protein phosphatase SHP-1 in a PLC-β3-dependent manner. PLC-β3 can form the multimolecular SPS complex together with SHP-1 and Stat5. The close physical proximity of SHP-1 and Stat5 brought about by interacting with the C-terminal segment of PLC-β3 (PLC-β3-CT accelerates SHP-1-mediated dephosphorylation of Stat5. Here we identify the minimal sequences within PLC-β3-CT required for its tumor suppressor function. Two of the three Stat5-binding noncontiguous regions, one of which also binds SHP-1, substantially inhibited in vitro proliferation of Ba/F3 cells. Surprisingly, an 11-residue Stat5-binding peptide (residues 988-998 suppressed Stat5 activity in Ba/F3 cells and in vivo proliferation and myeloid differentiation of hematopoietic stem/progenitor cells. Therefore, this study further defines PLC-β3-CT as the Stat5- and SHP-1-binding domain by identifying minimal functional sequences of PLC-β3 for its tumor suppressor function and implies their potential utility in the control of hematopoietic malignancies.

  17. Study of the fast inversion recovery pulse sequence. With reference to fast fluid attenuated inversion recovery and fast short TI inversion recovery pulse sequence

    International Nuclear Information System (INIS)

    Tsuchihashi, Toshio; Maki, Toshio; Suzuki, Takeshi

    1997-01-01

    The fast inversion recovery (fast IR) pulse sequence was evaluated. We compared the fast fluid attenuated inversion recovery (fast FLAIR) pulse sequence in which inversion time (TI) was established as equal to the water null point for the purpose of the water-suppressed T 2 -weighted image, with the fast short TI inversion recovery (fast STIR) pulse sequence in which TI was established as equal to the fat null point for purpose of fat suppression. In the fast FLAIR pulse sequence, the water null point was increased by making TR longer. In the FLAIR pulse sequence, the longitudinal magnetization contrast is determined by TI. If TI is increased, T 2 -weighted contrast improves in the same way as increasing TR for the SE pulse sequence. Therefore, images should be taken with long TR and long TI, which are longer than TR and longer than the water null point. On the other hand, the fat null point is not affected by TR in the fast STIR pulse sequence. However, effective TE was affected by variation of the null point. This increased in proportion to the increase in effective TE. Our evaluation indicated that the fast STIR pulse sequence can control the extensive signals from fat in a short time. (author)

  18. Random sequences are an abundant source of bioactive RNAs or peptides

    DEFF Research Database (Denmark)

    Neme, Rafik; Amador, Cristina; Yildirim, Burcin

    2017-01-01

    It is generally assumed that new genes arise through duplication and/or recombination of existing genes. The probability that a new functional gene could arise out of random non-coding DNA is so far considered to be negligible, as it seems unlikely that such an RNA or protein sequence could have ...

  19. N-terminal diproline and charge group effects on the stabilization of helical conformation in alanine-based short peptides: CD studies with water and methanol as solvent.

    Science.gov (United States)

    Goyal, Bhupesh; Srivastava, Kinshuk Raj; Durani, Susheel

    2017-06-01

    Protein folding problem remains a formidable challenge as main chain, side chain and solvent interactions remain entangled and have been difficult to resolve. Alanine-based short peptides are promising models to dissect protein folding initiation and propagation structurally as well as energetically. The effect of N-terminal diproline and charged side chains is assessed on the stabilization of helical conformation in alanine-based short peptides using circular dichroism (CD) with water and methanol as solvent. A1 (Ac-Pro-Pro-Ala-Lys-Ala-Lys-Ala-Lys-Ala-NH 2 ) is designed to assess the effect of N-terminal homochiral diproline and lysine side chains to induce helical conformation. A2 (Ac-Pro-Pro-Glu-Glu-Ala-Ala-Lys-Lys-Ala-NH 2 ) and A3 (Ac-dPro-Pro-Glu-Glu-Ala-Ala-Lys-Lys-Ala-NH 2 ) with N-terminal homochiral and heterochiral diproline, respectively, are designed to assess the effect of Glu...Lys (i, i + 4) salt bridge interactions on the stabilization of helical conformation. The CD spectra of A1, A2 and A3 in water manifest different amplitudes of the observed polyproline II (PPII) signals, which indicate different conformational distributions of the polypeptide structure. The strong effect of solvent substitution from water to methanol is observed for the peptides, and CD spectra in methanol evidence A2 and A3 as helical folds. Temperature-dependent CD spectra of A1 and A2 in water depict an isodichroic point reflecting coexistence of two conformations, PPII and β-strand conformation, which is consistent with the previous studies. The results illuminate the effect of N-terminal diproline and charged side chains in dictating the preferences for extended-β, semi-extended PPII and helical conformation in alanine-based short peptides. The results of the present study will enhance our understanding on stabilization of helical conformation in short peptides and hence aid in the design of novel peptides with helical structures. Copyright © 2017 European Peptide

  20. Choice of reference sequence and assembler for alignment of Listeria monocytogenes short-read sequence data greatly influences rates of error in SNP analyses.

    Directory of Open Access Journals (Sweden)

    Arthur W Pightling

    Full Text Available The wide availability of whole-genome sequencing (WGS and an abundance of open-source software have made detection of single-nucleotide polymorphisms (SNPs in bacterial genomes an increasingly accessible and effective tool for comparative analyses. Thus, ensuring that real nucleotide differences between genomes (i.e., true SNPs are detected at high rates and that the influences of errors (such as false positive SNPs, ambiguously called sites, and gaps are mitigated is of utmost importance. The choices researchers make regarding the generation and analysis of WGS data can greatly influence the accuracy of short-read sequence alignments and, therefore, the efficacy of such experiments. We studied the effects of some of these choices, including: i depth of sequencing coverage, ii choice of reference-guided short-read sequence assembler, iii choice of reference genome, and iv whether to perform read-quality filtering and trimming, on our ability to detect true SNPs and on the frequencies of errors. We performed benchmarking experiments, during which we assembled simulated and real Listeria monocytogenes strain 08-5578 short-read sequence datasets of varying quality with four commonly used assemblers (BWA, MOSAIK, Novoalign, and SMALT, using reference genomes of varying genetic distances, and with or without read pre-processing (i.e., quality filtering and trimming. We found that assemblies of at least 50-fold coverage provided the most accurate results. In addition, MOSAIK yielded the fewest errors when reads were aligned to a nearly identical reference genome, while using SMALT to align reads against a reference sequence that is ∼0.82% distant from 08-5578 at the nucleotide level resulted in the detection of the greatest numbers of true SNPs and the fewest errors. Finally, we show that whether read pre-processing improves SNP detection depends upon the choice of reference sequence and assembler. In total, this study demonstrates that researchers

  1. Novel nonphosphorylated peptides with conserved sequences selectively bind to Grb7 SH2 domain with affinity comparable to its phosphorylated ligand.

    Directory of Open Access Journals (Sweden)

    Dan Zhang

    Full Text Available The Grb7 (growth factor receptor-bound 7 protein, a member of the Grb7 protein family, is found to be highly expressed in such metastatic tumors as breast cancer, esophageal cancer, liver cancer, etc. The src-homology 2 (SH2 domain in the C-terminus is reported to be mainly involved in Grb7 signaling pathways. Using the random peptide library, we identified a series of Grb7 SH2 domain-binding nonphosphorylated peptides in the yeast two-hybrid system. These peptides have a conserved GIPT/K/N sequence at the N-terminus and G/WD/IP at the C-terminus, and the region between the N-and C-terminus contains fifteen amino acids enriched with serines, threonines and prolines. The association between the nonphosphorylated peptides and the Grb7 SH2 domain occurred in vitro and ex vivo. When competing for binding to the Grb7 SH2 domain in a complex, one synthesized nonphosphorylated ligand, containing the twenty-two amino acid-motif sequence, showed at least comparable affinity to the phosphorylated ligand of ErbB3 in vitro, and its overexpression inhibited the proliferation of SK-BR-3 cells. Such nonphosphorylated peptides may be useful for rational design of drugs targeted against cancers that express high levels of Grb7 protein.

  2. Short- and long-term evolutionary dynamics of bacterial insertion sequences: insights from Wolbachia endosymbionts.

    Science.gov (United States)

    Cerveau, Nicolas; Leclercq, Sébastien; Leroy, Elodie; Bouchon, Didier; Cordaux, Richard

    2011-01-01

    Transposable elements (TE) are one of the major driving forces of genome evolution, raising the question of the long-term dynamics underlying their evolutionary success. Long-term TE evolution can readily be reconstructed in eukaryotes, thanks to many degraded copies constituting genomic fossil records of past TE proliferations. By contrast, bacterial genomes usually experience high sequence turnover and short TE retention times, thereby obscuring ancient TE evolutionary patterns. We found that Wolbachia bacterial genomes contain 52-171 insertion sequence (IS) TEs. IS account for 11% of Wolbachia wRi, which is one of the highest IS genomic coverage reported in prokaryotes to date. We show that many IS groups are currently expanding in various Wolbachia genomes and that IS horizontal transfers are frequent among strains, which can explain the apparent synchronicity of these IS proliferations. Remarkably, >70% of Wolbachia IS are nonfunctional. They constitute an unusual bacterial IS genomic fossil record providing direct empirical evidence for a long-term IS evolutionary dynamics following successive periods of intense transpositional activity. Our results show that comprehensive IS annotations have the potential to provide new insights into prokaryote TE evolution and, more generally, prokaryote genome evolution. Indeed, the identification of an important IS genomic fossil record in Wolbachia demonstrates that IS elements are not always of recent origin, contrary to the conventional view of TE evolution in prokaryote genomes. Our results also raise the question whether the abundance of IS fossils is specific to Wolbachia or it may be a general, albeit overlooked, feature of prokaryote genomes.

  3. Effect of amino acid sequence and pH on nanofiber formation of self-assembling peptides EAK16-II and EAK16-IV.

    Science.gov (United States)

    Hong, Yooseong; Legge, Raymond L; Zhang, S; Chen, P

    2003-01-01

    Atomic force microscopy (AFM) and axisymmetric drop shape analysis-profile (ASDA-P) were used to investigate the mechanism of self-assembly of peptides. The peptides chosen consisted of 16 alternating hydrophobic and hydrophilic amino acids, where the hydrophilic residues possess alternating negative and positive charges. Two types of peptides, AEAEAKAKAEAEAKAK (EAK16-II) and AEAEAEAEAKAKAKAK (EAK16-IV), were investigated in terms of nanostructure formation through self-assembly. The experimental results, which focused on the effects of the amino acid sequence and pH, show that the nanostructures formed by the peptides are dependent on the amino acid sequence and the pH of the solution. For pH conditions around neutrality, one of the peptides used in this study, EAK16-IV, forms globular assemblies and has lower surface tension at air-water interfaces than another peptide, EAK16-II, which forms fibrillar assemblies at the same pH. When the pH is lowered below 6.5 or raised above 7.5, there is a transition from globular to fibrillar structures for EAK16-IV, but EAK16-II does not show any structural transition. Surface tension measurements using ADSA-P showed different surface activities of peptides at air-water interfaces. EAK16-II does not show a significant difference in surface tension for the pH range between 4 and 9. However, EAK16-IV shows a noticeable decrease in surface tension at pH around neutrality, indicating that the formation of globular assemblies is related to the molecular hydrophobicity.

  4. NNAlign: A Web-Based Prediction Method Allowing Non-Expert End-User Discovery of Sequence Motifs in Quantitative Peptide Data

    DEFF Research Database (Denmark)

    Andreatta, Massimo; Schafer-Nielsen, Claus; Lund, Ole

    2011-01-01

    Recent advances in high-throughput technologies have made it possible to generate both gene and protein sequence data at an unprecedented rate and scale thereby enabling entirely new "omics"-based approaches towards the analysis of complex biological processes. However, the amount and complexity...... to interpret large data sets. We have recently developed a method, NNAlign, which is generally applicable to any biological problem where quantitative peptide data is available. This method efficiently identifies underlying sequence patterns by simultaneously aligning peptide sequences and identifying motifs...... associated with quantitative readouts. Here, we provide a web-based implementation of NNAlign allowing non-expert end-users to submit their data (optionally adjusting method parameters), and in return receive a trained method (including a visual representation of the identified motif) that subsequently can...

  5. Human T-cell recognition of synthetic peptides representing conserved and variant sequences from the merozoite surface protein 2 of Plasmodium falciparum

    DEFF Research Database (Denmark)

    Theander, T G; Hviid, L; Dodoo, D

    1997-01-01

    Merozoite surface protein 2 (MSP2) is a malaria vaccine candidate currently undergoing clinical trials. We analyzed the peripheral blood mononuclear cell (PBMC) response to synthetic peptides corresponding to conserved and variant regions of the FCQ-27 allelic form of MSP2 in Ghanaian individuals....... The findings are encouraging for the development of a vaccine based on these T-epitope containing regions of MSP2, as the peptides were broadly recognized suggesting that they can bind to diverse HLA alleles and also because they include conserved MSP2 sequences. Immunisation with a vaccine construct...

  6. Beta-blockers influence the short-term and long-term prognostic information of natriuretic peptides and catecholamines in chronic heart failure independent from specific agents.

    Science.gov (United States)

    Frankenstein, Lutz; Nelles, Manfred; Slavutsky, Maxim; Schellberg, Dieter; Doesch, Andreas; Katus, Hugo; Remppis, Andrew; Zugck, Christian

    2007-10-01

    In chronic heart failure (CHF), the physiologic effects of natriuretic peptides and catecholamines are interdependent. Furthermore, reports state an agent-dependent effect of individual beta-blockers on biomarkers. Data on the short-term and long-term predictive power comparing these biomarkers as well as accounting for the influence of beta-blocker treatment both on the marker or the resultant prognostic information are scarce. We included 513 consecutive patients with systolic CHF, measured atrial natriuretic peptide (ANP), N-terminal prohormone brain natriuretic peptide (NTproBNP), noradrenaline, and adrenaline, and monitored them for 90 +/- 25 months. Death or the combination of death and cardiac transplantation at 1 year, 5 years, and overall follow-up were considered end points. Compared with patients not taking beta-blockers, patients taking beta-blockers had significantly lower levels of catecholamines but not natriuretic peptides. Only for adrenaline was the amount of this effect related to the specific beta-blocker chosen. Receiver operating characteristic curves demonstrated superior prognostic accuracy for NTproBNP both at the 1- and 5-year follow-up compared with ANP, noradrenaline, and adrenaline. In multivariate analysis including established risk markers (New York Heart Association functional class, left ventricular ejection fraction, peak oxygen uptake, and 6-minute walk test), of all neurohumoral parameters, only NTproBNP remained an independent predictor for both end points. Long-term beta-blocker therapy is associated with decreased levels of plasma catecholamines but not natriuretic peptides. This effect is independent from the actual beta-blocker chosen for natriuretic peptides and noradrenaline. In multivariate analysis, both for short-term and long-term prediction of mortality or the combined end point of death and cardiac transplantation, only NTproBNP remained independent from established clinical risk markers.

  7. 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.

  8. Assessing the 5S ribosomal RNA heterogeneity in Arabidopsis thaliana using short RNA next generation sequencing data.

    Science.gov (United States)

    Szymanski, Maciej; Karlowski, Wojciech M

    2016-01-01

    In eukaryotes, ribosomal 5S rRNAs are products of multigene families organized within clusters of tandemly repeated units. Accumulation of genomic data obtained from a variety of organisms demonstrated that the potential 5S rRNA coding sequences show a large number of variants, often incompatible with folding into a correct secondary structure. Here, we present results of an analysis of a large set of short RNA sequences generated by the next generation sequencing techniques, to address the problem of heterogeneity of the 5S rRNA transcripts in Arabidopsis and identification of potentially functional rRNA-derived fragments.

  9. Short-term glucagon stimulation test of C-peptide effect on glucose utilization in patients with type 1 diabetes mellitus.

    Science.gov (United States)

    Mojto, Viliam; Rausova, Zuzana; Chrenova, Jana; Dedik, Ladislav

    2015-12-01

    This work aimed to evaluate the use of a four-point glucagon stimulation test of C-peptide effect on glucose utilization in type 1 diabetic patients using a new mathematical model. A group of 32 type 1 diabetic patients and a group of 10 healthy control subjects underwent a four-point glucagon stimulation test with blood sampling at 0, 6, 15 and 30 min after 1 mg glucagon bolus intravenous administration. Pharmacokinetic and pharmacokinetic/pharmacodynamic models of C-peptide effect on glucose utilization versus area under curve (AUC) were used. A two-sample t test and ANOVA with Bonferroni correction were used to test the significance of differences between parameters. A significant difference between control and patient groups regarding the coefficient of whole-body glucose utilization and AUC C-peptide/AUC glucose ratio (p ≪ 0.001 and p = 0.002, respectively) was observed. The high correlation (r = 0.97) between modeled coefficient of whole-body glucose utilization and numerically calculated AUC C-peptide/AUC glucose ratio related to entire cohort indicated the stability of used method. The short-term four-point glucagon stimulation test allows the numerically calculated AUC C-peptide/AUC glucose ratio and/or the coefficient of whole-body glucose utilization calculated from model to be used to diagnostically identify type 1 diabetic patients.

  10. Use of eluted peptide sequence data to identify the binding characteristics of peptides to the insulin-dependent diabetes susceptibility allele HLA-DQ8 (DQ 3.2).

    Science.gov (United States)

    Godkin, A; Friede, T; Davenport, M; Stevanovic, S; Willis, A; Jewell, D; Hill, A; Rammensee, H G

    1997-06-01

    HLA-DQ8 (A1*0301, B1*0302) and -DQ2 (A1*0501, B1*0201) are both associated with diseases such as insulin-dependent diabetes mellitus and coeliac disease. We used the technique of pool sequencing to look at the requirements of peptides binding to HLA-DQ8, and combined these data with naturally sequenced ligands and in vitro binding assays to describe a novel motif for HLA-DQ8. The motif, which has the same basic format as many HLA-DR molecules, consists of four or five anchor regions, in the positions from the N-terminus of the binding core of n, n + 3, n + 5/6 and n + 8, i.e. P1, P4, P6/7 and P9. P1 and P9 require negative or polar residues, with mainly aliphatic residues at P4 and P6/7. The features of the HLA-DQ8 motif were then compared to a pool sequence of peptides eluted from HLA-DQ2. A consensus motif for the binding of a common peptide which may be involved in disease pathogenesis is described. Neither of the disease-associated alleles HLA-DQ2 and -DQ8 have Asp at position 57 of the beta-chain. This Asp, if present, may form a salt bridge with an Arg at position 79 of the alpha-chain and so alter the binding specificity of P9. HLA-DQ2 and -DQ8 both appear to prefer negatively charged amino acids at P9. In contrast, HLA-DQ7 (A1*0301, B1*0301), which is not associated with diabetes, has Asp at beta 57, allowing positively charged amino acids at P9. This analysis of the sequence features of DQ-binding peptides suggests molecular characteristics which may be useful to predict epitopes involved in disease pathogenesis.

  11. Resonant magnetoelectric response of composite cantilevers: Theory of short vs. open circuit operation and layer sequence effects

    Directory of Open Access Journals (Sweden)

    Matthias C. Krantz

    2015-11-01

    Full Text Available The magnetoelectric effect in layered composite cantilevers consisting of strain coupled layers of magnetostrictive (MS, piezoelectric (PE, and substrate materials is investigated for magnetic field excitation at bending resonance. Analytic theories are derived for the transverse magnetoelectric (ME response in short and open circuit operation for three different layer sequences and results presented and discussed for the FeCoBSi-AlN-Si and the FeCoBSi-PZT-Si composite systems. Response optimized PE-MS layer thickness ratios are found to greatly change with operation mode shifting from near equal MS and PE layer thicknesses in the open circuit mode to near vanishing PE layer thicknesses in short circuit operation for all layer sequences. In addition the substrate layer thickness is found to differently affect the open and short circuit ME response producing shifts and reversal between ME response maxima depending on layer sequence. The observed rich ME response behavior for different layer thicknesses, sequences, operating modes, and PE materials can be explained by common neutral plane effects and different elastic compliance effects in short and open circuit operation.

  12. [Possibilities in the differential diagnosis of brain neoplasms using the long and short time sequences of proton magnetic resonance spectroscopy

    NARCIS (Netherlands)

    Gajewicz, W.; Goraj, B.M.

    2004-01-01

    Currently to perform proton magnetic resonance spectroscopy (1H MRS) with single voxel spectroscopy (SVS) technique long and/or short echo time sequences are used in order to provide complementary information. PURPOSE: The aim of the study was to compare the usefulness of STEAM (time echo, TE, 20

  13. NetMHCpan, a method for quantitative predictions of peptide binding to any HLA-A and -B locus protein of known sequence

    DEFF Research Database (Denmark)

    Nielsen, Morten; Lundegaard, Claus; Blicher, Thomas

    2007-01-01

    BACKGROUND: Binding of peptides to Major Histocompatibility Complex (MHC) molecules is the single most selective step in the recognition of pathogens by the cellular immune system. The human MHC class I system (HLA-I) is extremely polymorphic. The number of registered HLA-I molecules has now surp...... to provide new basic insights into HLA structure-function relationships. The method is available at http://www.cbs.dtu.dk/services/NetMHCpan....... surpassed 1500. Characterizing the specificity of each separately would be a major undertaking. PRINCIPAL FINDINGS: Here, we have drawn on a large database of known peptide-HLA-I interactions to develop a bioinformatics method, which takes both peptide and HLA sequence information into account...... successfully validate this method. We further demonstrate that the method can be applied to perform a clustering analysis of MHC specificities and suggest using this clustering to select particularly informative novel MHC molecules for future biochemical and functional analysis. CONCLUSIONS: Encompassing all...

  14. A sensitive mass spectrometric method for hypothesis-driven detection of peptide post-translational modifications: multiple reaction monitoring-initiated detection and sequencing (MIDAS).

    Science.gov (United States)

    Unwin, Richard D; Griffiths, John R; Whetton, Anthony D

    2009-01-01

    The application of a targeted mass spectrometric workflow to the sensitive identification of post-translational modifications is described. This protocol employs multiple reaction monitoring (MRM) to search for all putative peptides specifically modified in a target protein. Positive MRMs trigger an MS/MS experiment to confirm the nature and site of the modification. This approach, termed MIDAS (MRM-initiated detection and sequencing), is more sensitive than approaches using neutral loss scanning or precursor ion scanning methodologies, due to a more efficient use of duty cycle along with a decreased background signal associated with MRM. We describe the use of MIDAS for the identification of phosphorylation, with a typical experiment taking just a couple of hours from obtaining a peptide sample. With minor modifications, the MIDAS method can be applied to other protein modifications or unmodified peptides can be used as a MIDAS target.

  15. 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.

  16. Short (

    NARCIS (Netherlands)

    Telleman, Gerdien; den Hartog, Laurens

    2013-01-01

    Aim: This systematic review assessed the implant survival rate of short (<10 mm) dental implants installed in partially edentulous patients. A case report of a short implant in the posterior region have been added. Materials and methods: A search was conducted in the electronic databases of MEDLINE

  17. Purification, characterization, and sequencing of novel antimicrobial peptides, Tu-AMP 1 and Tu-AMP 2, from bulbs of tulip (Tulipa gesneriana L.).

    Science.gov (United States)

    Fujimura, Masatoshi; Ideguchi, Mineo; Minami, Yuji; Watanabe, Keiichi; Tadera, Kenjiro

    2004-03-01

    Novel antimicrobial peptides (AMP), designated Tu-AMP 1 and Tu-AMP 2, were purified from the bulbs of tulip (Tulipa gesneriana L.) by chitin affinity chromatography and reverse-phase high-performance liquid chromatography (HPLC). They bind to chitin in a reversible way. They were basic peptides having isoelectric points of over 12. Tu-AMP 1 and Tu-AMP 2 had molecular masses of 4,988 Da and 5,006 Da on MALDI-TOF MS analysis, and their extinction coefficients of 1% aqueous solutions at 280 nm were 3.3 and 3.4, respectively. Half of all amino acid residues of Tu-AMP 1 and Tu-AMP 2 were occupied by cysteine, arginine, lysine, and proline. The concentrations of peptides required for 50% inhibition (IC(50)) of the growth of plant pathogenic bacteria and fungi were 2 to 20 microg/ml. The structural characteristics of Tu-AMP 1 and Tu-AMP 2 indicated that they were novel thionin-like antimicrobial peptides, though Tu-AMP 2 was a heterodimer composes of two short peptides joined with disulfide bonds.

  18. 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.

  19. Exploiting BAC-end sequences for the mining, characterization and utility of new short sequences repeat (SSR) markers in Citrus.

    Science.gov (United States)

    Biswas, Manosh Kumar; Chai, Lijun; Mayer, Christoph; Xu, Qiang; Guo, Wenwu; Deng, Xiuxin

    2012-05-01

    The aim of this study was to develop a large set of microsatellite markers based on publicly available BAC-end sequences (BESs), and to evaluate their transferability, discriminating capacity of genotypes and mapping ability in Citrus. A set of 1,281 simple sequence repeat (SSR) markers were developed from the 46,339 Citrus clementina BAC-end sequences (BES), of them 20.67% contained SSR longer than 20 bp, corresponding to roughly one perfect SSR per 2.04 kb. The most abundant motifs were di-nucleotide (16.82%) repeats. Among all repeat motifs (TA/AT)n is the most abundant (8.38%), followed by (AG/CT)n (4.51%). Most of the BES-SSR are located in the non-coding region, but 1.3% of BES-SSRs were found to be associated with transposable element (TE). A total of 400 novel SSR primer pairs were synthesized and their transferability and polymorphism tested on a set of 16 Citrus and Citrus relative's species. Among these 333 (83.25%) were successfully amplified and 260 (65.00%) showed cross-species transferability with Poncirus trifoliata and Fortunella sp. These cross-species transferable markers could be useful for cultivar identification, for genomic study of Citrus, Poncirus and Fortunella sp. Utility of the developed SSR marker was demonstrated by identifying a set of 118 markers each for construction of linkage map of Citrus reticulata and Poncirus trifoliata. Genetic diversity and phylogenetic relationship among 40 Citrus and its related species were conducted with the aid of 25 randomly selected SSR primer pairs and results revealed that citrus genomic SSRs are superior to genic SSR for genetic diversity and germplasm characterization of Citrus spp.

  20. Procedures of amino acid sequencing of peptides in natural proteins collection of knowledge and intelligence for construction of reliable chemical inference system

    OpenAIRE

    Kudo, Yoshihiro; Kanaya, Shigehiko

    1994-01-01

    In order to establish a reliable chemical inference system on amino acid sequencing of natural peptides, as various kinds of relevant knowledge and intelligence as possible are collected. Topics are on didemnins, dolastatin 3, TL-119 and/or A-3302-B, mycosubtilin, patellamide A, duramycin (and cinnamycin), bottoromycin A 2, A19009, galantin I, vancomycin, stenothricin, calf speleen profilin, neocarzinostatin, pancreatic spasmolytic polypeptide, cerebratulus toxin B-IV, RNAase U 2, ferredoxin ...

  1. Cationic antimicrobial peptides inactivate Shiga toxin-encoding bacteriophages

    Science.gov (United States)

    Del Cogliano, Manuel E.; Hollmann, Axel; Martinez, Melina; Semorile, Liliana; Ghiringhelli, Pablo D.; Maffía, Paulo C.; Bentancor, Leticia V.

    2017-12-01

    Shiga toxin (Stx) is the principal virulence factor during Shiga toxin-producing Escherichia coli (STEC) infections. We have previously reported the inactivation of bacteriophage encoding Stx after treatment with chitosan, a linear polysaccharide polymer with cationic properties. Cationic antimicrobial peptides (cAMPs) are short linear aminoacidic sequences, with a positive net charge, which display bactericidal or bacteriostatic activity against a wide range of bacterial species. They are promising novel antibiotics since they have shown bactericidal effects against multiresistant bacteria. To evaluate whether cationic properties are responsible for bacteriophage inactivation, we tested seven cationic peptides with proven antimicrobial activity as anti-bacteriophage agents, and one random sequence cationic peptide with no antimicrobial activity as a control. We observed bacteriophage inactivation after incubation with five cAMPs, but no inactivating activity was observed with the random sequence cationic peptide or with the non alpha helical cAMP Omiganan. Finally, to confirm peptide-bacteriophage interaction, zeta potential was analyzed by following changes on bacteriophage surface charges after peptide incubation. According to our results we could propose that: 1) direct interaction of peptides with phage is a necessary step for bacteriophage inactivation, 2) cationic properties are necessary but not sufficient for bacteriophage inactivation, and 3) inactivation by cationic peptides could be sequence (or structure) specific. Overall our data suggest that these peptides could be considered a new family of molecules potentially useful to decrease bacteriophage replication and Stx expression.

  2. Cationic Antimicrobial Peptides Inactivate Shiga Toxin-Encoding Bacteriophages

    Directory of Open Access Journals (Sweden)

    Manuel E. Del Cogliano

    2017-12-01

    Full Text Available Shiga toxin (Stx is the principal virulence factor during Shiga toxin-producing Escherichia coli (STEC infections. We have previously reported the inactivation of bacteriophage encoding Stx after treatment with chitosan, a linear polysaccharide polymer with cationic properties. Cationic antimicrobial peptides (cAMPs are short linear aminoacidic sequences, with a positive net charge, which display bactericidal or bacteriostatic activity against a wide range of bacterial species. They are promising novel antibiotics since they have shown bactericidal effects against multiresistant bacteria. To evaluate whether cationic properties are responsible for bacteriophage inactivation, we tested seven cationic peptides with proven antimicrobial activity as anti-bacteriophage agents, and one random sequence cationic peptide with no antimicrobial activity as a control. We observed bacteriophage inactivation after incubation with five cAMPs, but no inactivating activity was observed with the random sequence cationic peptide or with the non-alpha helical cAMP Omiganan. Finally, to confirm peptide-bacteriophage interaction, zeta potential was analyzed by following changes on bacteriophage surface charges after peptide incubation. According to our results we could propose that: (1 direct interaction of peptides with phage is a necessary step for bacteriophage inactivation, (2 cationic properties are necessary but not sufficient for bacteriophage inactivation, and (3 inactivation by cationic peptides could be sequence (or structure specific. Overall our data suggest that these peptides could be considered a new family of molecules potentially useful to decrease bacteriophage replication and Stx expression.

  3. Use of short tandem repeat sequences to study Mycobacterium leprae in leprosy patients in Malawi and India.

    Directory of Open Access Journals (Sweden)

    Saroj K Young

    2008-04-01

    Full Text Available Inadequate understanding of the transmission of Mycobacterium leprae makes it difficult to predict the impact of leprosy control interventions. Genotypic tests that allow tracking of individual bacterial strains would strengthen epidemiological studies and contribute to our understanding of the disease.Genotyping assays based on variation in the copy number of short tandem repeat sequences were applied to biopsies collected in population-based epidemiological studies of leprosy in northern Malawi, and from members of multi-case households in Hyderabad, India. In the Malawi series, considerable genotypic variability was observed between patients, and also within patients, when isolates were collected at different times or from different tissues. Less within-patient variability was observed when isolates were collected from similar tissues at the same time. Less genotypic variability was noted amongst the closely related Indian patients than in the Malawi series.Lineages of M. leprae undergo changes in their pattern of short tandem repeat sequences over time. Genetic divergence is particularly likely between bacilli inhabiting different (e.g., skin and nerve tissues. Such variability makes short tandem repeat sequences unsuitable as a general tool for population-based strain typing of M. leprae, or for distinguishing relapse from reinfection. Careful use of these markers may provide insights into the development of disease within individuals and for tracking of short transmission chains.

  4. Identification of a haemolysin-like peptide with antibacterial activity using the draft genome sequence of Staphylococcus epidermidis strain A487.

    Science.gov (United States)

    Al-Mahrous, Mohammed M; Jack, Ralph W; Sandiford, Stephanie K; Tagg, John R; Beatson, Scott A; Upton, Mathew

    2011-08-01

    Our interest in Staphylococcus epidermidis strain A487 was prompted by the unusual nature of its inhibitory activity in screening tests against methicillin-resistant Staphylococcus aureus isolates. The inhibitory activity was detected in deferred antagonism tests only if the agar plate was preheated for at least 35 min at ≥ 55 °C before inoculation of the indicator bacteria, this phenomenon indicating possible involvement of a heat-labile immunity agent or protease. The inhibitor was purified to homogeneity by ammonium sulphate precipitation, followed by cation-exchange and reversed-phase chromatography. Tandem MS revealed a novel peptide of molecular weight 2588.4 Da. The draft genome sequence of strain A487 was determined using 454 GS FLX technology, allowing the identification of the structural gene (hlp) encoding the mature peptide MQFITDLIKKAVDFFKGLFGNK. The deduced amino acid sequence of peptide 487 exhibited 70.8% similarity to that of a putative haemolysin from Staphylococcus cohnii. Analysis of the genome of strain A487 showed several additional inhibitor-encoding genes, including hld, the determinant for staphylococcal δ-lysin. This work indicates that potentially useful inhibitors could be overlooked in agar-based inhibitor screening programmes lacking a heat pretreatment step and also highlights the utility of draft genome sequence examination in antibacterial agent discovery. © 2011 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

  5. Anticancer activities of bovine and human lactoferricin-derived peptides.

    Science.gov (United States)

    Arias, Mauricio; Hilchie, Ashley L; Haney, Evan F; Bolscher, Jan G M; Hyndman, M Eric; Hancock, Robert E W; Vogel, Hans J

    2017-02-01

    Lactoferrin (LF) is a mammalian host defense glycoprotein with diverse biological activities. Peptides derived from the cationic region of LF possess cytotoxic activity against cancer cells in vitro and in vivo. Bovine lactoferricin (LFcinB), a peptide derived from bovine LF (bLF), exhibits broad-spectrum anticancer activity, while a similar peptide derived from human LF (hLF) is not as active. In this work, several peptides derived from the N-terminal regions of bLF and hLF were studied for their anticancer activities against leukemia and breast-cancer cells, as well as normal peripheral blood mononuclear cells. The cyclized LFcinB-CLICK peptide, which possesses a stable triazole linkage, showed improved anticancer activity, while short peptides hLF11 and bLF10 were not cytotoxic to cancer cells. Interestingly, hLF11 can act as a cell-penetrating peptide; when combined with the antimicrobial core sequence of LFcinB (RRWQWR) through either a Pro or Gly-Gly linker, toxicity to Jurkat cells increased. Together, our work extends the library of LF-derived peptides tested for anticancer activity, and identified new chimeric peptides with high cytotoxicity towards cancerous cells. Additionally, these results support the notion that short cell-penetrating peptides and antimicrobial peptides can be combined to create new adducts with increased potency.

  6. Peptide and nucleotide sequences of rat CD4 (W3/25) antigen: evidence for derivation from a structure with four immunoglobulin-related domains

    International Nuclear Information System (INIS)

    Clark, S.J.; Jefferies, W.A.; Barclay, A.N.; Gagnon, J.; Williams, A.F.

    1987-01-01

    The rat W3/25 antigen was the first marker antigen of helper T lymphocytes to be identified. Subsequently, the human OKT4 antigen (now called CD4) was described, and cell distribution and functional data suggested that W3/25 and OKT4 antigens were homologous. This is now confirmed by the matching of peptide sequences from W3/25 antigen with sequence predicted from rat cDNA clones detected by cross-hybridization with a cDNA probe for human CD4. Analysis of the two sequences suggests an evolutionary origin from a structure with four immunoglobulin-related domains, although only domain 1 at the NH 2 terminus meets the standard criteria for an immunoglobulin-related sequence. CD4 domains 2 and 4 contain disulfide bonds but seem like truncated immunoglobulin domains, whereas domain 3 may have a pattern of β-strands like an immunoglobulin variable domain, but without the disulfide bond

  7. Optimization of antibacterial peptides by genetic algorithms and cheminformatics

    DEFF Research Database (Denmark)

    Fjell, Christopher D.; Jenssen, Håvard; Cheung, Warren A.

    2011-01-01

    Pathogens resistant to available drug therapies are a pressing global health problem. Short, cationic peptides represent a novel class of agents that have lower rates of drug resistance than derivatives of current antibiotics. Previously, we created a software system utilizing artificial neural...... 47 of the top rated 50 peptides chosen from an in silico library of nearly 100 000 sequences. Here, we report a method of generating candidate peptide sequences using the heuristic evolutionary programming method of genetic algorithms (GA), which provided a large (19-fold) improvement...

  8. Isolation and amino acid sequence of a short-chain neurotoxin from an Australian elapid snake, Pseudechis australis.

    OpenAIRE

    Takasaki, C; Tamiya, N

    1985-01-01

    A short-chain neurotoxin Pseudechis australis a (toxin Pa a) was isolated from the venom of an Australian elapid snake Pseudechis australis (king brown snake) by sequential chromatography on CM-cellulose, Sephadex G-50 and CM-cellulose columns. Toxin Pa a has an LD50 (intravenous) value of 76 micrograms/kg body wt. in mice and consists of 62 amino acid residues. The amino acid sequence of Pa a shows considerable homology with those of short-chain neurotoxins of elapid snakes, especially of tr...

  9. Recurrent Neural Network Model for Constructive Peptide Design.

    Science.gov (United States)

    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.

  10. Identification of snake bradykinin-potentiating peptides (BPPs)-simile sequences in rat brain--Potential BPP-like precursor protein?

    Science.gov (United States)

    Campeiro, Joana D'Arc; Neshich, Izabella P; Sant'Anna, Osvaldo A; Lopes, Robson; Ianzer, Danielle; Assakura, Marina T; Neshich, Goran; Hayashi, Mirian A F

    2015-08-01

    Bradykinin-potentiating peptides (BPPs) from the South American pit viper snake venom were the first natural inhibitors of the human angiotensin I-converting enzyme (ACE) described. The pioneer characterization of the BPPs precursor from the snake venom glands by our group showed for the first time the presence of the C-type natriuretic peptide (CNP) in this same viper precursor protein. The confirmation of the BPP/CNP expression in snake brain regions correlated with neuroendocrine functions stimulated us to pursue the physiological correlates of these vasoactive peptides in mammals. Notably, several snake toxins were shown to have endogenous physiological correlates in mammals. In the present work, we expressed in bacteria the BPPs domain of the snake venom gland precursor protein, and this purified recombinant protein was used to raise specific polyclonal anti-BPPs antibodies. The correspondent single protein band immune-recognized in adult rat brain cytosol was isolated by 2D-SDS/PAGE and/or HPLC, before characterization by MS fingerprint analysis, which identified this protein as superoxide dismutase (SOD, EC 1.15.1.1), a classically known enzyme with antioxidant activity and important roles in the blood pressure modulation. In silico analysis showed the exposition of the BPP-like peptide sequences on the surface of the 3D structure of rat SOD. These peptides were chemically synthesized to show the BPP-like biological activities in ex vivo and in vivo pharmacological bioassays. Taken together, our data suggest that SOD protein have the potential to be a source for putative BPP-like bioactive peptides, which once released may contribute to the blood pressure control in mammals. Copyright © 2015 Elsevier Inc. All rights reserved.

  11. The differences in heparin binding for the C-terminal basic-sequence-rich peptides of HPV-16 and HPV-18 capsid protein L1

    International Nuclear Information System (INIS)

    Sun Jian; Yu Jisheng; Yu Zhiwu; Zha Xiao; Wu Yuqing

    2012-01-01

    Graphial abstract: The differences in heparin binding for the C-terminal basic-sequence-rich peptides of HPV-16 and HPV-18 capsid protein L1. Highlights: ► Several driving forces contribute to the interaction between heparin and peptides. ► C-terminal of HPV L1 is a potential candidate for the attachment to host cells. ► The C-terminal peptides of HPV-16 and -18 L1 have different heparin-binding. ► The different heparin-binding provides an explanation for the distinct prevalences. - Abstract: The high-risk types of human papillomaviruses (HPV) HPV-16 and -18 are the predominant types associated with cervical cancer. HPV-16 and -18 account for about 50% and 20%, respectively, of cervical cancers worldwide. While the reason and molecular mechanism of the distinct prevalence and distributions between them remain poorly understood, the binding affinity of cell surface receptor with capsid proteins, especially L1, may be involved. We examined heparin binding with two synthetic peptides corresponding to the 14 amino acid C-terminal peptides of HPV-16 and -18 L1 with the goal of comparing the equivalent residues in different HPV types. Using isothermal titration calorimetry (ITC) and static right-angle light scattering (SLS), we determined the binding constant K, reaction enthalpy ΔH, and other thermodynamic parameters in the interaction. Especially, we assessed the role of specific residues in binding with heparin by comparing the NMR spectra of free and heparin-bound peptides.

  12. Short Interspersed Nuclear Element (SINE Sequences in the Genome of the Human Pathogenic Fungus Aspergillus fumigatus Af293.

    Directory of Open Access Journals (Sweden)

    Lakkhana Kanhayuwa

    Full Text Available Novel families of short interspersed nuclear element (SINE sequences in the human pathogenic fungus Aspergillus fumigatus, clinical isolate Af293, were identified and categorised into tRNA-related and 5S rRNA-related SINEs. Eight predicted tRNA-related SINE families originating from different tRNAs, and nominated as AfuSINE2 sequences, contained target site duplications of short direct repeat sequences (4-14 bp flanking the elements, an extended tRNA-unrelated region and typical features of RNA polymerase III promoter sequences. The elements ranged in size from 140-493 bp and were present in low copy number in the genome and five out of eight were actively transcribed. One putative tRNAArg-derived sequence, AfuSINE2-1a possessed a unique feature of repeated trinucleotide ACT residues at its 3'-terminus. This element was similar in sequence to the I-4_AO element found in A. oryzae and an I-1_AF long nuclear interspersed element-like sequence identified in A. fumigatus Af293. Families of 5S rRNA-related SINE sequences, nominated as AfuSINE3, were also identified and their 5'-5S rRNA-related regions show 50-65% and 60-75% similarity to respectively A. fumigatus 5S rRNAs and SINE3-1_AO found in A. oryzae. A. fumigatus Af293 contains five copies of AfuSINE3 sequences ranging in size from 259-343 bp and two out of five AfuSINE3 sequences were actively transcribed. Investigations on AfuSINE distribution in the fungal genome revealed that the elements are enriched in pericentromeric and subtelomeric regions and inserted within gene-rich regions. We also demonstrated that some, but not all, AfuSINE sequences are targeted by host RNA silencing mechanisms. Finally, we demonstrated that infection of the fungus with mycoviruses had no apparent effects on SINE activity.

  13. Short Interspersed Nuclear Element (SINE) Sequences in the Genome of the Human Pathogenic Fungus Aspergillus fumigatus Af293.

    Science.gov (United States)

    Kanhayuwa, Lakkhana; Coutts, Robert H A

    2016-01-01

    Novel families of short interspersed nuclear element (SINE) sequences in the human pathogenic fungus Aspergillus fumigatus, clinical isolate Af293, were identified and categorised into tRNA-related and 5S rRNA-related SINEs. Eight predicted tRNA-related SINE families originating from different tRNAs, and nominated as AfuSINE2 sequences, contained target site duplications of short direct repeat sequences (4-14 bp) flanking the elements, an extended tRNA-unrelated region and typical features of RNA polymerase III promoter sequences. The elements ranged in size from 140-493 bp and were present in low copy number in the genome and five out of eight were actively transcribed. One putative tRNAArg-derived sequence, AfuSINE2-1a possessed a unique feature of repeated trinucleotide ACT residues at its 3'-terminus. This element was similar in sequence to the I-4_AO element found in A. oryzae and an I-1_AF long nuclear interspersed element-like sequence identified in A. fumigatus Af293. Families of 5S rRNA-related SINE sequences, nominated as AfuSINE3, were also identified and their 5'-5S rRNA-related regions show 50-65% and 60-75% similarity to respectively A. fumigatus 5S rRNAs and SINE3-1_AO found in A. oryzae. A. fumigatus Af293 contains five copies of AfuSINE3 sequences ranging in size from 259-343 bp and two out of five AfuSINE3 sequences were actively transcribed. Investigations on AfuSINE distribution in the fungal genome revealed that the elements are enriched in pericentromeric and subtelomeric regions and inserted within gene-rich regions. We also demonstrated that some, but not all, AfuSINE sequences are targeted by host RNA silencing mechanisms. Finally, we demonstrated that infection of the fungus with mycoviruses had no apparent effects on SINE activity.

  14. Hybridization Capture Using Short PCR Products Enriches Small Genomes by Capturing Flanking Sequences (CapFlank)

    DEFF Research Database (Denmark)

    Tsangaras, Kyriakos; Wales, Nathan; Sicheritz-Pontén, Thomas

    2014-01-01

    nucleotides) can result in enrichment across entire mitochondrial and bacterial genomes. Our findings suggest that some of the off-target sequences derived in capture experiments are non-randomly enriched, and that CapFlank will facilitate targeted enrichment of large contiguous sequences with minimal prior...

  15. Genomic Sequence of a Ranavirus Isolated from Short-Finned Eel (Anguilla australis)

    DEFF Research Database (Denmark)

    Subramaniam, Kuttichantran; Toffan, Anna; Cappellozza, Elisabetta

    2016-01-01

    The short-finned eel ranavirus (SERV) was isolated from short-finned eel imported to Italy from New Zealand. Phylogenomic analyses revealed that SERV is a unique member of the genus Ranavirus, family Iridoviridae, branching at the base of the tree near other fish ranaviruses....

  16. Short bowel patients treated for two years with glucagon-like peptide 2 (GLP-2): compliance, safety, and effects on quality of life

    DEFF Research Database (Denmark)

    Jeppesen, P B; Lund, P; Gottschalck, I B

    2009-01-01

    BACKGROUND AND AIMS: Glucagon-like peptide 2 (GLP-2) has been shown to improve intestinal absorption in short bowel syndrome (SBS) patients in a short-term study. This study describes safety, compliance, and changes in quality of life in 11 SBS patients at baseline, week 13, 26, and 52 during two...... years of subcutaneous GLP-2 treatment, 400 microgram TID, intermitted by an 8-week washout period. METHODS: Safety and compliance was evaluated during the admissions. The Sickness Impact Profile (SIP), Short Form 36 (SF 36), and Inflammatory Bowel Disease Questionnaire (IBDQ) evaluated quality of life......-ascendo-anastomosis. The investigator excluded a patient due to unreliable feedback. Stoma nipple enlargement was seen in all 9 jejunostomy patients. Reported GLP-2 compliance was excellent (>93%). GLP-2 improved the overall quality of life VAS-score (4.1 +/- 2.8 cm versus 6.0 +/- 2.4 cm, P

  17. BarraCUDA - a fast short read sequence aligner using graphics processing units

    Directory of Open Access Journals (Sweden)

    Klus Petr

    2012-01-01

    Full Text Available Abstract Background With the maturation of next-generation DNA sequencing (NGS technologies, the throughput of DNA sequencing reads has soared to over 600 gigabases from a single instrument run. General purpose computing on graphics processing units (GPGPU, extracts the computing power from hundreds of parallel stream processors within graphics processing cores and provides a cost-effective and energy efficient alternative to traditional high-performance computing (HPC clusters. In this article, we describe the implementation of BarraCUDA, a GPGPU sequence alignment software that is based on BWA, to accelerate the alignment of sequencing reads generated by these instruments to a reference DNA sequence. Findings Using the NVIDIA Compute Unified Device Architecture (CUDA software development environment, we ported the most computational-intensive alignment component of BWA to GPU to take advantage of the massive parallelism. As a result, BarraCUDA offers a magnitude of performance boost in alignment throughput when compared to a CPU core while delivering the same level of alignment fidelity. The software is also capable of supporting multiple CUDA devices in parallel to further accelerate the alignment throughput. Conclusions BarraCUDA is designed to take advantage of the parallelism of GPU to accelerate the alignment of millions of sequencing reads generated by NGS instruments. By doing this, we could, at least in part streamline the current bioinformatics pipeline such that the wider scientific community could benefit from the sequencing technology. BarraCUDA is currently available from http://seqbarracuda.sf.net

  18. BarraCUDA - a fast short read sequence aligner using graphics processing units

    LENUS (Irish Health Repository)

    Klus, Petr

    2012-01-13

    Abstract Background With the maturation of next-generation DNA sequencing (NGS) technologies, the throughput of DNA sequencing reads has soared to over 600 gigabases from a single instrument run. General purpose computing on graphics processing units (GPGPU), extracts the computing power from hundreds of parallel stream processors within graphics processing cores and provides a cost-effective and energy efficient alternative to traditional high-performance computing (HPC) clusters. In this article, we describe the implementation of BarraCUDA, a GPGPU sequence alignment software that is based on BWA, to accelerate the alignment of sequencing reads generated by these instruments to a reference DNA sequence. Findings Using the NVIDIA Compute Unified Device Architecture (CUDA) software development environment, we ported the most computational-intensive alignment component of BWA to GPU to take advantage of the massive parallelism. As a result, BarraCUDA offers a magnitude of performance boost in alignment throughput when compared to a CPU core while delivering the same level of alignment fidelity. The software is also capable of supporting multiple CUDA devices in parallel to further accelerate the alignment throughput. Conclusions BarraCUDA is designed to take advantage of the parallelism of GPU to accelerate the alignment of millions of sequencing reads generated by NGS instruments. By doing this, we could, at least in part streamline the current bioinformatics pipeline such that the wider scientific community could benefit from the sequencing technology. BarraCUDA is currently available from http:\\/\\/seqbarracuda.sf.net

  19. Binding of cholesterol and inhibitory peptide derivatives with the fusogenic hydrophobic sequence of F-glycoprotein of HVJ (Sendai virus): possible implication in the fusion reaction

    International Nuclear Information System (INIS)

    Asano, K.; Asano, A.

    1988-01-01

    Specificity of the binding of sterols and related compounds with purified F-protein (fusion protein) of the HVJ (Sendai virus) was studied by binding competition with [ 3 H] cholesterol. Requirement for cholesterol or the A/B ring trans structure and nonrequirement for the 3-hydroxyl group were found in this binding. Binding of 125 I-labeled Z-Phe-Tyr, an inhibitory peptide of viral membrane-cell membrane fusion, was studied by using purified proteins and virions. F-Protein and virions showed a specific binding with the peptide, whereas the result was negative with hemagglutinin and neuraminidase protein. Thermolysin-truncated F-protein (an F-protein derivative deprived of a 2.5-kDa fragment from the N-terminal of the F 1 subunit and without fusogenic activity) exhibited a considerably diminished binding ability both to cholesterol and to inhibitory peptides. Therefore, the N-terminal hydrophobic sequence that was previously assigned as fusogenic seems to be the binding site of these molecules. In support of this, the binding of cholesterol with F-protein was inhibited by Z-Phe-Tyr and other fusion inhibitory peptides, whereas it was not affected with non-fusion-inhibitory Z-Gly-Phe. These results are discussed in relation to the notion that the binding of the N-terminal portion of the F 1 subunit of F-protein with cholesterol in the target cell membranes facilitiates the fusion reaction

  20. A Short Peptide That Mimics the Binding Domain of TGF-β1 Presents Potent Anti-Inflammatory Activity.

    Directory of Open Access Journals (Sweden)

    Emília R Vaz

    Full Text Available The transforming growth factor beta 1 (TGF-β1 is a pleiotropic cytokine with multiple roles in development, wound healing, and immune regulation. TGF-β1-mediated immune dysfunction may lead to pathological conditions, such as inflammation. Chronic inflammatory process is characterized by a continuous release of pro-inflammatory cytokines, and the inhibition or the blockage of these cytokines signaling pathways are considered a target treatment. In this context, despite the high numbers of TGF-β-targeted pathways, the inducible regulatory T cells (iTreg to control inflammation seems to be a promising approach. Our aim was to develop novel peptides through phage display (PhD technology that could mimic TGF-β1 function with higher potency. Specific mimetic peptides were obtained through a PhD subtraction strategy from whole cell binding using TGF-β1 recombinant as a competitor during elution step. We have selected a peptide that seems to play an important role on cellular differentiation and modulation of TNF-α and IL-10 cytokines. The synthetic pm26TGF-β1 peptide tested in PBMC significantly down-modulated TNF-α and up-regulated IL-10 responses, leading to regulatory T cells (Treg phenotype differentiation. Furthermore, the synthetic peptide was able to decrease leukocytes rolling in BALB/C mice and neutrophils migration during inflammatory process in C57BL/6 mice. These data suggest that this peptide may be useful for the treatment of inflammatory diseases, especially because it displays potent anti-inflammatory properties and do not exhibit neutrophils' chemoattraction.

  1. Musicians' and nonmusicians' short-term memory for verbal and musical sequences: comparing phonological similarity and pitch proximity.

    Science.gov (United States)

    Williamson, Victoria J; Baddeley, Alan D; Hitch, Graham J

    2010-03-01

    Language-music comparative studies have highlighted the potential for shared resources or neural overlap in auditory short-term memory. However, there is a lack of behavioral methodologies for comparing verbal and musical serial recall. We developed a visual grid response that allowed both musicians and nonmusicians to perform serial recall of letter and tone sequences. The new method was used to compare the phonological similarity effect with the impact of an operationalized musical equivalent-pitch proximity. Over the course of three experiments, we found that short-term memory for tones had several similarities to verbal memory, including limited capacity and a significant effect of pitch proximity in nonmusicians. Despite being vulnerable to phonological similarity when recalling letters, however, musicians showed no effect of pitch proximity, a result that we suggest might reflect strategy differences. Overall, the findings support a limited degree of correspondence in the way that verbal and musical sounds are processed in auditory short-term memory.

  2. Sorting of a HaloTag protein that has only a signal peptide sequence into exocrine secretory granules without protein aggregation.

    Science.gov (United States)

    Fujita-Yoshigaki, Junko; Matsuki-Fukushima, Miwako; Yokoyama, Megumi; Katsumata-Kato, Osamu

    2013-11-15

    The mechanism involved in the sorting and accumulation of secretory cargo proteins, such as amylase, into secretory granules of exocrine cells remains to be solved. To clarify that sorting mechanism, we expressed a reporter protein HaloTag fused with partial sequences of salivary amylase protein in primary cultured parotid acinar cells. We found that a HaloTag protein fused with only the signal peptide sequence (Met(1)-Ala(25)) of amylase, termed SS25H, colocalized well with endogenous amylase, which was confirmed by immunofluorescence microscopy. Percoll-density gradient centrifugation of secretory granule fractions shows that the distributions of amylase and SS25H were similar. These results suggest that SS25H is transported to secretory granules and is not discriminated from endogenous amylase by the machinery that functions to remove proteins other than granule cargo from immature granules. Another reporter protein, DsRed2, that has the same signal peptide sequence also colocalized with amylase, suggesting that the sorting to secretory granules is not dependent on a characteristic of the HaloTag protein. Whereas Blue Native PAGE demonstrates that endogenous amylase forms a high-molecular-weight complex, SS25H does not participate in the complex and does not form self-aggregates. Nevertheless, SS25H was released from cells by the addition of a β-adrenergic agonist, isoproterenol, which also induces amylase secretion. These results indicate that addition of the signal peptide sequence, which is necessary for the translocation in the endoplasmic reticulum, is sufficient for the transportation and storage of cargo proteins in secretory granules of exocrine cells.

  3. Disruption of the Putative Vascular Leak Peptide Sequence in the Stabilized Ricin Vaccine Candidate RTA1-33/44-198

    Directory of Open Access Journals (Sweden)

    Charles B. Millard

    2013-01-01

    Full Text Available Vitetta and colleagues identified and characterized a putative vascular leak peptide (VLP consensus sequence in recombinant ricin toxin A-chain (RTA that contributed to dose-limiting human toxicity when RTA was administered intravenously in large quantities during chemotherapy. We disrupted this potentially toxic site within the more stable RTA1-33/44-198 vaccine immunogen and determined the impact of these mutations on protein stability, structure and protective immunogenicity using an experimental intranasal ricin challenge model in BALB/c mice to determine if the mutations were compatible. Single amino acid substitutions at the positions corresponding with RTA D75 (to A, or N and V76 (to I, or M had minor effects on the apparent protein melting temperature of RTA1-33/44-198 but all four variants retained greater apparent stability than the parent RTA. Moreover, each VLP(− variant tested provided protection comparable with that of RTA1-33/44-198 against supralethal intranasal ricin challenge as judged by animal survival and several biomarkers. To understand better how VLP substitutions and mutations near the VLP site impact epitope structure, we introduced a previously described thermal stabilizing disulfide bond (R48C/T77C along with the D75N or V76I substitutions in RTA1-33/44-198. The D75N mutation was compatible with the adjacent stabilizing R48C/T77C disulfide bond and the Tm was unaffected, whereas the V76I mutation was less compatible with the adjacent disulfide bond involving C77. A crystal structure of the RTA1-33/44-198 R48C/T77C/D75N variant showed that the structural integrity of the immunogen was largely conserved and that a stable immunogen could be produced from E. coli. We conclude that it is feasible to disrupt the VLP site in RTA1-33/44-198 with little or no impact on apparent protein stability or protective efficacy in mice and such variants can be stabilized further by introduction of a disulfide bond.

  4. Antimicrobial Activity and Stability of Short and Long Based Arachnid Synthetic Peptides in the Presence of Commercial Antibiotics.

    Science.gov (United States)

    Arenas, Ivan; Villegas, Elba; Walls, Oliver; Barrios, Humberto; Rodríguez, Ramon; Corzo, Gerardo

    2016-02-17

    Four antimicrobial peptides (AMPs) named Pin2[G], Pin2[14], P18K and FA1 were chemically synthesized and purified. The four peptides were evaluated in the presence of eight commercial antibiotics against four microorganisms of medical importance: Escherichia coli, Staphylococcus aureus, Pseudomonas aeruginosa, and Klebsiella pneumoniae. The commercial antibiotics used were amoxicillin, azithromycin, ceftriaxone, gentamicin, levofloxacin, sulfamethoxazole, trimethoprim and vancomycin. The best AMP against P. aeruginosa was the peptide FA1, and the best AMP against S. aureus was Pin2[G]. Both FA1 and Pin2[G] were efficient against E. coli, but they were not effective against K. pneumoniae. As K. pneumoniae was resistant to most of the commercial antibiotics, combinations of the AMPs FA1 and Pin2[G] were prepared with these antibiotics. According to the fractional inhibitory concentration (FIC) index, the best antimicrobial combinations were obtained with concomitant applications of mixtures of FA1 with levofloxacin and sulfamethoxazole. However, combinations of FA1 or Pin2[G] with other antibiotics showed that total inhibitory effect of the combinations were greater than the sum of the individual effects of either the antimicrobial peptide or the antibiotic. We also evaluated the stability of the AMPs. The AMP Pin2[G] manifested the best performance in saline buffer, in supernatants of bacterial growth and in human blood plasma. Nevertheless, all AMPs were cleaved using endoproteolytic enzymes. These data show advantages and disadvantages of AMPs for potential clinical treatments of bacterial infections, using them in conjunction with commercial antibiotics.

  5. Glucagon-like peptide 2 improves nutrient absorption and nutritional status in short-bowel patients with no colon

    DEFF Research Database (Denmark)

    Jeppesen, P B; Hartmann, B; Thulesen, J

    2001-01-01

    Glucagon-like peptide 2 (GLP-2) is intestinotrophic, antisecretory, and transit-modulating in rodents, and it is mainly secreted from the intestinal mucosa of the terminal ileum and colon after food ingestion. We assessed the effect of GLP-2 on the gastrointestinal function in patients without a ...

  6. Role of solution conformation and flexibility of short peptide ligands that bind to the p56(lck) SH2 domain

    NARCIS (Netherlands)

    Dekker, Frank J; de Mol, Nico J; Bultinck, Patrick; Kemmink, Johan; Hilbers, Hans W; Liskamp, Rob M J; Dekker, Frank

    2003-01-01

    A general approach in drug design is making ligands more rigid in order to avoid loss in conformational entropy (deltaS(conf)) upon receptor binding. We hypothesized that in the high affinity binding of pYEEI peptide ligands to the p56(lck) SH2 domain this loss in deltaS(conf) might be diminished

  7. Synergistic effect of supplemental enteral nutrients and exogenous glucagon-like peptide 2 on intestinal adaptation in a rat model of short bowel syndrome

    DEFF Research Database (Denmark)

    Liu, Xiaowen; Nelson, David W; Holst, Jens Juul

    2006-01-01

    BACKGROUND: Short bowel syndrome (SBS) can lead to intestinal failure and require total or supplemental parenteral nutrition (TPN or PN, respectively). Glucagon-like peptide 2 (GLP-2) is a nutrient-dependent, proglucagon-derived gut hormone that stimulates intestinal adaptation. OBJECTIVE: Our...... objective was to determine whether supplemental enteral nutrients (SEN) modulate the intestinotrophic response to a low dose of GLP-2 coinfused with PN in a rat model of SBS (60% jejunoileal resection plus cecectomy). DESIGN: Rats were randomly assigned to 8 treatments by using a 2 x 2 x 2 factorial design...

  8. RNA-Seq analysis and gene discovery of Andrias davidianus using Illumina short read sequencing.

    Directory of Open Access Journals (Sweden)

    Fenggang Li

    Full Text Available The Chinese giant salamander, Andrias davidianus, is an important species in the course of evolution; however, there is insufficient genomic data in public databases for understanding its immunologic mechanisms. High-throughput transcriptome sequencing is necessary to generate an enormous number of transcript sequences from A. davidianus for gene discovery. In this study, we generated more than 40 million reads from samples of spleen and skin tissue using the Illumina paired-end sequencing technology. De novo assembly yielded 87,297 transcripts with a mean length of 734 base pairs (bp. Based on the sequence similarities, searching with known proteins, 38,916 genes were identified. Gene enrichment analysis determined that 981 transcripts were assigned to the immune system. Tissue-specific expression analysis indicated that 443 of transcripts were specifically expressed in the spleen and skin. Among these transcripts, 147 transcripts were found to be involved in immune responses and inflammatory reactions, such as fucolectin, β-defensins and lymphotoxin beta. Eight tissue-specific genes were selected for validation using real time reverse transcription quantitative PCR (qRT-PCR. The results showed that these genes were significantly more expressed in spleen and skin than in other tissues, suggesting that these genes have vital roles in the immune response. This work provides a comprehensive genomic sequence resource for A. davidianus and lays the foundation for future research on the immunologic and disease resistance mechanisms of A. davidianus and other amphibians.

  9. ReadDepth: a parallel R package for detecting copy number alterations from short sequencing reads.

    Directory of Open Access Journals (Sweden)

    Christopher A Miller

    2011-01-01

    Full Text Available Copy number alterations are important contributors to many genetic diseases, including cancer. We present the readDepth package for R, which can detect these aberrations by measuring the depth of coverage obtained by massively parallel sequencing of the genome. In addition to achieving higher accuracy than existing packages, our tool runs much faster by utilizing multi-core architectures to parallelize the processing of these large data sets. In contrast to other published methods, readDepth does not require the sequencing of a reference sample, and uses a robust statistical model that accounts for overdispersed data. It includes a method for effectively increasing the resolution obtained from low-coverage experiments by utilizing breakpoint information from paired end sequencing to do positional refinement. We also demonstrate a method for inferring copy number using reads generated by whole-genome bisulfite sequencing, thus enabling integrative study of epigenomic and copy number alterations. Finally, we apply this tool to two genomes, showing that it performs well on genomes sequenced to both low and high coverage. The readDepth package runs on Linux and MacOSX, is released under the Apache 2.0 license, and is available at http://code.google.com/p/readdepth/.

  10. Short-read reading-frame predictors are not created equal: sequence error causes loss of signal

    Directory of Open Access Journals (Sweden)

    Trimble William L

    2012-07-01

    Full Text Available Abstract Background Gene prediction algorithms (or gene callers are an essential tool for analyzing shotgun nucleic acid sequence data. Gene prediction is a ubiquitous step in sequence analysis pipelines; it reduces the volume of data by identifying the most likely reading frame for a fragment, permitting the out-of-frame translations to be ignored. In this study we evaluate five widely used ab initio gene-calling algorithms—FragGeneScan, MetaGeneAnnotator, MetaGeneMark, Orphelia, and Prodigal—for accuracy on short (75–1000 bp fragments containing sequence error from previously published artificial data and “real” metagenomic datasets. Results While gene prediction tools have similar accuracies predicting genes on error-free fragments, in the presence of sequencing errors considerable differences between tools become evident. For error-containing short reads, FragGeneScan finds more prokaryotic coding regions than does MetaGeneAnnotator, MetaGeneMark, Orphelia, or Prodigal. This improved detection of genes in error-containing fragments, however, comes at the cost of much lower (50% specificity and overprediction of genes in noncoding regions. Conclusions Ab initio gene callers offer a significant reduction in the computational burden of annotating individual nucleic acid reads and are used in many metagenomic annotation systems. For predicting reading frames on raw reads, we find the hidden Markov model approach in FragGeneScan is more sensitive than other gene prediction tools, while Prodigal, MGA, and MGM are better suited for higher-quality sequences such as assembled contigs.

  11. Synthesis of Mikto-Arm Star Peptide Conjugates.

    Science.gov (United States)

    Koo, Jin Mo; Su, Hao; Lin, Yi-An; Cui, Honggang

    2018-01-01

    Mikto-arm star peptide conjugates are an emerging class of self-assembling peptide-based structural units that contain three or more auxiliary segments of different chemical compositions and/or functionalities. This group of molecules exhibit interesting self-assembly behavior in solution due to their chemically asymmetric topology. Here we describe the detailed procedure for synthesis of an ABC Mikto-arm star peptide conjugate in which two immiscible entities (a saturated hydrocarbon and a hydrophobic and lipophobic fluorocarbon) are conjugated onto a short β-sheet forming peptide sequence, GNNQQNY, derived from the Sup35 prion, through a lysine junction. Automated and manual Fmoc-solid phase synthesis techniques are used to synthesize the Mikto-arm star peptide conjugates, followed by HPLC purification. We envision that this set of protocols can afford a versatile platform to synthesize a new class of peptidic building units for diverse applications.

  12. 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....

  13. B and T Cell Epitope-Based Peptides Predicted from Evolutionarily Conserved and Whole Protein Sequences of Ebola Virus as Vaccine Targets.

    Science.gov (United States)

    Yasmin, T; Nabi, A H M Nurun

    2016-05-01

    Ebola virus (EBV) has become a serious threat to public health. Different approaches were applied to predict continuous and discontinuous B cell epitopes as well as T cell epitopes from the sequence-based and available three-dimensional structural analyses of each protein of EBV. Peptides '(79) VPSATKRWGFRSGVPP(94) ' from GP1 and '(515) LHYWTTQDEGAAIGLA(530) ' from GP2 of Ebola were found to be the consensus peptidic sequences predicted as linear B cell epitope of which the latter contains a region (519) TTQDEG(524) that fulfilled all the criteria of accessibility, hydrophilicity, flexibility and beta turn region for becoming an ideal B cell epitope. Different nonamers as T cell epitopes were obtained that interacted with different numbers of MHC class I and class II alleles with a binding affinity of <100 nm. Interestingly, these alleles also bound to the MHC class I alleles mostly prevalent in African and South Asian regions. Of these, 'LANETTQAL' and 'FLYDRLAST' nonamers were predicted to be the most potent T cell epitopes and they, respectively, interacted with eight and twelve class I alleles that covered 63.79% and 54.16% of world population, respectively. These nonamers were found to be the core sequences of 15mer peptides that interacted with the most common class II allele, HLA-DRB1*01:01. They were further validated for their binding to specific class I alleles using docking technique. Thus, these predicted epitopes may be used as vaccine targets against EBV and can be validated in model hosts to verify their efficacy as vaccine. © 2016 The Foundation for the Scandinavian Journal of Immunology.

  14. Temporal Clustering and Sequencing in Short-Term Memory and Episodic Memory

    Science.gov (United States)

    Farrell, Simon

    2012-01-01

    A model of short-term memory and episodic memory is presented, with the core assumptions that (a) people parse their continuous experience into episodic clusters and (b) items are clustered together in memory as episodes by binding information within an episode to a common temporal context. Along with the additional assumption that information…

  15. Sequence and expression pattern of a novel human orphan G-protein-coupled receptor, GPRC5B, a family C receptor with a short amino-terminal domain

    DEFF Research Database (Denmark)

    Bräuner-Osborne, Hans; Krogsgaard-Larsen, P

    2000-01-01

    Query of GenBank with the amino acid sequence of human metabotropic glutamate receptor subtype 2 (mGluR2) identified a predicted gene product of unknown function on BAC clone CIT987SK-A-69G12 (located on chromosome band 16p12) as a homologous protein. The transcript, entitled GPRC5B, was cloned f...... from an expressed sequence tag clone that contained the entire open reading frame of the transcript encoding a protein of 395 amino acids. Analysis of the protein sequence reveal that GPRC5B contains a signal peptide and seven transmembrane alpha-helices, which is a hallmark of G...

  16. Identification of a Short Cell-Penetrating Peptide from Bovine Lactoferricin for Intracellular Delivery of DNA in Human A549 Cells.

    Directory of Open Access Journals (Sweden)

    Betty R Liu

    Full Text Available Cell-penetrating peptides (CPPs have been shown to deliver cargos, including protein, DNA, RNA, and nanomaterials, in fully active forms into live cells. Most of the CPP sequences in use today are based on non-native proteins that may be immunogenic. Here we demonstrate that the L5a CPP (RRWQW from bovine lactoferricin (LFcin, stably and noncovalently complexed with plasmid DNA and prepared at an optimal nitrogen/phosphate ratio of 12, is able to efficiently enter into human lung cancer A549 cells. The L5a CPP delivered a plasmid containing the enhanced green fluorescent protein (EGFP coding sequence that was subsequently expressed in cells, as revealed by real-time PCR and fluorescent microscopy at the mRNA and protein levels, respectively. Treatment with calcium chloride increased the level of gene expression, without affecting CPP-mediated transfection efficiency. Zeta-potential analysis revealed that positively electrostatic interactions of CPP/DNA complexes correlated with CPP-mediated transport. The L5a and L5a/DNA complexes were not cytotoxic. This biomimetic LFcin L5a represents one of the shortest effective CPPs and could be a promising lead peptide with less immunogenic for DNA delivery in gene therapy.

  17. Identification of a Short Cell-Penetrating Peptide from Bovine Lactoferricin for Intracellular Delivery of DNA in Human A549 Cells.

    Science.gov (United States)

    Liu, Betty R; Huang, Yue-Wern; Aronstam, Robert S; Lee, Han-Jung

    2016-01-01

    Cell-penetrating peptides (CPPs) have been shown to deliver cargos, including protein, DNA, RNA, and nanomaterials, in fully active forms into live cells. Most of the CPP sequences in use today are based on non-native proteins that may be immunogenic. Here we demonstrate that the L5a CPP (RRWQW) from bovine lactoferricin (LFcin), stably and noncovalently complexed with plasmid DNA and prepared at an optimal nitrogen/phosphate ratio of 12, is able to efficiently enter into human lung cancer A549 cells. The L5a CPP delivered a plasmid containing the enhanced green fluorescent protein (EGFP) coding sequence that was subsequently expressed in cells, as revealed by real-time PCR and fluorescent microscopy at the mRNA and protein levels, respectively. Treatment with calcium chloride increased the level of gene expression, without affecting CPP-mediated transfection efficiency. Zeta-potential analysis revealed that positively electrostatic interactions of CPP/DNA complexes correlated with CPP-mediated transport. The L5a and L5a/DNA complexes were not cytotoxic. This biomimetic LFcin L5a represents one of the shortest effective CPPs and could be a promising lead peptide with less immunogenic for DNA delivery in gene therapy.

  18. Comparison Review of Short-Acting and Long-Acting Glucagon-like Peptide-1 Receptor Agonists

    OpenAIRE

    Uccellatore, Annachiara; Genovese, Stefano; Dicembrini, Ilaria; Mannucci, Edoardo; Ceriello, Antonio

    2015-01-01

    Glucagon-like peptide-1 (GLP-1) receptor agonists (GLP-1 RAs) are useful tools for treating type 2 diabetes mellitus. In their recent position statement, the American Diabetes Association and European Association for the Study of Diabetes recommend GLP1-RAs as add-on to metformin when therapeutic goals are not achieved with monotherapy, particularly for patients who wish to avoid weight gain or hypoglycemia. GLP1-RAs differ substantially in their duration of action, frequency of administratio...

  19. Antimicrobial Activity and Stability of Short and Long Based Arachnid Synthetic Peptides in the Presence of Commercial Antibiotics

    Directory of Open Access Journals (Sweden)

    Ivan Arenas

    2016-02-01

    Full Text Available Four antimicrobial peptides (AMPs named Pin2[G], Pin2[14], P18K and FA1 were chemically synthesized and purified. The four peptides were evaluated in the presence of eight commercial antibiotics against four microorganisms of medical importance: Escherichia coli, Staphylococcus aureus, Pseudomonas aeruginosa, and Klebsiella pneumoniae. The commercial antibiotics used were amoxicillin, azithromycin, ceftriaxone, gentamicin, levofloxacin, sulfamethoxazole, trimethoprim and vancomycin. The best AMP against P. aeruginosa was the peptide FA1, and the best AMP against S. aureus was Pin2[G]. Both FA1 and Pin2[G] were efficient against E. coli, but they were not effective against K. pneumoniae. As K. pneumoniae was resistant to most of the commercial antibiotics, combinations of the AMPs FA1 and Pin2[G] were prepared with these antibiotics. According to the fractional inhibitory concentration (FIC index, the best antimicrobial combinations were obtained with concomitant applications of mixtures of FA1 with levofloxacin and sulfamethoxazole. However, combinations of FA1 or Pin2[G] with other antibiotics showed that total inhibitory effect of the combinations were greater than the sum of the individual effects of either the antimicrobial peptide or the antibiotic. We also evaluated the stability of the AMPs. The AMP Pin2[G] manifested the best performance in saline buffer, in supernatants of bacterial growth and in human blood plasma. Nevertheless, all AMPs were cleaved using endoproteolytic enzymes. These data show advantages and disadvantages of AMPs for potential clinical treatments of bacterial infections, using them in conjunction with commercial antibiotics.

  20. Peptides in headlock ? a novel high-affinity and versatile peptide-binding nanobody for proteomics and microscopy

    OpenAIRE

    Braun, Michael B.; Traenkle, Bjoern; Koch, Philipp A.; Emele, Felix; Weiss, Frederik; Poetz, Oliver; Stehle, Thilo; Rothbauer, Ulrich

    2016-01-01

    Nanobodies are highly valuable tools for numerous bioanalytical and biotechnical applications. Here, we report the characterization of a nanobody that binds a short peptide epitope with extraordinary affinity. Structural analysis reveals an unusual binding mode where the extended peptide becomes part of a ?-sheet structure in the nanobody. This interaction relies on sequence-independent backbone interactions augmented by a small number of specificity-determining side chain contacts. Once boun...

  1. Detection of short repeated genomic sequences on metaphase chromosomes using padlock probes and target primed rolling circle DNA synthesis

    Directory of Open Access Journals (Sweden)

    Stougaard Magnus

    2007-11-01

    Full Text Available Abstract Background In situ detection of short sequence elements in genomic DNA requires short probes with high molecular resolution and powerful specific signal amplification. Padlock probes can differentiate single base variations. Ligated padlock probes can be amplified in situ by rolling circle DNA synthesis and detected by fluorescence microscopy, thus enhancing PRINS type reactions, where localized DNA synthesis reports on the position of hybridization targets, to potentially reveal the binding of single oligonucleotide-size probe molecules. Such a system has been presented for the detection of mitochondrial DNA in fixed cells, whereas attempts to apply rolling circle detection to metaphase chromosomes have previously failed, according to the literature. Methods Synchronized cultured cells were fixed with methanol/acetic acid to prepare chromosome spreads in teflon-coated diagnostic well-slides. Apart from the slide format and the chromosome spreading everything was done essentially according to standard protocols. Hybridization targets were detected in situ with padlock probes, which were ligated and amplified using target primed rolling circle DNA synthesis, and detected by fluorescence labeling. Results An optimized protocol for the spreading of condensed metaphase chromosomes in teflon-coated diagnostic well-slides was developed. Applying this protocol we generated specimens for target primed rolling circle DNA synthesis of padlock probes recognizing a 40 nucleotide sequence in the male specific repetitive satellite I sequence (DYZ1 on the Y-chromosome and a 32 nucleotide sequence in the repetitive kringle IV domain in the apolipoprotein(a gene positioned on the long arm of chromosome 6. These targets were detected with good efficiency, but the efficiency on other target sites was unsatisfactory. Conclusion Our aim was to test the applicability of the method used on mitochondrial DNA to the analysis of nuclear genomes, in particular as

  2. Far-UV-induced dimeric photoproducts in short oligonucleotides: sequence effects

    International Nuclear Information System (INIS)

    Douki, T.; Zalizniak, T.; Cadet, J.

    1997-01-01

    Cyclobutane pyrimidine dimers and pyrimidine (6-4)pyrimidone adducts represent the two major classes of far-UV-induced DNA photoproducts. Because of the lack of appropriate detection methods for each individual photoproduct, little is known about the effect of the sequence on their formaiton. In the present work, the photoproduct distribution obtained upon exposure of a series of dinucleoside monophosphate to 254 nm light was determined. (author)

  3. [Topographic mapping of retinal function with a scanning laser ophthalmoscope and multifocal electroretinography using short M-sequences].

    Science.gov (United States)

    Rudolph, G; Bechmann, M; Berninger, T; Kutschbach, E; Held, U; Tornow, R P; Kalpadakis, P; Zol'nikova, I V; Shamshinova, A M

    2001-01-01

    A new method of multifocal electroretinography making use of scanning laser ophthalmoscope with a wavelength of 630 nm (SLO-m-ERG), evoking short spatial visual stimuli on the retina, is proposed. Algorithm of presenting the visual stimuli and analysis of distribution of local electroretinograms on the surface of the retina is based on short m-sequences. Mathematical cross correlation analysis shows a three-dimensional distribution of bioelectrical activity of the retina in the central visual field. In normal subjects the cone bioelectrical activity is the maximum in the macular area (corresponding to the density of cone distribution) and absent in the blind spot. The method detects the slightest pathological changes in the retina under control of the site of stimulation and ophthalmoscopic picture of the fundus oculi. The site of the pathological process correlates with the topography of changes in bioelectrical activity of the examined retinal area in diseases of the macular area and pigmented retinitis detectable by ophthalmoscopy.

  4. Analysing breast tissue composition with MRI using currently available short, simple sequences

    International Nuclear Information System (INIS)

    Chau, A.C.M.; Hua, J.; Taylor, D.B.

    2016-01-01

    Aim: To determine the most robust commonly available magnetic resonance imaging (MRI) sequence to quantify breast tissue composition at 1.5 T. Materials and methods: Two-dimensional (2D) T1-weighted, Dixon fat, Dixon water and SPAIR images were obtained from five participants and a breast phantom using a 1.5 T Siemens Aera MRI system. Manual segmentation of the breasts was performed, and an in-house computer program was used to generate signal intensity histograms. Relative trough depth and relative peak separation were used to determine the robustness of the images for quantifying the two breast tissues. Total breast volumes and percentage breast densities calculated using the four sequences were compared. Results: Dixon fat histograms had consistently low relative trough depth and relative peak separation compared to those obtained using other sequences. There was no significant difference in total breast volumes and percentage breast densities of the participants or breast phantom using Dixon fat and 2D T1-weighted histograms. Dixon water and SPAIR histograms were not suitable for quantifying breast tissue composition. Conclusion: Dixon fat images are the most robust for the quantification of breast tissue composition using a signal intensity histogram. - Highlights: • Signal intensity histogram analysis can determine robustness of images for quantification of breast tissue composition. • Dixon fat images are the most robust. • The characteristics of the signal intensity histograms from Dixon water and SPAIR images make quantification unsuitable.

  5. [Short interspersed repetitive sequences (SINEs) and their use as a phylogenetic tool].

    Science.gov (United States)

    Kramerov, D A; Vasetskiĭ, N S

    2009-01-01

    The data on one of the most common repetitive elements of eukaryotic genomes, short interspersed elements (SINEs), are reviewed. Their structure, origin, and functioning in the genome are discussed. The variation and abundance of these neutral genomic markers makes them a convenient and reliable tool for phylogenetic analysis. The main methods of such analysis are presented, and the potential and limitations of this approach are discussed using specific examples.

  6. Whole Genome Sequencing Identifies a Missense Mutation in HES7 Associated with Short Tails in Asian Domestic Cats.

    Science.gov (United States)

    Xu, Xiao; Sun, Xin; Hu, Xue-Song; Zhuang, Yan; Liu, Yue-Chen; Meng, Hao; Miao, Lin; Yu, He; Luo, Shu-Jin

    2016-08-25

    Domestic cats exhibit abundant variations in tail morphology and serve as an excellent model to study the development and evolution of vertebrate tails. Cats with shortened and kinked tails were first recorded in the Malayan archipelago by Charles Darwin in 1868 and remain quite common today in Southeast and East Asia. To elucidate the genetic basis of short tails in Asian cats, we built a pedigree of 13 cats segregating at the trait with a founder from southern China and performed linkage mapping based on whole genome sequencing data from the pedigree. The short-tailed trait was mapped to a 5.6 Mb region of Chr E1, within which the substitution c. 5T > C in the somite segmentation-related gene HES7 was identified as the causal mutation resulting in a missense change (p.V2A). Validation in 245 unrelated cats confirmed the correlation between HES7-c. 5T > C and Chinese short-tailed feral cats as well as the Japanese Bobtail breed, indicating a common genetic basis of the two. In addition, some of our sampled kinked-tailed cats could not be explained by either HES7 or the Manx-related T-box, suggesting at least three independent events in the evolution of domestic cats giving rise to short-tailed traits.

  7. 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.

  8. A short autocomplementary sequence plays an essential role in avian sarcoma-leukosis virus RNA dimerization.

    Science.gov (United States)

    Fossé, P; Motté, N; Roumier, A; Gabus, C; Muriaux, D; Darlix, J L; Paoletti, J

    1996-12-24

    Retroviral genomes consist of two identical RNA molecules joined noncovalently near their 5'-ends. Recently, two models have been proposed for RNA dimer formation on the basis of results obtained in vitro with human immunodeficiency virus type 1 RNA and Moloney murine leukemia virus RNA. It was first proposed that viral RNA dimerizes by forming an interstrand quadruple helix with purine tetrads. The second model postulates that RNA dimerization is initiated by a loop-loop interaction between the two RNA molecules. In order to better characterize the dimerization process of retroviral genomic RNA, we analyzed the in vitro dimerization of avian sarcoma-leukosis virus (ASLV) RNA using different transcripts. We determined the requirements for heterodimer formation, the thermal dissociation of RNA dimers, and the influence of antisense DNA oligonucleotides on dimer formation. Our results strongly suggest that purine tetrads are not involved in dimer formation. Data show that an autocomplementary sequence located upstream from the splice donor site and within a major packaging signal plays a crucial role in ASLV RNA dimer formation in vitro. This sequence is able to form a stem-loop structure, and phylogenetic analysis reveals that it is conserved in 28 different avian sarcoma and leukosis viruses. These results suggest that dimerization of ASLV RNA is initiated by a loop-loop interaction between two RNA molecules and provide an additional argument for the ubiquity of the dimerization process via loop-loop interaction.

  9. Peptide Vaccine: Progress and Challenges

    Directory of Open Access Journals (Sweden)

    Weidang Li

    2014-07-01

    Full Text Available Conventional vaccine strategies have been highly efficacious for several decades in reducing mortality and morbidity due to infectious diseases. The bane of conventional vaccines, such as those that include whole organisms or large proteins, appear to be the inclusion of unnecessary antigenic load that, not only contributes little to the protective immune response, but complicates the situation by inducing allergenic and/or reactogenic responses. Peptide vaccines are an attractive alternative strategy that relies on usage of short peptide fragments to engineer the induction of highly targeted immune responses, consequently avoiding allergenic and/or reactogenic sequences. Conversely, peptide vaccines used in isolation are often weakly immunogenic and require particulate carriers for delivery and adjuvanting. In this article, we discuss the specific advantages and considerations in targeted induction of immune responses by peptide vaccines and progresses in the development of such vaccines against various diseases. Additionally, we also discuss the development of particulate carrier strategies and the inherent challenges with regard to safety when combining such technologies with peptide vaccines.

  10. Improvement of an enzyme-linked immunosorbent assay for equine herpesvirus type 4 by using a synthetic-peptide 24-mer repeat sequence of glycoprotein G as an antigen

    Science.gov (United States)

    BANNAI, Hiroshi; NEMOTO, Manabu; TSUJIMURA, Koji; YAMANAKA, Takashi; MAEDA, Ken; KONDO, Takashi

    2015-01-01

    To increase the sensitivity of an enzyme-linked immunosorbent assay (ELISA) for equine herpesvirus type 4 (EHV-4) that uses a 12-mer peptide of glycoprotein G (gG4-12-mer: MKNNPIYSEGSL) [4], we used a longer peptide consisting of a 24-mer repeat sequence (gG4-24-mer: MKNNPIYSEGSLMLNVQHDDSIHT) as an antigen. Sera of horses experimentally infected with EHV-4 reacted much more strongly to the gG4-24-mer peptide than to the gG4-12-mer peptide. We used peptide ELISAs to test paired sera from horses naturally infected with EHV-4 (n=40). gG4-24-mer ELISA detected 37 positive samples (92.5%), whereas gG4-12-mer ELISA detected only 28 (70.0%). gG4-24-mer ELISA was much more sensitive than gG4-12-mer ELISA. PMID:26424485

  11. Improvement of an enzyme-linked immunosorbent assay for equine herpesvirus type 4 by using a synthetic-peptide 24-mer repeat sequence of glycoprotein G as an antigen.

    Science.gov (United States)

    Bannai, Hiroshi; Nemoto, Manabu; Tsujimura, Koji; Yamanaka, Takashi; Maeda, Ken; Kondo, Takashi

    2016-02-01

    To increase the sensitivity of an enzyme-linked immunosorbent assay (ELISA) for equine herpesvirus type 4 (EHV-4) that uses a 12-mer peptide of glycoprotein G (gG4-12-mer: MKNNPIYSEGSL) [4], we used a longer peptide consisting of a 24-mer repeat sequence (gG4-24-mer: MKNNPIYSEGSLMLNVQHDDSIHT) as an antigen. Sera of horses experimentally infected with EHV-4 reacted much more strongly to the gG4-24-mer peptide than to the gG4-12-mer peptide. We used peptide ELISAs to test paired sera from horses naturally infected with EHV-4 (n=40). gG4-24-mer ELISA detected 37 positive samples (92.5%), whereas gG4-12-mer ELISA detected only 28 (70.0%). gG4-24-mer ELISA was much more sensitive than gG4-12-mer ELISA.

  12. Efficient Subcellular Targeting to the Cell Nucleus of Quantum Dots Densely Decorated with a Nuclear Localization Sequence Peptide.

    Science.gov (United States)

    Maity, Amit Ranjan; Stepensky, David

    2016-01-27

    Organelle-targeted drug delivery can enhance the efficiency of the intracellularly acting drugs and reduce their toxicity. We generated core-shell type CdSe-ZnS quantum dots (QDs) densely decorated with NLS peptidic targeting residues using a 3-stage decoration approach and investigated their endocytosis and nuclear targeting efficiencies. The diameter of the generated QDs increased following the individual decoration stages (16.3, 18.9, and 21.9 nm), the ζ-potential became less negative (-33.2, -17.5, and -11.9 mV), and characteristic changes appeared in the FTIR spectra following decoration with the linker and NLS peptides. Quantitative analysis of the last decoration stage revealed that 37.9% and 33.2% of the alkyne-modified NLS groups that were added to the reaction mix became covalently attached or adsorbed to the QDs surface, respectively. These numbers correspond to 63.6 and 55.7 peptides conjugated or adsorbed to a single QD (the surface density of 42 and 37 conjugated and adsorbed peptides per 1000 nm(2) of the QDs surface), which is higher than in the majority of previous studies that reported decoration efficiencies of formulations intended for nuclear-targeted drug delivery. QDs decorated with NLS peptides undergo more efficient endocytosis, as compared to other investigated QDs formulations, and accumulated to a higher extent in the cell nucleus or in close vicinity to it (11.9%, 14.6%, and 56.1% of the QDs endocytosed by an average cell for the QD-COOH, QD-azide, and QD-NLS formulations, respectively). We conclude that dense decoration of QDs with NLS residues increased their endocytosis and led to their nuclear targeting (preferential accumulation in the cells nuclei or in close vicinity to them). The experimental system and research tools that were used in this study allow quantitative investigation of the mechanisms that govern the QDs nuclear targeting and their dependence on the formulation properties. These findings will contribute to the

  13. Draft Genome Sequence of Lactobacillus delbrueckii Strain #22 Isolated from a Patient with Short Bowel Syndrome and Previous d-Lactic Acidosis and Encephalopathy.

    Science.gov (United States)

    Domann, Eugen; Fischer, Florence; Glowatzki, Fabian; Fritzenwanker, Moritz; Hain, Torsten; Zechel-Gran, Silke; Giffhorn-Katz, Susanne; Neubauer, Bernd A

    2016-07-28

    d-Lactic acidosis with associated encephalopathy caused by overgrowth of intestinal lactic acid bacteria is a rarely diagnosed neurological complication of patients with short bowel syndrome. Here, we report the draft genome sequence of Lactobacillus delbrueckii strain #22 isolated from a patient with short bowel syndrome and previous d-lactic acidosis/encephalopathy. Copyright © 2016 Domann et al.

  14. 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.

  15. Chimeric mitochondrial peptides from contiguous regular and swinger RNA.

    Science.gov (United States)

    Seligmann, Hervé

    2016-01-01

    Previous mass spectrometry analyses described human mitochondrial peptides entirely translated from swinger RNAs, RNAs where polymerization systematically exchanged nucleotides. Exchanges follow one among 23 bijective transformation rules, nine symmetric exchanges (X ↔ Y, e.g. A ↔ C) and fourteen asymmetric exchanges (X → Y → Z → X, e.g. A → C → G → A), multiplying by 24 DNA's protein coding potential. Abrupt switches from regular to swinger polymerization produce chimeric RNAs. Here, human mitochondrial proteomic analyses assuming abrupt switches between regular and swinger transcriptions, detect chimeric peptides, encoded by part regular, part swinger RNA. Contiguous regular- and swinger-encoded residues within single peptides are stronger evidence for translation of swinger RNA than previously detected, entirely swinger-encoded peptides: regular parts are positive controls matched with contiguous swinger parts, increasing confidence in results. Chimeric peptides are 200 × rarer than swinger peptides (3/100,000 versus 6/1000). Among 186 peptides with > 8 residues for each regular and swinger parts, regular parts of eleven chimeric peptides correspond to six among the thirteen recognized, mitochondrial protein-coding genes. Chimeric peptides matching partly regular proteins are rarer and less expressed than chimeric peptides matching non-coding sequences, suggesting targeted degradation of misfolded proteins. Present results strengthen hypotheses that the short mitogenome encodes far more proteins than hitherto assumed. Entirely swinger-encoded proteins could exist.

  16. Dynamical system modeling to simulate donor T cell response to whole exome sequencing-derived recipient peptides: Understanding randomness in alloreactivity incidence following stem cell transplantation.

    Directory of Open Access Journals (Sweden)

    Vishal Koparde

    Full Text Available Quantitative relationship between the magnitude of variation in minor histocompatibility antigens (mHA and graft versus host disease (GVHD pathophysiology in stem cell transplant (SCT donor-recipient pairs (DRP is not established. In order to elucidate this relationship, whole exome sequencing (WES was performed on 27 HLA matched related (MRD, & 50 unrelated donors (URD, to identify nonsynonymous single nucleotide polymorphisms (SNPs. An average 2,463 SNPs were identified in MRD, and 4,287 in URD DRP (p<0.01; resulting peptide antigens that may be presented on HLA class I molecules in each DRP were derived in silico (NetMHCpan ver2.0 and the tissue expression of proteins these were derived from determined (GTex. MRD DRP had an average 3,670 HLA-binding-alloreactive peptides, putative mHA (pmHA with an IC50 of <500 nM, and URD, had 5,386 (p<0.01. To simulate an alloreactive donor cytotoxic T cell response, the array of pmHA in each patient was considered as an operator matrix modifying a hypothetical cytotoxic T cell clonal vector matrix; each responding T cell clone's proliferation was determined by the logistic equation of growth, accounting for HLA binding affinity and tissue expression of each alloreactive peptide. The resulting simulated organ-specific alloreactive T cell clonal growth revealed marked variability, with the T cell count differences spanning orders of magnitude between different DRP. Despite an estimated, uniform set of constants used in the model for all DRP, and a heterogeneously treated group of patients, higher total and organ-specific T cell counts were associated with cumulative incidence of moderate to severe GVHD in recipients. In conclusion, exome wide sequence differences and the variable alloreactive peptide binding to HLA in each DRP yields a large range of possible alloreactive donor T cell responses. Our findings also help understand the apparent randomness observed in the development of alloimmune responses.

  17. Foreshock sequences and short-term earthquake predictability on East Pacific Rise transform faults.

    Science.gov (United States)

    McGuire, Jeffrey J; Boettcher, Margaret S; Jordan, Thomas H

    2005-03-24

    East Pacific Rise transform faults are characterized by high slip rates (more than ten centimetres a year), predominantly aseismic slip and maximum earthquake magnitudes of about 6.5. Using recordings from a hydroacoustic array deployed by the National Oceanic and Atmospheric Administration, we show here that East Pacific Rise transform faults also have a low number of aftershocks and high foreshock rates compared to continental strike-slip faults. The high ratio of foreshocks to aftershocks implies that such transform-fault seismicity cannot be explained by seismic triggering models in which there is no fundamental distinction between foreshocks, mainshocks and aftershocks. The foreshock sequences on East Pacific Rise transform faults can be used to predict (retrospectively) earthquakes of magnitude 5.4 or greater, in narrow spatial and temporal windows and with a high probability gain. The predictability of such transform earthquakes is consistent with a model in which slow slip transients trigger earthquakes, enrich their low-frequency radiation and accommodate much of the aseismic plate motion.

  18. VARIOUS SPEECH SEQUENCES OF ENGLISH DEPARTMENT STUDENTS IN DOING REQUEST VIA SHORT MESSAGE SERVICE

    Directory of Open Access Journals (Sweden)

    Ike Revita

    2015-09-01

    Full Text Available Interaction, if not wisely considered, may be very risky. The unwise utterances may lead to misunderstanding. When this happens, pragmatic failure is of great possibility occur. This writing is aimed at describing the some variations of speech seqeunces in doing request and the reasons of using each variation. The data are the request uttered by English Department students Andalas University to their friends, their lecturer and vice versa at the campus. Data are collected by observational method, interviewing and note-taking technique. To analyze the data, pragmatic and referential identity method is used. The result of analysis is naratively and descriptively presented. Having been related to the concept of speech act of request (Revita, 2008 and context (Yule, 1986, it is found that there are four variations of speech sequence when English Department students do request. They are (a (1 1 in 1; (b 2 in 1; (c 3 in 1; and (d multi acts in 1. The choice of these variations is basically based on several reasons, namely (i social; (iipsychological; (iii cultural ; and (iv religious aspect.

  19. Convolutional neural network regression for short-axis left ventricle segmentation in cardiac cine MR sequences.

    Science.gov (United States)

    Tan, Li Kuo; Liew, Yih Miin; Lim, Einly; McLaughlin, Robert A

    2017-07-01

    Automated left ventricular (LV) segmentation is crucial for efficient quantification of cardiac function and morphology to aid subsequent management of cardiac pathologies. In this paper, we parameterize the complete (all short axis slices and phases) LV segmentation task in terms of the radial distances between the LV centerpoint and the endo- and epicardial contours in polar space. We then utilize convolutional neural network regression to infer these parameters. Utilizing parameter regression, as opposed to conventional pixel classification, allows the network to inherently reflect domain-specific physical constraints. We have benchmarked our approach primarily against the publicly-available left ventricle segmentation challenge (LVSC) dataset, which consists of 100 training and 100 validation cardiac MRI cases representing a heterogeneous mix of cardiac pathologies and imaging parameters across multiple centers. Our approach attained a .77 Jaccard index, which is the highest published overall result in comparison to other automated algorithms. To test general applicability, we also evaluated against the Kaggle Second Annual Data Science Bowl, where the evaluation metric was the indirect clinical measures of LV volume rather than direct myocardial contours. Our approach attained a Continuous Ranked Probability Score (CRPS) of .0124, which would have ranked tenth in the original challenge. With this we demonstrate the effectiveness of convolutional neural network regression paired with domain-specific features in clinical segmentation. Copyright © 2017 Elsevier B.V. All rights reserved.

  20. Effects of treatment with glucagon-like peptide-2 on bone resorption in colectomized patients with distal ileostomy or jejunostomy and short-bowel syndrome

    DEFF Research Database (Denmark)

    Gottschalck, Ida B; Jeppesen, Palle B; Hartmann, Bolette

    2008-01-01

    OBJECTIVE: The gut hormone GLP-2 (glucagon-like peptide-2) seems to be involved in the circadian pattern of bone resorption, whereas parathyroid hormone (PTH) is an established key hormone in bone turnover. Endogenous GLP-2 secretion is lacking in colectomized patients with short-bowel syndrome...... (SBS) and they have reduced bone mineral density (BMD). The aim of the study was to investigate the anti-resorptive effect (assessed by s-CTX) of 14 days of GLP-2 treatment in these patients and to determine whether 56 days of treatment would improve BMD. PTH secretion in response to GLP-2 was also...... in the SBS patients, and after 56 days of GLP-2 treatment there was no improvement in BMD. A significant reduction in PTH secretion in response to GLP-2 was observed only in patients with ileostomy. CONCLUSIONS: The decreased bone resorption in response to GLP-2 injections cannot be elicited in SBS patients...

  1. Short-acting glucagon-like peptide-1 receptor agonists as add-on to insulin therapy in type 1 diabetes

    DEFF Research Database (Denmark)

    Albèr, Anders; Brønden, Andreas; Knop, Filip K

    2017-01-01

    emptying in patients with type 1 diabetes, which could translate into effective lowering of postprandial glucose excursions; however, these observations regarding short-acting GLP-1RAs are all derived from small open-label trials and should thus be interpreted with caution. In the present paper we review......A large proportion of patients with type 1 diabetes do not reach their glycaemic target of glycated hemoglobin (HbA1c) type 1 diabetes are overweight and obese. Treatment of type 1 diabetes is based on insulin therapy......, which is associated with well-described and unfortunate adverse effects such as hypoglycaemia and increased body weight. Glucagon-like peptide-1 (GLP-1) receptor agonists (RAs) are the focus of increasing interest as a possible adjunctive treatment to insulin in type 1 diabetes because...

  2. [Effects of introduction of short peptides before carotid artery occlusion on behaviour and caspase-3 activity in the brain of old rats].

    Science.gov (United States)

    Mendzheritskiĭ, A M; Karantysh, G V; Ivonina, K O

    2011-01-01

    The comparative research of effect of Pinealon and Cortexin on behavior and activity of caspase-3 in a brain of old rats in a model of carotid arteries occlusion was conducted. It is shown that introduction of short peptides promotes a survival rate of the animals that have modeled occlusion of carotid arteries. Under Pinealon before occlusion of carotid arteries, behavioral dream has been increased and a position-finding behavior, a motivational behavior and a motor performance have been reduced. The rats that were introduced Cortexin before carotid arteries occlusion demonstrated the raise of behavioral dream time. At introduction of Pinealon activity of caspase-3 moderately raises in false-operated animals and in a model of occlusion of carotid arteries.

  3. Clinical trial simulations in pediatric patients using realistic covariates: application to teduglutide, a glucagon-like peptide-2 analog in neonates and infants with short-bowel syndrome.

    Science.gov (United States)

    Mouksassi, M S; Marier, J F; Cyran, J; Vinks, A A

    2009-12-01

    Teduglutide, a synthetic glucagon-like peptide-2 (GLP-2) analog with activity relating to the regeneration, maintenance, and repair of the intestinal epithelium, is currently being evaluated for the treatment of short-bowel syndrome (SBS), Crohn's disease, and other gastrointestinal disorders. On the basis of promising results from teduglutide studies in adults with SBS and from studies in neonatal and juvenile animal models, a pediatric multiple-dose phase I clinical study was designed to determine the safety, efficacy, and pharmacokinetics of teduglutide in pediatric patients with SBS who have undergone resection for necrotizing enterocolitis, malrotation, or intestinal atresia. This report details the application of clinical trial simulations coupled with a novel approach using generalized additive modeling for location, scale, and shape (GAMLSS) that facilitates the simulation of demographic covariates specific to the targeted patient populations. The goal was to optimize phase I dosing strategies and the likelihood of achieving target exposure and therapeutic effect.

  4. Identification of a Bacteria Using Phylogenetic Relationships Revealed by MS/MS Sequencing of Tryptic Peptides Derived From Cellular Proteins

    National Research Council Canada - National Science Library

    Dworzanski, J. P; Wick, H; Snyder, A. P; Deshpande, S. V; Chen, R; Li, L

    2004-01-01

    .... In addition four hundred bacterial genome-sequencing projects are in progress. These achievements provide new possibilities for reliable identification of bacteria on a molecular level by retrieving their genomic information...

  5. Transepithelial transport of milk-derived angiotensin I-converting enzyme inhibitory peptide with the RLSFNP sequence.

    Science.gov (United States)

    Guo, Yuxing; Gan, Junai; Zhu, Qian; Zeng, Xiaoqun; Sun, Yangying; Wu, Zhen; Pan, Daodong

    2018-02-01

    To exert an antihypertensive effect after oral administration, angiotensin I-converting enzyme (ACE)-inhibitory peptides must remain active after intestinal transport. The purpose of this article is to elucidate the transport permeability and route of ACE-inhibitory peptide Arg-Leu-Ser-Phe-Asn-Pro (RLSFNP) across the intestinal epithelium using Caco-2 cell monolayers. Intact RLSFNP and RLSFNP breakdown fragments F, FNP, SFNP and RLSF were found in RLSFNP transport solution across Caco-2 cell monolayers using ultra-performance liquid chromatography-tandem mass spectrometry. RLSFNP fragments FNP, SFNP and RLSF also contributed to ACE inhibitory effects. Protease inhibitors (bacitracin and leupeptin) and absorption enhancers (sodium glycocholate hydrate, sodium deoxycholate and Na 2 EDTA) improved the transport flux of RLSFNP. A transport inhibitor experiment showed that intact RLSFNP may be transported via the paracellular route. Intact RLSFNP can be transported across the Caco-2 cell monolayers via the paracellular route. Extensive hydrolysis was the chief reason for the low permeability of RLSFNP. © 2017 Society of Chemical Industry. © 2017 Society of Chemical Industry.

  6. Long-term earthquake forecasts based on the epidemic-type aftershock sequence (ETAS model for short-term clustering

    Directory of Open Access Journals (Sweden)

    Jiancang Zhuang

    2012-07-01

    Full Text Available Based on the ETAS (epidemic-type aftershock sequence model, which is used for describing the features of short-term clustering of earthquake occurrence, this paper presents some theories and techniques related to evaluating the probability distribution of the maximum magnitude in a given space-time window, where the Gutenberg-Richter law for earthquake magnitude distribution cannot be directly applied. It is seen that the distribution of the maximum magnitude in a given space-time volume is determined in the longterm by the background seismicity rate and the magnitude distribution of the largest events in each earthquake cluster. The techniques introduced were applied to the seismicity in the Japan region in the period from 1926 to 2009. It was found that the regions most likely to have big earthquakes are along the Tohoku (northeastern Japan Arc and the Kuril Arc, both with much higher probabilities than the offshore Nankai and Tokai regions.

  7. Short communication: Promotion of glucagon-like peptide-2 secretion in dairy calves with a bioactive extract from Olea europaea.

    Science.gov (United States)

    Morrison, S Y; Pastor, J J; Quintela, J C; Holst, J J; Hartmann, B; Drackley, J K; Ipharraguerre, I R

    2017-03-01

    Diarrhea episodes in dairy calves involve profound alterations in the mechanism controlling gut barrier function that ultimately compromise intestinal permeability to macromolecules, including pathogenic bacteria. Intestinal dysfunction models suggest that a key element of intestinal adaptation during the neonatal phase is the nutrient-induced secretion of glucagon-like peptide (GLP)-2 and associated effects on mucosal cell proliferation, barrier function, and inflammatory response. Bioactive molecules found in Olea europaea have been shown to induce the release of regulatory peptides from model enteroendocrine cells. The ability to enhance GLP-2 secretion via the feeding of putative GLP-2 secretagogues is untested in newborn calves. The objectives of this study were to determine whether feeding a bioactive extract from Olea europaea (OBE) mixed in the milk replacer (1) can stimulate GLP-2 secretion beyond the response elicited by enteral nutrients and, thereby, (2) improve intestinal permeability and animal growth as well as (3) reduce the incidence of diarrhea in preweaning dairy calves. Holstein heifer calves (n = 60) were purchased, transported to the research facility, and blocked by body weight and total serum protein and assigned to 1 of 3 treatments. Treatments were control (CON), standard milk replacer (MR) and ad libitum starter; CON plus OBE added into MR at 30 mg/kg of body weight (OBE30); and CON plus OBE added into MR at 60 mg/kg of body weight (OBE60). The concentration of GLP-2 was measured at the end of wk 2. Intestinal permeability was measured at the onset of the study and the end of wk 2 and 6, with lactulose and d-mannitol as markers. Treatments did not affect calf growth and starter intake. Compared with CON, administration of OBE60 increased the nutrient-induced response in GLP-2 by about 1 fold and reduced MR intake during the second week of study. Throughout the study, however, all calves had compromised intestinal permeability and a high

  8. [3H]Azidodantrolene photoaffinity labeling, synthetic domain peptides and monoclonal antibody reactivity identify the dantrolene binding sequence on RyR1

    Energy Technology Data Exchange (ETDEWEB)

    Paul-Pletzer, Kalanethee; Yamamoto, Takeshi; Bhat, Manju B.; Ma, Jianjie; Ikemoto, Noriaki; Jimenez, Leslie S.; Morimoto, Hiromi; Williams, Philip G.; Parness, Jerome

    2002-06-14

    Dantrolene is a drug that suppresses intracellular Ca2+ release from sarcoplasmic reticulum in normal skeletal muscle and is used as a therapeutic agent in individuals susceptible to malignant hyperthermia. Though its precise mechanism of action has not been elucidated, we have identified the N-terminal region (amino acids 1-1400) of the skeletal muscle isoform of the ryanodine receptor (RyR1), the primary Ca2+ release channel in sarcoplasmic reticulum, as a molecular target for dantrolene using the photoaffinity analog [3H]azidodantrolene(1). Here, we demonstrate that heterologously expressed RyR1 retains its capacity to be specifically labeled with [3H]azidodantrolene,indicating that muscle specific factors are not required for this ligand-receptor interaction. Synthetic domain peptides of RyR1, previously shown to affect RyR1 function in vitro and in vivo, were exploited as potential drug binding site mimics and used in photoaffinity labeling experiments. Only DP1 and DP1-2, peptide s containing the amino acid sequence corresponding to RyR1 residues 590-609, were specifically labeled by [3H]azidodantrolene. A monoclonal anti-RyR1 antibody which recognizes RyR1 and its 1400 amino acid N-terminal fragment, recognizes DP1 and DP1-2 in both Western blots and immunoprecipitation assays, and specifically inhibits [3H]azidodantrolene photolabeling of RyR1 and its N-terminal fragment in sarcoplasmic reticulum. Our results indicate that synthetic domain peptides can mimic a native, ligand binding conformation in vitro, and that the dantrolene binding site and the epitope for the monoclonal antibody on RyR1 are equivalent and composed of amino-acids 590-609.

  9. Alignment of Short Reads: A Crucial Step for Application of Next-Generation Sequencing Data in Precision Medicine

    Directory of Open Access Journals (Sweden)

    Hao Ye

    2015-11-01

    Full Text Available Precision medicine or personalized medicine has been proposed as a modernized and promising medical strategy. Genetic variants of patients are the key information for implementation of precision medicine. Next-generation sequencing (NGS is an emerging technology for deciphering genetic variants. Alignment of raw reads to a reference genome is one of the key steps in NGS data analysis. Many algorithms have been developed for alignment of short read sequences since 2008. Users have to make a decision on which alignment algorithm to use in their studies. Selection of the right alignment algorithm determines not only the alignment algorithm but also the set of suitable parameters to be used by the algorithm. Understanding these algorithms helps in selecting the appropriate alignment algorithm for different applications in precision medicine. Here, we review current available algorithms and their major strategies such as seed-and-extend and q-gram filter. We also discuss the challenges in current alignment algorithms, including alignment in multiple repeated regions, long reads alignment and alignment facilitated with known genetic variants.

  10. 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.

  11. Short term reproducibility of a high contrast 3-D isotropic optic nerve imaging sequence in healthy controls

    Science.gov (United States)

    Harrigan, Robert L.; Smith, Alex K.; Mawn, Louise A.; Smith, Seth A.; Landman, Bennett A.

    2016-03-01

    The optic nerve (ON) plays a crucial role in human vision transporting all visual information from the retina to the brain for higher order processing. There are many diseases that affect the ON structure such as optic neuritis, anterior ischemic optic neuropathy and multiple sclerosis. Because the ON is the sole pathway for visual information from the retina to areas of higher level processing, measures of ON damage have been shown to correlate well with visual deficits. Increased intracranial pressure has been shown to correlate with the size of the cerebrospinal fluid (CSF) surrounding the ON. These measures are generally taken at an arbitrary point along the nerve and do not account for changes along the length of the ON. We propose a high contrast and high-resolution 3-D acquired isotropic imaging sequence optimized for ON imaging. We have acquired scan-rescan data using the optimized sequence and a current standard of care protocol for 10 subjects. We show that this sequence has superior contrast-to-noise ratio to the current standard of care while achieving a factor of 11 higher resolution. We apply a previously published automatic pipeline to segment the ON and CSF sheath and measure the size of each individually. We show that these measures of ON size have lower short- term reproducibility than the population variance and the variability along the length of the nerve. We find that the proposed imaging protocol is (1) useful in detecting population differences and local changes and (2) a promising tool for investigating biomarkers related to structural changes of the ON.

  12. Induction of porcine host defense peptide gene expression by short-chain fatty acids and their analogs.

    Directory of Open Access Journals (Sweden)

    Xiangfang Zeng

    Full Text Available Dietary modulation of the synthesis of endogenous host defense peptides (HDPs represents a novel antimicrobial approach for disease control and prevention, particularly against antibiotic-resistant infections. However, HDP regulation by dietary compounds such as butyrate is species-dependent. To examine whether butyrate could induce HDP expression in pigs, we evaluated the expressions of a panel of porcine HDPs in IPEC-J2 intestinal epithelial cells, 3D4/31 macrophages, and primary monocytes in response to sodium butyrate treatment by real-time PCR. We revealed that butyrate is a potent inducer of multiple, but not all, HDP genes. Porcine β-defensin 2 (pBD2, pBD3, epididymis protein 2 splicing variant C (pEP2C, and protegrins were induced markedly in response to butyrate, whereas pBD1 expression remained largely unaltered in any cell type. Additionally, a comparison of the HDP-inducing efficacy among saturated free fatty acids of different aliphatic chain lengths revealed that fatty acids containing 3-8 carbons showed an obvious induction of HDP expression in IPEC-J2 cells, with butyrate being the most potent and long-chain fatty acids having only a marginal effect. We further investigated a panel of butyrate analogs for their efficacy in HDP induction, and found glyceryl tributyrate, benzyl butyrate, and 4-phenylbutyrate to be comparable with butyrate. Identification of butyrate and several analogs with a strong capacity to induce HDP gene expression in pigs provides attractive candidates for further evaluation of their potential as novel alternatives to antibiotics in augmenting innate immunity and disease resistance of pigs.

  13. A dose-equivalent comparison of the effects of continuous subcutaneous glucagon-like peptide 2 (GLP-2) infusions versus meal related GLP-2 injections in the treatment of short bowel syndrome (SBS) patients

    DEFF Research Database (Denmark)

    Naimi, R M; Madsen, K B; Askov-Hansen, C

    2013-01-01

    Glucagon-like peptide 2 (GLP-2), secreted endogenously from L-cells in the distal bowel in relation to meals, modulates intestinal absorption by adjusting gastric emptying and secretion and intestinal growth. Short bowel syndrome (SBS) patients with distal intestinal resections have attenuated...

  14. Two short basic sequences surrounding the zinc finger of nucleocapsid protein NCp10 of Moloney murine leukemia virus are critical for RNA annealing activity.

    Science.gov (United States)

    De Rocquigny, H; Ficheux, D; Gabus, C; Allain, B; Fournie-Zaluski, M C; Darlix, J L; Roques, B P

    1993-02-25

    The 56 amino acid nucleocapsid protein (NCp10) of Moloney Murine Leukemia Virus, contains a CysX2CysX4HisX4Cys zinc finger flanked by basic residues. In vitro NCp10 promotes genomic RNA dimerization, a process most probably linked to genomic RNA packaging, and replication primer tRNA(Pro) annealing to the initiation site of reverse transcription. To characterize the amino-acid sequences involved in the various functions of NCp10, we have synthesized by solid phase method the native protein and a series of derived peptides shortened at the N- or C-terminus with or without the zinc finger domain. In the latter case, the two parts of the protein were linked by a Glycine - Glycine spacer. The in vitro studies of these peptides show that nucleic acid annealing activities of NCp10 do not require a zinc finger but are critically dependent on the presence of specific sequences located on each side of the CCHC domain and containing proline and basic residues. Thus, deletion of 11R or 49PRPQT, of the fully active 29 residue peptide 11RQGGERRRSQLDRDGGKKPRGPRGPRPQT53 leads to a complete loss of NCp10 activity. Therefore it is proposed that in NCp10, the zinc finger directs the spatial recognition of the target RNAs by the basic domains surrounding the zinc finger.

  15. Relationship between apoptosis and the BH2 domain sequence of the VP5 peptide of infectious pancreatic necrosis virus

    Directory of Open Access Journals (Sweden)

    Cesar Ortega S.

    2014-03-01

    Full Text Available Objective. To determine whether the level of apoptosis induced by infectious pancreatic necrosis virus (IPNV is related to the amino acid sequence of the BH2 domain of the VP5 protein and the level of infectivity. Materials and methods. Three IPNV strains were used, the VP2 protein gene was amplified for genotyping and the VP5 sequence was also obtained. The infectivity of the strains was calculated using the viral titer obtained at 12, 24, 36 and 45 hpi in CHSE-214 cells. The percentage of apoptosis in infected cells was visualized by TUNEL assay and immunohistochemistry (caspase 3 detection. Results. The V70/06 and V33/98 strains corresponded to genotype Sp, while V112/06 to VR-299; the amino acid analysis of the V70/06 strain allows its classification as middle virulent strain and V33/98 and V112/06 strains as low virulent ones; infection with the V112/06 strain produced a lower viral titer (p0.05. Conclusions. The results showed that the differences in the BH2 sequence of the VP5 protein, infectivity and the VP2 sequence are not associated with the modulation of apoptosis.

  16. De novo assembly of a 40 Mb eukaryotic genome from short sequence reads: Sordaria macrospora, a model organism for fungal morphogenesis.

    Science.gov (United States)

    Nowrousian, Minou; Stajich, Jason E; Chu, Meiling; Engh, Ines; Espagne, Eric; Halliday, Karen; Kamerewerd, Jens; Kempken, Frank; Knab, Birgit; Kuo, Hsiao-Che; Osiewacz, Heinz D; Pöggeler, Stefanie; Read, Nick D; Seiler, Stephan; Smith, Kristina M; Zickler, Denise; Kück, Ulrich; Freitag, Michael

    2010-04-08

    Filamentous fungi are of great importance in ecology, agriculture, medicine, and biotechnology. Thus, it is not surprising that genomes for more than 100 filamentous fungi have been sequenced, most of them by Sanger sequencing. While next-generation sequencing techniques have revolutionized genome resequencing, e.g. for strain comparisons, genetic mapping, or transcriptome and ChIP analyses, de novo assembly of eukaryotic genomes still presents significant hurdles, because of their large size and stretches of repetitive sequences. Filamentous fungi contain few repetitive regions in their 30-90 Mb genomes and thus are suitable candidates to test de novo genome assembly from short sequence reads. Here, we present a high-quality draft sequence of the Sordaria macrospora genome that was obtained by a combination of Illumina/Solexa and Roche/454 sequencing. Paired-end Solexa sequencing of genomic DNA to 85-fold coverage and an additional 10-fold coverage by single-end 454 sequencing resulted in approximately 4 Gb of DNA sequence. Reads were assembled to a 40 Mb draft version (N50 of 117 kb) with the Velvet assembler. Comparative analysis with Neurospora genomes increased the N50 to 498 kb. The S. macrospora genome contains even fewer repeat regions than its closest sequenced relative, Neurospora crassa. Comparison with genomes of other fungi showed that S. macrospora, a model organism for morphogenesis and meiosis, harbors duplications of several genes involved in self/nonself-recognition. Furthermore, S. macrospora contains more polyketide biosynthesis genes than N. crassa. Phylogenetic analyses suggest that some of these genes may have been acquired by horizontal gene transfer from a distantly related ascomycete group. Our study shows that, for typical filamentous fungi, de novo assembly of genomes from short sequence reads alone is feasible, that a mixture of Solexa and 454 sequencing substantially improves the assembly, and that the resulting data can be used for

  17. De novo assembly of a 40 Mb eukaryotic genome from short sequence reads: Sordaria macrospora, a model organism for fungal morphogenesis.

    Directory of Open Access Journals (Sweden)

    Minou Nowrousian

    2010-04-01

    Full Text Available Filamentous fungi are of great importance in ecology, agriculture, medicine, and biotechnology. Thus, it is not surprising that genomes for more than 100 filamentous fungi have been sequenced, most of them by Sanger sequencing. While next-generation sequencing techniques have revolutionized genome resequencing, e.g. for strain comparisons, genetic mapping, or transcriptome and ChIP analyses, de novo assembly of eukaryotic genomes still presents significant hurdles, because of their large size and stretches of repetitive sequences. Filamentous fungi contain few repetitive regions in their 30-90 Mb genomes and thus are suitable candidates to test de novo genome assembly from short sequence reads. Here, we present a high-quality draft sequence of the Sordaria macrospora genome that was obtained by a combination of Illumina/Solexa and Roche/454 sequencing. Paired-end Solexa sequencing of genomic DNA to 85-fold coverage and an additional 10-fold coverage by single-end 454 sequencing resulted in approximately 4 Gb of DNA sequence. Reads were assembled to a 40 Mb draft version (N50 of 117 kb with the Velvet assembler. Comparative analysis with Neurospora genomes increased the N50 to 498 kb. The S. macrospora genome contains even fewer repeat regions than its closest sequenced relative, Neurospora crassa. Comparison with genomes of other fungi showed that S. macrospora, a model organism for morphogenesis and meiosis, harbors duplications of several genes involved in self/nonself-recognition. Furthermore, S. macrospora contains more polyketide biosynthesis genes than N. crassa. Phylogenetic analyses suggest that some of these genes may have been acquired by horizontal gene transfer from a distantly related ascomycete group. Our study shows that, for typical filamentous fungi, de novo assembly of genomes from short sequence reads alone is feasible, that a mixture of Solexa and 454 sequencing substantially improves the assembly, and that the resulting data

  18. Peptide Nucleic Acids

    DEFF Research Database (Denmark)

    2004-01-01

    A novel class of compounds known as peptide nucleic acids, bind complementary DNA and RNA strands, and generally do so more strongly than the corresponding DNA or RNA strands while exhibiting increased sequence specificity and solubility. The peptide nucleic acids comprise ligands selected from...

  19. Fine de novo sequencing of a fungal genome using only SOLiD short read data: verification on Aspergillus oryzae RIB40.

    Directory of Open Access Journals (Sweden)

    Myco Umemura

    Full Text Available The development of next-generation sequencing (NGS technologies has dramatically increased the throughput, speed, and efficiency of genome sequencing. The short read data generated from NGS platforms, such as SOLiD and Illumina, are quite useful for mapping analysis. However, the SOLiD read data with lengths of <60 bp have been considered to be too short for de novo genome sequencing. Here, to investigate whether de novo sequencing of fungal genomes is possible using only SOLiD short read sequence data, we performed de novo assembly of the Aspergillus oryzae RIB40 genome using only SOLiD read data of 50 bp generated from mate-paired libraries with 2.8- or 1.9-kb insert sizes. The assembled scaffolds showed an N50 value of 1.6 Mb, a 22-fold increase than those obtained using only SOLiD short read in other published reports. In addition, almost 99% of the reference genome was accurately aligned by the assembled scaffold fragments in long lengths. The sequences of secondary metabolite biosynthetic genes and clusters, whose products are of considerable interest in fungal studies due to their potential medicinal, agricultural, and cosmetic properties, were also highly reconstructed in the assembled scaffolds. Based on these findings, we concluded that de novo genome sequencing using only SOLiD short reads is feasible and practical for molecular biological study of fungi. We also investigated the effect of filtering low quality data, library insert size, and k-mer size on the assembly performance, and recommend for the assembly use of mild filtered read data where the N50 was not so degraded and the library has an insert size of ∼2.0 kb, and k-mer size 33.

  20. Effects of sequence changes in the HIV-1 gp41 fusion peptide on CCR5 inhibitor resistance

    International Nuclear Information System (INIS)

    Anastassopoulou, Cleo G.; Ketas, Thomas J.; Sanders, Rogier W.; Johan Klasse, Per; Moore, John P.

    2012-01-01

    A rare pathway of HIV-1 resistance to small molecule CCR5 inhibitors such as Vicriviroc (VCV) involves changes solely in the gp41 fusion peptide (FP). Here, we show that the G516V change is critical to VCV resistance in PBMC and TZM-bl cells, although it must be accompanied by either M518V or F519I to have a substantial impact. Modeling VCV inhibition data from the two cell types indicated that G516V allows both double mutants to use VCV-CCR5 complexes for entry. The model further identified F519I as an independent determinant of preference for the unoccupied, high-VCV affinity form of CCR5. From inhibitor-free reversion cultures, we also identified a substitution in the inner domain of gp120, T244A, which appears to counter the resistance phenotype created by the FP substitutions. Examining the interplay of these changes will enhance our understanding of Env complex interactions that influence both HIV-1 entry and resistance to CCR5 inhibitors.

  1. Serodiagnosis of cutaneous leishmaniasis: assessment of an enzyme-linked immunosorbent assay using a peptide sequence from gene B protein

    DEFF Research Database (Denmark)

    Jensen, A T; Gaafar, A; Ismail, A

    1996-01-01

    An enzyme-linked immunosorbent assay (ELISA) using a 28 amino acid sequence of the repetitive element of gene B protein (GBP) from Leishmania major was developed for serodiagnosis of cutaneous leishmaniasis (CL). The assay was compared to ELISAs using crude amastigote and promastigote antigens from...... samples from healthy Sudanese individuals living in an area endemic for malaria but free of leish-maniasis were negative in all the assays. Significantly higher levels of antibodies were found in the patients who had suffered from the disease for more than eight weeks than in patients with a shorter...

  2. 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.

  3. Evaluation of short repetition time, partial flip angle, gradient recalled echo pulse sequences in cervical spine imaging

    International Nuclear Information System (INIS)

    Enzmann, D.; Rubin, J.B.

    1987-01-01

    A short repetition time (TR), partial flip angle, gradient recalled echo pulse sequence (GRASS) was prospectively studied to optimize it for the diagnosis of cervical disk and cord disease in 98 patients. Changes in signal-to-noise ratio (SNR) and contrast were measured as the following parameters were varied: flip angle (3 0 to 18 0 ), TR (22-60 msec), and echo time (TE) (12.5-25 msec). Flip angle was the single most important parameter. For disk disease, cerebrospinal fluid (CSF) SNR peaked at an 8 0 flip angle in the axial view but at a 4 0 flip angle in the sagittal view. In the sagittal view, disk-CSF contrast decreased progressively from a flip angle of 3 0 , while in the axial view it peaked at 10 0 . For cord lesions the findings were similar except that lesion-cord contrast could be increased by lengthening both TR and TE. No one combination of parameters proved greatly superior for either disk disease or cord disease. The selection of parameters required balancing of several factors that often had opposing effects

  4. A novel family of sequence-specific endoribonucleases associated with the clustered regularly interspaced short palindromic repeats.

    Science.gov (United States)

    Beloglazova, Natalia; Brown, Greg; Zimmerman, Matthew D; Proudfoot, Michael; Makarova, Kira S; Kudritska, Marina; Kochinyan, Samvel; Wang, Shuren; Chruszcz, Maksymilian; Minor, Wladek; Koonin, Eugene V; Edwards, Aled M; Savchenko, Alexei; Yakunin, Alexander F

    2008-07-18

    Clustered regularly interspaced short palindromic repeats (CRISPRs) together with the associated CAS proteins protect microbial cells from invasion by foreign genetic elements using presently unknown molecular mechanisms. All CRISPR systems contain proteins of the CAS2 family, suggesting that these uncharacterized proteins play a central role in this process. Here we show that the CAS2 proteins represent a novel family of endoribonucleases. Six purified CAS2 proteins from diverse organisms cleaved single-stranded RNAs preferentially within U-rich regions. A representative CAS2 enzyme, SSO1404 from Sulfolobus solfataricus, cleaved the phosphodiester linkage on the 3'-side and generated 5'-phosphate- and 3'-hydroxyl-terminated oligonucleotides. The crystal structure of SSO1404 was solved at 1.6A resolution revealing the first ribonuclease with a ferredoxin-like fold. Mutagenesis of SSO1404 identified six residues (Tyr-9, Asp-10, Arg-17, Arg-19, Arg-31, and Phe-37) that are important for enzymatic activity and suggested that Asp-10 might be the principal catalytic residue. Thus, CAS2 proteins are sequence-specific endoribonucleases, and we propose that their role in the CRISPR-mediated anti-phage defense might involve degradation of phage or cellular mRNAs.

  5. Short Bowel Patients Treated for Two Years with Glucagon-Like Peptide 2: Effects on Intestinal Morphology and Absorption, Renal Function, Bone and Body Composition, and Muscle Function

    Directory of Open Access Journals (Sweden)

    P. B. Jeppesen

    2009-01-01

    Full Text Available Background and aims. In a short-term study, Glucagon-like peptide 2 (GLP-2 has been shown to improve intestinal absorption in short bowel syndrome (SBS patients. This study describes longitudinal changes in relation to GLP-2 treatment for two years. Methods. GLP-2, 400 micrograms, s.c.,TID, were offered, to eleven SBS patients keeping parenteral support constant. 72-hour nutritional balance studies were performed at baseline, weeks 13, 26, 52 during two years intermitted by an 8-week washout period. In addition, mucosal morphometrics, renal function (by creatinine clearance, body composition and bone mineral density (by DEXA, biochemical markers of bone turnover (by s-CTX and osteocalcin, PTH and vitamin D, and muscle function (NMR, lungfunction, exercise test were measured. Results. GLP-2 compliance was >93%. Three of eleven patients did not complete the study. In the remaining 8 patients, GLP-2 significantly reduced the fecal wet weight from approximately 3.0 to approximately 2.0 kg/day. This was accompanied by a decline in the oral wet weight intake, maintaining intestinal wet weight absorption and urinary weight constant. Renal function improved. No significant changes were demonstrated in energy intake or absorption, and GLP-2 did not significantly affect mucosal morphology, body composition, bone mineral density or muscle function. Conclusions. GLP-2 treatment reduces fecal weight by approximately 1000 g/d and enables SBS patients to maintain their intestinal fluid and electrolyte absorption at lower oral intakes. This was accompanied by a 28% improvement in creatinine clearance.

  6. MetaVelvet: An Extension of Velvet Assembler to de novo Metagenome Assembly from Short Sequence Reads (Metagenomics Informatics Challenges Workshop: 10K Genomes at a Time)

    Energy Technology Data Exchange (ETDEWEB)

    Sakakibara, Yasumbumi

    2011-10-13

    Keio University's Yasumbumi Sakakibara on "MetaVelvet: An Extension of Velvet Assembler to de novo Metagenome Assembly from Short Sequence Reads" at the Metagenomics Informatics Challenges Workshop held at the DOE JGI on October 12-13, 2011.

  7. ARA-PEPs: a repository of putative sORF-encoded peptides in Arabidopsis thaliana.

    Science.gov (United States)

    Hazarika, Rashmi R; De Coninck, Barbara; Yamamoto, Lidia R; Martin, Laura R; Cammue, Bruno P A; van Noort, Vera

    2017-01-17

    Many eukaryotic RNAs have been considered non-coding as they only contain short open reading frames (sORFs). However, there is increasing evidence for the translation of these sORFs into bioactive peptides with potent signaling, antimicrobial, developmental, antioxidant roles etc. Yet only a few peptides encoded by sORFs are annotated in the model organism Arabidopsis thaliana. To aid the functional annotation of these peptides, we have developed ARA-PEPs (available at http://www.biw.kuleuven.be/CSB/ARA-PEPs ), a repository of putative peptides encoded by sORFs in the A. thaliana genome starting from in-house Tiling arrays, RNA-seq data and other publicly available datasets. ARA-PEPs currently lists 13,748 sORF-encoded peptides with transcriptional evidence. In addition to existing data, we have identified 100 novel transcriptionally active regions (TARs) that might encode 341 novel stress-induced peptides (SIPs). To aid in identification of bioactivity, we add functional annotation and sequence conservation to predicted peptides. To our knowledge, this is the largest repository of plant peptides encoded by sORFs with transcript evidence, publicly available and this resource will help scientists to effortlessly navigate the list of experimentally studied peptides, the experimental and computational evidence supporting the activity of these peptides and gain new perspectives for peptide discovery.

  8. Ranked solutions to a class of combinatorial optimizations—with applications in mass spectrometry based peptide sequencing and a variant of directed paths in random media

    Science.gov (United States)

    Doerr, Timothy P.; Alves, Gelio; Yu, Yi-Kuo

    2005-08-01

    Typical combinatorial optimizations are NP-hard; however, for a particular class of cost functions the corresponding combinatorial optimizations can be solved in polynomial time using the transfer matrix technique or, equivalently, the dynamic programming approach. This suggests a way to efficiently find approximate solutions-find a transformation that makes the cost function as similar as possible to that of the solvable class. After keeping many high-ranking solutions using the approximate cost function, one may then re-assess these solutions with the full cost function to find the best approximate solution. Under this approach, it is important to be able to assess the quality of the solutions obtained, e.g., by finding the true ranking of the kth best approximate solution when all possible solutions are considered exhaustively. To tackle this statistical issue, we provide a systematic method starting with a scaling function generated from the finite number of high-ranking solutions followed by a convergent iterative mapping. This method, useful in a variant of the directed paths in random media problem proposed here, can also provide a statistical significance assessment for one of the most important proteomic tasks-peptide sequencing using tandem mass spectrometry data. For directed paths in random media, the scaling function depends on the particular realization of randomness; in the mass spectrometry case, the scaling function is spectrum-specific.

  9. The cytoplasmic domain close to the transmembrane region of the glucagon-like peptide-1 receptor contains sequence elements that regulate agonist-dependent internalisation.

    Science.gov (United States)

    Vázquez, Patricia; Roncero, Isabel; Blázquez, Enrique; Alvarez, Elvira

    2005-07-01

    In order to gain better insight into the molecular events involved in the signal transduction generated through glucagon-like peptide-1 (GLP-1) receptors, we tested the effect of deletions and point mutations within the cytoplasmic tail of this receptor with a view to establishing relationships between signal transduction desensitisation and receptor internalisation. Wild-type and truncated (deletion of the last 27 amino acids (GLPR 435R) and deletion of 44 amino acids (GLPR 418R)) GLP-1 receptors bound the agonist with similar affinity. Deletion of the last 27 amino acids decreased the internalisation rate by 78%, while deletion of 44 amino acids containing all the phosphorylation sites hitherto described in this receptor decreased the internalisation rate by only 47%. Binding of the ligand to both receptors stimulated adenylyl cyclase. In contrast, deletion of the region containing amino acids 419 to 435 (GLPR 419delta435) increased the internalisation rate by 268%, and the replacement of EVQ(408-410) by alanine (GLPR A(408-410)) increased this process to 296%. In both receptors, the efficacy in stimulating adenylate cyclase was decreased. All the receptors studied were internalised by coated pits, except for the receptor with a deletion of the last 44 amino acids, which also had a faster resensitisation rate. Our findings indicate that the neighbouring trans-membrane domain of the carboxyl-terminal tail of the GLP-1 receptor contains sequence elements that regulate agonist-dependent internalisation and transmembrane signalling.

  10. Characterization of synthetic peptides by mass spectrometry

    DEFF Research Database (Denmark)

    Prabhala, Bala Krishna; Mirza, Osman Asghar; Højrup, Peter

    2015-01-01

    Mass spectrometry (MS) is well suited for analysis of the identity and purity of synthetic peptides. The sequence of a synthetic peptide is most often known, so the analysis is mainly used to confirm the identity and purity of the peptide. Here, simple procedures are described for MALDI......-TOF-MS and LC-MS of synthetic peptides....

  11. Effects of phosphorylatable short peptide-conjugated chitosan-mediated IL-1Ra and igf-1 gene transfer on articular cartilage defects in rabbits.

    Directory of Open Access Journals (Sweden)

    Ronglan Zhao

    Full Text Available Previously, we reported an improvement in the transfection efficiency of the plasmid DNA-chitosan (pDNA/CS complex by the utilization of phosphorylatable short peptide-conjugated chitosan (pSP-CS. In this study, we investigated the effects of pSP-CS-mediated gene transfection of interleukin-1 receptor antagonist protein (IL-1Ra combined with insulin-like growth factor-1 (IGF-1 in rabbit chondrocytes and in a rabbit model of cartilage defects. pBudCE4.1-IL-1Ra+igf-1, pBudCE4.1-IL-1Ra and pBudCE4.1-igf-1 were constructed and combined with pSP-CS to form pDNA/pSP-CS complexes. These complexes were transfected into rabbit primary chondrocytes or injected into the joint cavity. Seven weeks after treatment, all rabbits were sacrificed and analyzed. High levels of IL-1Ra and igf-1 expression were detected both in the cell culture supernatant and in the synovial fluid. In vitro, the transgenic complexes caused significant proliferation of chondrocytes, promotion of glycosaminoglycan (GAG and collagen II synthesis, and inhibition of chondrocyte apoptosis and nitric oxide (NO synthesis. In vivo, the exogenous genes resulted in increased collagen II synthesis and reduced NO and GAG concentrations in the synovial fluid; histological studies revealed that pDNA/pSP-CS treatment resulted in varying degrees of hyaline-like cartilage repair and Mankin score decrease. The co-expression of both genes produced greater effects than each single gene alone both in vitro and in vivo. The results suggest that pSP-CS is a good candidate for use in gene therapy for the treatment of cartilage defects and that igf-1 and IL-1Ra co-expression produces promising biologic effects on cartilage defects.

  12. Pro B-type natriuretic peptide plasma value: a new criterion for the prediction of short- and long-term outcomes after transcatheter aortic valve implantation.

    Science.gov (United States)

    López-Otero, Diego; Trillo-Nouche, Ramiro; Gude, Francisco; Cid-Álvarez, Belen; Ocaranza-Sanchez, Raimundo; Alvarez, Melisa Santas; Lear, Pamela V; Gonzalez-Juanatey, José R

    2013-09-30

    To determine the prognostic value of pro B-type natriuretic peptide (pro-BNP) to predict mortality after transcatheter aortic valve implantation (TAVI). Logistic EuroSCORE (LES) overestimates observed mortality after TAVI. A new risk score specific to TAVI is needed to accurately assess mortality and outcome. Eighty-five patients were included. Indications for TAVI were nonoperable or surgically high-risk patients (LES>20%). Pro-BNP was measured 24h before the procedure. Cox proportional hazards model was used to evaluate clinical factors. The predictive accuracy of these Cox models was determined by using time-dependent receiver operating characteristic (ROC) curves. Pro-BNP levels (log-transformed) were significantly higher in non-survivors than in survivors at 30 days (3.36 ± 0.43 vs. 3.81 ± 0.43, p<0.004) and at the end of follow-up (3.34 ± 0.42 vs. 3.63 ± 0.48, p<0.011). Multivariate analysis revealed that only increased log pro-BNP levels were associated with higher mortality rate at short [hazard ratio (HR) (95% confidence intervals (CI)]=5.35 (1.74-16.5), p=0.003] and long-term follow-ups [HR=11 (CI: 1.51-81.3), p=0.018]. LES was not associated with increased mortality at either time point [HR=1.03 (CI: 0.95-1.10), p=0.483 and HR=1.03 (CI: 0.98-1.07), p=0.230, respectively]. At 30, 90, 180, and 365 days, the c-index was 0.72 for log pro-BNP and 0.63 for LES (p=0.044). Pre-procedure log transform of plasma pro-BNP levels are an independent and strong predictor of short- and long-term outcomes after TAVI and are more discriminatory than LES. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

  13. 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

  14. 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

  15. Shotgun protein sequencing.

    Energy Technology Data Exchange (ETDEWEB)

    Faulon, Jean-Loup Michel; Heffelfinger, Grant S.

    2009-06-01

    A novel experimental and computational technique based on multiple enzymatic digestion of a protein or protein mixture that reconstructs protein sequences from sequences of overlapping peptides is described in this SAND report. This approach, analogous to shotgun sequencing of DNA, is to be used to sequence alternative spliced proteins, to identify post-translational modifications, and to sequence genetically engineered proteins.

  16. Photodissociative Cross-Linking of Non-covalent Peptide-Peptide Ion Complexes in the Gas Phase

    Science.gov (United States)

    Nguyen, Huong T. H.; Andrikopoulos, Prokopis C.; Rulíšek, Lubomír; Shaffer, Christopher J.; Tureček, František

    2018-05-01

    We report a gas-phase UV photodissociation study investigating non-covalent interactions between neutral hydrophobic pentapeptides and peptide ions incorporating a diazirine-tagged photoleucine residue. Phenylalanine (Phe) and proline (Pro) were chosen as the conformation-affecting residues that were incorporated into a small library of neutral pentapeptides. Gas-phase ion-molecule complexes of these peptides with photo-labeled pentapeptides were subjected to photodissociation. Selective photocleavage of the diazirine ring at 355 nm formed short-lived carbene intermediates that underwent cross-linking by insertion into H-X bonds of the target peptide. The cross-link positions were established from collision-induced dissociation tandem mass spectra (CID-MS3) providing sequence information on the covalent adducts. Effects of the amino acid residue (Pro or Phe) and its position in the target peptide sequence were evaluated. For proline-containing peptides, interactions resulting in covalent cross-links in these complexes became more prominent as proline was moved towards the C-terminus of the target peptide sequence. The photocross-linking yields of phenylalanine-containing peptides depended on the position of both phenylalanine and photoleucine. Density functional theory calculations were used to assign structures of low-energy conformers of the (GLPMG + GLL*LK + H)+ complex. Born-Oppenheimer molecular dynamics trajectory calculations were used to capture the thermal motion in the complexes within 100 ps and determine close contacts between the incipient carbene and the H-X bonds in the target peptide. This provided atomic-level resolution of potential cross-links that aided spectra interpretation and was in agreement with experimental data. [Figure not available: see fulltext.

  17. GenHtr: a tool for comparative assessment of genetic heterogeneity in microbial genomes generated by massive short-read sequencing

    Directory of Open Access Journals (Sweden)

    Yu GongXin

    2010-10-01

    Full Text Available Abstract Background Microevolution is the study of short-term changes of alleles within a population and their effects on the phenotype of organisms. The result of the below-species-level evolution is heterogeneity, where populations consist of subpopulations with a large number of structural variations. Heterogeneity analysis is thus essential to our understanding of how selective and neutral forces shape bacterial populations over a short period of time. The Solexa Genome Analyzer, a next-generation sequencing platform, allows millions of short sequencing reads to be obtained with great accuracy, allowing for the ability to study the dynamics of the bacterial population at the whole genome level. The tool referred to as GenHtr was developed for genome-wide heterogeneity analysis. Results For particular bacterial strains, GenHtr relies on a set of Solexa short reads on given bacteria pathogens and their isogenic reference genome to identify heterogeneity sites, the chromosomal positions with multiple variants of genes in the bacterial population, and variations that occur in large gene families. GenHtr accomplishes this by building and comparatively analyzing genome-wide heterogeneity genotypes for both the newly sequenced genomes (using massive short-read sequencing and their isogenic reference (using simulated data. As proof of the concept, this approach was applied to SRX007711, the Solexa sequencing data for a newly sequenced Staphylococcus aureus subsp. USA300 cell line, and demonstrated that it could predict such multiple variants. They include multiple variants of genes critical in pathogenesis, e.g. genes encoding a LysR family transcriptional regulator, 23 S ribosomal RNA, and DNA mismatch repair protein MutS. The heterogeneity results in non-synonymous and nonsense mutations, leading to truncated proteins for both LysR and MutS. Conclusion GenHtr was developed for genome-wide heterogeneity analysis. Although it is much more time

  18. Biomedical Applications of Self-Assembling Peptides

    NARCIS (Netherlands)

    Radmalekshahi, Mazda; Lempsink, Ludwijn; Amidi, Maryam; Hennink, Wim E.; Mastrobattista, Enrico

    2016-01-01

    Self-assembling peptides have gained increasing attention as versatile molecules to generate diverse supramolecular structures with tunable functionality. Because of the possibility to integrate a wide range of functional domains into self-assembling peptides including cell attachment sequences,

  19. A rice chloroplast transit peptide sequence does not alter the cytoplasmic localization of sheep serotonin N-acetyltransferase expressed in transgenic rice plants.

    Science.gov (United States)

    Byeon, Yeong; Lee, Hyoung Yool; Lee, Kyungjin; Back, Kyoungwhan

    2014-09-01

    Ectopic overexpression of melatonin biosynthetic genes of animal origin has been used to generate melatonin-rich transgenic plants to examine the functional roles of melatonin in plants. However, the subcellular localization of these proteins expressed in the transgenic plants remains unknown. We studied the localization of sheep (Ovis aries) serotonin N-acetyltransferase (OaSNAT) and a translational fusion of a rice SNAT transit peptide to OaSNAT (TS:OaSNAT) in plants. Laser confocal microscopy analysis revealed that both OaSNAT and TS:OaSNAT proteins were localized to the cytoplasm even with the addition of the transit sequence to OaSNAT. Transgenic rice plants overexpressing the TS:OaSNAT fusion transgene exhibited high SNAT enzyme activity relative to untransformed wild-type plants, but lower activity than transgenic rice plants expressing the wild-type OaSNAT gene. Melatonin levels in both types of transgenic rice plant corresponded well with SNAT enzyme activity levels. The TS:OaSNAT transgenic lines exhibited increased seminal root growth relative to wild-type plants, but less than in the OaSNAT transgenic lines, confirming that melatonin promotes root growth. Seed-specific OaSNAT expression under the control of a rice prolamin promoter did not confer high levels of melatonin production in transgenic rice seeds compared with seeds from transgenic plants expressing OaSNAT under the control of the constitutive maize ubiquitin promoter. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  20. Drastic Effect of the Peptide Sequence on the Copper-Binding Properties of Tripeptides and the Electrochemical Behaviour of Their Copper(II) Complexes.

    Science.gov (United States)

    Mena, Silvia; Mirats, Andrea; Caballero, Ana B; Guirado, Gonzalo; Barrios, Leoní A; Teat, Simon J; Rodriguez-Santiago, Luis; Sodupe, Mariona; Gamez, Patrick

    2018-04-06

    The binding and electrochemical properties of the complexes Cu II -HAH, Cu II -HWH, Cu II -Ac-HWH, Cu II -HHW, and Cu II -WHH have been studied by using NMR and UV/Vis spectroscopies, CV, and density functional calculations. The results obtained highlight the importance of the peptidic sequence on the coordination properties and, consequently, on the redox properties of their Cu II complexes. For Cu II -HAH and Cu II -HWH, no cathodic processes are observed up to -1.2 V; that is, the complexes exhibit very high stability towards copper reduction. This behaviour is associated with the formation of very stable square-planar (5,5,6)-membered chelate rings (ATCUN motif), which enclose two deprotonated amides. In contrast, for non-ATCUN Cu II -Ac-HWH, Cu II -HHW complexes, simulations seem to indicate that only one deprotonated amide is enclosed in the coordination sphere. In these cases, the main electrochemical feature is a reductive irreversible one electron-transfer process from Cu II to Cu I , accompanied with structural changes of the metal coordination sphere and reprotonation of the amide. Finally, for Cu II -WHH, two major species have been detected: one at low pH (10) with an ATCUN motif, both species coexisting at intermediate pH. The present study shows that the use of CV, using glassy carbon as a working electrode, is an ideal and rapid tool for the determination of the redox properties of Cu II metallopeptides. © 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  1. Meat and fermented meat products as a source of bioactive peptides.

    Science.gov (United States)

    Stadnik, Joanna; Kęska, Paulina

    2015-01-01

    Bioactive peptides are short amino acid sequences, that upon release from the parent protein may play different physiological roles, including antioxidant, antihypertensive, antimicrobial, and other bioactivities. They have been identified from a range of foods, including those of animal origin, e.g., milk and muscle sources (with pork, beef, or chicken and various species of fish and marine organism). Bioactive peptides are encrypted within the sequence of the parent protein molecule and latent until released and activated by enzymatic proteolysis, e.g. during gastrointestinal digestion or food processing. Bioactive peptides derived from food sources have the potential for incorporation into functional foods and nutraceuticals. The aim of this paper is to present an overview of the muscle-derived bioactive peptides, especially those of fermented meats and the potential benefits of these bioactive compounds to human health.

  2. Antimicrobial Peptides from Plants

    Directory of Open Access Journals (Sweden)

    James P. Tam

    2015-11-01

    Full Text Available Plant antimicrobial peptides (AMPs have evolved differently from AMPs from other life forms. They are generally rich in cysteine residues which form multiple disulfides. In turn, the disulfides cross-braced plant AMPs as cystine-rich peptides to confer them with extraordinary high chemical, thermal and proteolytic stability. The cystine-rich or commonly known as cysteine-rich peptides (CRPs of plant AMPs are classified into families based on their sequence similarity, cysteine motifs that determine their distinctive disulfide bond patterns and tertiary structure fold. Cystine-rich plant AMP families include thionins, defensins, hevein-like peptides, knottin-type peptides (linear and cyclic, lipid transfer proteins, α-hairpinin and snakins family. In addition, there are AMPs which are rich in other amino acids. The ability of plant AMPs to organize into specific families with conserved structural folds that enable sequence variation of non-Cys residues encased in the same scaffold within a particular family to play multiple functions. Furthermore, the ability of plant AMPs to tolerate hypervariable sequences using a conserved scaffold provides diversity to recognize different targets by varying the sequence of the non-cysteine residues. These properties bode well for developing plant AMPs as potential therapeutics and for protection of crops through transgenic methods. This review provides an overview of the major families of plant AMPs, including their structures, functions, and putative mechanisms.

  3. A novel monoclonal antibody to a defined peptide epitope in MUC16

    DEFF Research Database (Denmark)

    Marcos-Silva, Lara; Ricardo, Sara; Chen, Kowa

    2015-01-01

    with the tandem-repeat region, their epitopes appear to be conformational dependent and not definable by a short peptide. Aberrant glycoforms of MUC16 may constitute promising targets for diagnostic and immunotherapeutic intervention, and it is important to develop well-defined immunogens for induction of potent...... immunodominant linear peptide epitopes within the tandem repeat. We developed one monoclonal antibody, 5E11, reactive with a minimum epitope with the sequence FNTTER. This sequence contains potential N- and O-glycosylation sites and, interestingly, glycosylation blocked binding of 5E11. In immunochemistry...

  4. Genome-wide detection of chromosomal rearrangements, indels, and mutations in circular chromosomes by short read sequencing

    DEFF Research Database (Denmark)

    Skovgaard, Ole; Bak, Mads; Løbner-Olesen, Anders

    2011-01-01

    a combination of WGS and genome copy number analysis, for the identification of mutations that suppress the growth deficiency imposed by excessive initiations from the Escherichia coli origin of replication, oriC. The E. coli chromosome, like the majority of bacterial chromosomes, is circular, and DNA...... replication is initiated by assembling two replication complexes at the origin, oriC. These complexes then replicate the chromosome bidirectionally toward the terminus, ter. In a population of growing cells, this results in a copy number gradient, so that origin-proximal sequences are more frequent than...... origin-distal sequences. Major rearrangements in the chromosome are, therefore, readily identified by changes in copy number, i.e., certain sequences become over- or under-represented. Of the eight mutations analyzed in detail here, six were found to affect a single gene only, one was a large chromosomal...

  5. Dataset of the molecular dynamics simulations of bilayers consisting of short amyloidogenic peptide VDSWNVLVAG from Bgl2p–glucantransferase of S. cerevisiae cell wall

    Directory of Open Access Journals (Sweden)

    Anna V. Glyakina

    2016-12-01

    Full Text Available The amyloidogenic peptide VDSWNVLVAG from Bgl2p–glucantransferase of Saccharomyces cerevisiae cell wall and its modifying analog VESWNVLVAG were taken for the construction of four types of bilayers which differ by orientation of the peptides in the layers and of the layers relative to each other. These bilayers were used as starting models for the molecular dynamics (MD at three charge states (neutral, pH3, and pH5. The changes of the fraction of secondary structure during 1 ns simulations were received for 96 MD trajectories. The data article contains the necessary information for the construction of models of β-strands organization in the oligomer structure. These results were used in the associated research article “Structural model of amyloid fibrils for amyloidogenic peptide from Bgl2p–glucantransferase of S. cerevisiae cell wall and its modifying analog. New morphology of amyloid fibrils” (Selivanova et al., 2016 [1].

  6. Dynamical System Modeling to Simulate Donor T Cell Response to Whole Exome Sequencing-Derived Recipient Peptides Demonstrates Different Alloreactivity Potential in HLA-Matched and -Mismatched Donor-Recipient Pairs.

    Science.gov (United States)

    Abdul Razzaq, Badar; Scalora, Allison; Koparde, Vishal N; Meier, Jeremy; Mahmood, Musa; Salman, Salman; Jameson-Lee, Max; Serrano, Myrna G; Sheth, Nihar; Voelkner, Mark; Kobulnicky, David J; Roberts, Catherine H; Ferreira-Gonzalez, Andrea; Manjili, Masoud H; Buck, Gregory A; Neale, Michael C; Toor, Amir A

    2016-05-01

    Immune reconstitution kinetics and subsequent clinical outcomes in HLA-matched recipients of allogeneic stem cell transplantation (SCT) are variable and difficult to predict. Considering SCT as a dynamical system may allow sequence differences across the exomes of the transplant donors and recipients to be used to simulate an alloreactive T cell response, which may allow better clinical outcome prediction. To accomplish this, whole exome sequencing was performed on 34 HLA-matched SCT donor-recipient pairs (DRPs) and the nucleotide sequence differences translated to peptides. The binding affinity of the peptides to the relevant HLA in each DRP was determined. The resulting array of peptide-HLA binding affinity values in each patient was considered as an operator modifying a hypothetical T cell repertoire vector, in which each T cell clone proliferates in accordance with the logistic equation of growth. Using an iterating system of matrices, each simulated T cell clone's growth was calculated with the steady-state population being proportional to the magnitude of the binding affinity of the driving HLA-peptide complex. Incorporating competition between T cell clones responding to different HLA-peptide complexes reproduces a number of features of clinically observed T cell clonal repertoire in the simulated repertoire, including sigmoidal growth kinetics of individual T cell clones and overall repertoire, Power Law clonal frequency distribution, increase in repertoire complexity over time with increasing clonal diversity, and alteration of clonal dominance when a different antigen array is encountered, such as in SCT. The simulated, alloreactive T cell repertoire was markedly different in HLA-matched DRPs. The patterns were differentiated by rate of growth and steady-state magnitude of the simulated T cell repertoire and demonstrate a possible correlation with survival. In conclusion, exome wide sequence differences in DRPs may allow simulation of donor alloreactive T

  7. In situ detection of a heat-shock regulatory element binding protein using a soluble short synthetic enhancer sequence

    Energy Technology Data Exchange (ETDEWEB)

    Harel-Bellan, A; Brini, A T; Farrar, W L [National Cancer Institute, Frederick, MD (USA); Ferris, D K [Program Resources, Inc., Frederick, MD (USA); Robin, P [Institut Gustave Roussy, Villejuif (France)

    1989-06-12

    In various studies, enhancer binding proteins have been successfully absorbed out by competing sequences inserted into plasmids, resulting in the inhibition of the plasmid expression. Theoretically, such a result could be achieved using synthetic enhancer sequences not inserted into plasmids. In this study, a double stranded DNA sequence corresponding to the human heat shock regulatory element was chemically synthesized. By in vitro retardation assays, the synthetic sequence was shown to bind specifically a protein in extracts from the human T cell line Jurkat. When the synthetic enhancer was electroporated into Jurkat cells, not only the enhancer was shown to remain undegraded into the cells for up to 2 days, but also its was shown to bind intracellularly a protein. The binding was specific and was modulated upon heat shock. Furthermore, the binding protein was shown to be of the expected molecular weight by UV crosslinking. However, when the synthetic enhancer element was co-electroporated with an HSP 70-CAT reporter construct, the expression of the reporter plasmid was consistently enhanced in the presence of the exogenous synthetic enhancer.

  8. Predicting binding within disordered protein regions to structurally characterised peptide-binding domains.

    Directory of Open Access Journals (Sweden)

    Waqasuddin Khan

    Full Text Available Disordered regions of proteins often bind to structured domains, mediating interactions within and between proteins. However, it is difficult to identify a priori the short disordered regions involved in binding. We set out to determine if docking such peptide regions to peptide binding domains would assist in these predictions.We assembled a redundancy reduced dataset of SLiM (Short Linear Motif containing proteins from the ELM database. We selected 84 sequences which had an associated PDB structures showing the SLiM bound to a protein receptor, where the SLiM was found within a 50 residue region of the protein sequence which was predicted to be disordered. First, we investigated the Vina docking scores of overlapping tripeptides from the 50 residue SLiM containing disordered regions of the protein sequence to the corresponding PDB domain. We found only weak discrimination of docking scores between peptides involved in binding and adjacent non-binding peptides in this context (AUC 0.58.Next, we trained a bidirectional recurrent neural network (BRNN using as input the protein sequence, predicted secondary structure, Vina docking score and predicted disorder score. The results were very promising (AUC 0.72 showing that multiple sources of information can be combined to produce results which are clearly superior to any single source.We conclude that the Vina docking score alone has only modest power to define the location of a peptide within a larger protein region known to contain it. However, combining this information with other knowledge (using machine learning methods clearly improves the identification of peptide binding regions within a protein sequence. This approach combining docking with machine learning is primarily a predictor of binding to peptide-binding sites, and is not intended as a predictor of specificity of binding to particular receptors.

  9. 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

  10. Differential effects of glucagon-like peptide-1 on microvascular recruitment and glucose metabolism in short- and long-term Insulin resistance

    DEFF Research Database (Denmark)

    Sjøberg, Kim Anker; Rattigan, Stephen; Jeppesen, Jacob Fuglsbjerg

    2015-01-01

    Acute infusion of glucagon-like-peptide-1 (GLP-1) has potent effects on blood flow distribution through the microcirculation in healthy humans and rats. High fat diet induces impairments in insulin-mediated microvascular recruitment (MVR) and muscle glucose uptake, and here we examined whether......-mediated glucose uptake in skeletal muscle by 90% (Prights...

  11. Short communication: Is consumption of a cheese rich in angiotensin-converting enzyme-inhibiting peptides, such as the Norwegian cheese Gamalost, associated with reduced blood pressure?

    Science.gov (United States)

    Nilsen, R; Pripp, A H; Høstmark, A T; Haug, A; Skeie, S

    2014-05-01

    Epidemiological and clinical studies have shown that angiotensin-converting enzyme (ACE)-inhibiting peptides derived from dairy products may decrease blood pressure. These peptides have been identified in many cheeses, and Gamalost, a traditional Norwegian cheese, is particularly rich in these peptides. The aim of this cross-sectional study was to examine whether frequency of Gamalost intake was associated with blood pressure in a Norwegian population sample. Blood pressure and other clinical measurements, including the factors of metabolic syndrome, were obtained from 168 participants (56% female, mean age = 51 yr) who completed a questionnaire about dietary habits and other health-related factors. Mean Gamalost intake was 2 servings per week. The prevalence of hypertension was 23.8% in the population, with mean systolic and diastolic blood pressures of 128 and 78 mmHg, respectively. Intake of Gamalost was inversely associated with systolic blood pressure. Each increase in frequency unit of Gamalost intake corresponded to a reduction in systolic blood pressure of 0.72 mmHg, after controlling for sex, age, education, waist circumference, physical activity, smoking status, and dairy food intake. Results from this study indicate that consumption of Gamalost (or other foods rich in ACE-inhibiting peptides) may reduce blood pressure. Copyright © 2014 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

  12. 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

  13. Sequential immunization with V3 peptides from primary human immunodeficiency virus type 1 produces cross-neutralizing antibodies against primary isolates with a matching narrow-neutralization sequence motif.

    Science.gov (United States)

    Eda, Yasuyuki; Takizawa, Mari; Murakami, Toshio; Maeda, Hiroaki; Kimachi, Kazuhiko; Yonemura, Hiroshi; Koyanagi, Satoshi; Shiosaki, Kouichi; Higuchi, Hirofumi; Makizumi, Keiichi; Nakashima, Toshihiro; Osatomi, Kiyoshi; Tokiyoshi, Sachio; Matsushita, Shuzo; Yamamoto, Naoki; Honda, Mitsuo

    2006-06-01

    An antibody response capable of neutralizing not only homologous but also heterologous forms of the CXCR4-tropic human immunodeficiency virus type 1 (HIV-1) MNp and CCR5-tropic primary isolate HIV-1 JR-CSF was achieved through sequential immunization with a combination of synthetic peptides representing HIV-1 Env V3 sequences from field and laboratory HIV-1 clade B isolates. In contrast, repeated immunization with a single V3 peptide generated antibodies that neutralized only type-specific laboratory-adapted homologous viruses. To determine whether the cross-neutralization response could be attributed to a cross-reactive antibody in the immunized animals, we isolated a monoclonal antibody, C25, which neutralized the heterologous primary viruses of HIV-1 clade B. Furthermore, we generated a humanized monoclonal antibody, KD-247, by transferring the genes of the complementary determining region of C25 into genes of the human V region of the antibody. KD-247 bound with high affinity to the "PGR" motif within the HIV-1 Env V3 tip region, and, among the established reference antibodies, it most effectively neutralized primary HIV-1 field isolates possessing the matching neutralization sequence motif, suggesting its promise for clinical applications involving passive immunizations. These results demonstrate that sequential immunization with B-cell epitope peptides may contribute to a humoral immune-based HIV vaccine strategy. Indeed, they help lay the groundwork for the development of HIV-1 vaccine strategies that use sequential immunization with biologically relevant peptides to overcome difficulties associated with otherwise poorly immunogenic epitopes.

  14. Photoperiod Regulates vgf-Derived Peptide Processing in Siberian Hamsters.

    Directory of Open Access Journals (Sweden)

    Barbara Noli

    Full Text Available VGF mRNA is induced in specific hypothalamic areas of the Siberian hamster upon exposure to short photoperiods, which is associated with a seasonal decrease in appetite and weight loss. Processing of VGF generates multiple bioactive peptides, so the objective of this study was to determine the profile of the VGF-derived peptides in the brain, pituitary and plasma from Siberian hamsters, and to establish whether differential processing might occur in the short day lean state versus long day fat. Antisera against short sequences at the C- or N- termini of proVGF, as well as against NERP-1, TPGH and TLQP peptides, were used for analyses of tissues, and both immunohistochemistry and enzyme linked immunosorbent assay (ELISA coupled with high-performance liquid (HPLC or gel chromatography were carried out. VGF peptide immunoreactivity was found within cortex cholinergic perikarya, in multiple hypothalamic nuclei, including those containing vasopressin, and in pituitary gonadotrophs. ELISA revealed that exposure to short day photoperiod led to a down-regulation of VGF immunoreactivity in the cortex, and a less pronounced decrease in the hypothalamus and pituitary, while the plasma VGF levels were not affected by the photoperiod. HPLC and gel chromatography both confirmed the presence of multiple VGF-derived peptides in these tissues, while gel chromatography showed the presence of the VGF precursor in all tissues tested except for the cortex. These observations are consistent with the view that VGF-derived peptides have pleiotropic actions related to changing photoperiod, possibly by regulating cholinergic systems in the cortex, vasopressin hypothalamic pathways, and the reproductive axis.

  15. "I know your name, but not your number"--Patients with verbal short-term memory deficits are impaired in learning sequences of digits.

    Science.gov (United States)

    Bormann, Tobias; Seyboth, Margret; Umarova, Roza; Weiller, Cornelius

    2015-06-01

    Studies on verbal learning in patients with impaired verbal short-term memory (vSTM) have revealed dissociations among types of verbal information. Patients with impaired vSTM are able to learn lists of known words but fail to acquire new word forms. This suggests that vSTM is involved in new word learning. The present study assessed both new word learning and the learning of digit sequences in two patients with impaired vSTM. In two experiments, participants were required to learn people's names, ages and professions, or their four digit 'phone numbers'. The STM patients were impaired on learning unknown family names and phone numbers, but managed to acquire other verbal information. In contrast, a patient with a severe verbal episodic memory impairment was impaired across information types. These results indicate verbal STM involvement in the learning of digit sequences. Copyright © 2015 Elsevier Ltd. All rights reserved.

  16. MRI in multiple sclerosis of the spinal cord: evaluation of fast short-tan inversion-recovery and spin-echo sequences

    International Nuclear Information System (INIS)

    Dietemann, J.L.; Thibaut-Menard, A.; Neugroschl, C.; Gillis, C.; Abu Eid, M.; Bogorin, A.; Warter, J.M.; Tranchant, C.

    2000-01-01

    We compared the sensitivity of T2-weighted spin-echo (FSE) and fast short-tau inversion-recovery (fSTIR) sequences in detection of multiple sclerosis of the spinal cord in 100 consecutive patients with clinically confirmed multiple sclerosis (MS); 86 patients underwent also brain MRI. In all, 310 focal lesions were detected on fSTIR and 212 on T2-weighted FSE, spinal cord lesions were seen better on fSTIR images, with a higher contrast between the lesion and the normal spinal cord. In 24 patients in whom cord plaques were shown with both sequences, the cranial study was normal or inconclusive. Assessment of spinal plaques can be particularly important when MRI of the brain is inconclusive, and in there situations fSTIR can be helpful. (orig.)

  17. A simple, flexible and efficient PCR-fusion/Gateway cloning procedure for gene fusion, site-directed mutagenesis, short sequence insertion and domain deletions and swaps

    Directory of Open Access Journals (Sweden)

    Etchells J Peter

    2009-10-01

    Full Text Available Abstract Background The progress and completion of various plant genome sequencing projects has paved the way for diverse functional genomic studies that involve cloning, modification and subsequent expression of target genes. This requires flexible and efficient procedures for generating binary vectors containing: gene fusions, variants from site-directed mutagenesis, addition of protein tags together with domain swaps and deletions. Furthermore, efficient cloning procedures, ideally high throughput, are essential for pyramiding of multiple gene constructs. Results Here, we present a simple, flexible and efficient PCR-fusion/Gateway cloning procedure for construction of binary vectors for a range of gene fusions or variants with single or multiple nucleotide substitutions, short sequence insertions, domain deletions and swaps. Results from selected applications of the procedure which include ORF fusion, introduction of Cys>Ser mutations, insertion of StrepII tag sequence and domain swaps for Arabidopsis secondary cell wall AtCesA genes are demonstrated. Conclusion The PCR-fusion/Gateway cloning procedure described provides an elegant, simple and efficient solution for a wide range of diverse and complicated cloning tasks. Through streamlined cloning of sets of gene fusions and modification variants into binary vectors for systematic functional studies of gene families, our method allows for efficient utilization of the growing sequence and expression data.

  18. Subtyping Salmonella enterica serovar enteritidis isolates from different sources by using sequence typing based on virulence genes and clustered regularly interspaced short palindromic repeats (CRISPRs).

    Science.gov (United States)

    Liu, Fenyun; Kariyawasam, Subhashinie; Jayarao, Bhushan M; Barrangou, Rodolphe; Gerner-Smidt, Peter; Ribot, Efrain M; Knabel, Stephen J; Dudley, Edward G

    2011-07-01

    Salmonella enterica subsp. enterica serovar Enteritidis is a major cause of food-borne salmonellosis in the United States. Two major food vehicles for S. Enteritidis are contaminated eggs and chicken meat. Improved subtyping methods are needed to accurately track specific strains of S. Enteritidis related to human salmonellosis throughout the chicken and egg food system. A sequence typing scheme based on virulence genes (fimH and sseL) and clustered regularly interspaced short palindromic repeats (CRISPRs)-CRISPR-including multi-virulence-locus sequence typing (designated CRISPR-MVLST)-was used to characterize 35 human clinical isolates, 46 chicken isolates, 24 egg isolates, and 63 hen house environment isolates of S. Enteritidis. A total of 27 sequence types (STs) were identified among the 167 isolates. CRISPR-MVLST identified three persistent and predominate STs circulating among U.S. human clinical isolates and chicken, egg, and hen house environmental isolates in Pennsylvania, and an ST that was found only in eggs and humans. It also identified a potential environment-specific sequence type. Moreover, cluster analysis based on fimH and sseL identified a number of clusters, of which several were found in more than one outbreak, as well as 11 singletons. Further research is needed to determine if CRISPR-MVLST might help identify the ecological origins of S. Enteritidis strains that contaminate chickens and eggs.

  19. Characteristic interpersonal behavior in dependent and avoidant personality disorder can be observed within very short interaction sequences.

    Science.gov (United States)

    Leising, Daniel; Sporberg, Doreen; Rehbein, Diana

    2006-08-01

    We present a behavior observation study of interpersonal behavior in 96 female subjects, who had been screened for the presence of dependent, avoidant, narcissistic and histrionic personality disorder features. Each subject took part in three short role-plays, taken from assertiveness training. Afterwards, both the subject and her role-play partner judged, how assertive the subject had been. Although observation time was very short, dependent and avoidant subjects could be easily identified from their overly submissive behavior in the role-plays. Histrionic and narcissistic subjects did not show distinctive interpersonal behavior. Contrary to a common belief, higher scores on some personality disorder (PD) scales were positively related to cross-situational variability of behavior. Results are discussed with regard to their implications for clinical diagnostics, therapy and the methodology of personality disorder research in general.

  20. Comparative mode of action of novel hybrid peptide CS-1a and its rearranged amphipathic analogue CS-2a.

    Science.gov (United States)

    Joshi, Seema; Bisht, Gopal S; Rawat, Diwan S; Maiti, Souvik; Pasha, Santosh

    2012-10-01

    Cell selective, naturally occurring, host defence cationic peptides present a good template for the design of novel peptides with the aim of achieving a short length with improved antimicrobial potency and selectivity. A novel, short peptide CS-1a (14 residues) was derived using a sequence hybridization approach on sarcotoxin I (39 residues) and cecropin B (35 residues). The sequence of CS-1a was rearranged to enhance amphipathicity with the help of a Schiffer-Edmundson diagram to obtain CS-2a. Both peptides showed good antibacterial activity in the concentration range 4-16 μg·mL(-1) against susceptible as well as drug-resistant bacterial strains, including the clinically relevant pathogens Acenatobacter sp. and methicillin-resistant Staphylococcus aureus. The major thrust of these peptides is their nonhaemolytic activity against human red blood cells up to a high concentration of 512 μg·mL(-1). Compared to CS-1a, amphipathic peptide CS-2a showed a more pronounced α-helical conformation, along with a better membrane insertion depth in bacterial mimic 1,2-dipalmitoyl-sn-glycero-3-phosphocholine/1,2-dipalmitoyl-sn-glycero-3-phospho-(1'-rac-glycerol) small unilamellar vesicles. With equivalent lipid-binding affinity, the two peptides assumed different pathways of membrane disruption, as demonstrated by calcein leakage and the results of transmission electron microscopy on model bacterial mimic large unilamellar vesicles. Extending the work from model membranes to intact Escherichia coli cells, differences in membrane perturbation were visible in microscopic images of peptide-treated E. coli. The present study describes two novel short peptides with potent activity, cell selectivity and divergent modes of action that will aid in the future design of peptides with better therapeutic potential. © 2012 The Authors Journal compilation © 2012 FEBS.

  1. Multiple regulatory mechanisms of hepatocyte growth factor expression in malignant cells with a short poly(dA) sequence in the HGF gene promoter.

    Science.gov (United States)

    Sakai, Kazuko; Takeda, Masayuki; Okamoto, Isamu; Nakagawa, Kazuhiko; Nishio, Kazuto

    2015-01-01

    Hepatocyte growth factor (HGF) expression is a poor prognostic factor in various types of cancer. Expression levels of HGF have been reported to be regulated by shorter poly(dA) sequences in the promoter region. In the present study, the poly(dA) mononucleotide tract in various types of human cancer cell lines was examined and compared with the HGF expression levels in those cells. Short deoxyadenosine repeat sequences were detected in five of the 55 cell lines used in the present study. The H69, IM95, CCK-81, Sui73 and H28 cells exhibited a truncated poly(dA) sequence in which the number of poly(dA) repeats was reduced by ≥5 bp. Two of the cell lines exhibited high HGF expression, determined by reverse transcription quantitative polymerase chain reaction and enzyme-linked immunosorbent assay. The CCK-81, Sui73 and H28 cells with shorter poly(dA) sequences exhibited low HGF expression. The cause of the suppression of HGF expression in the CCK-81, Sui73 and H28 cells was clarified by two approaches, suppression by methylation and single nucleotide polymorphisms in the HGF gene. Exposure to 5-Aza-dC, an inhibitor of DNA methyltransferase 1, induced an increased expression of HGF in the CCK-81 cells, but not in the other cells. Single-nucleotide polymorphism (SNP) rs72525097 in intron 1 was detected in the Sui73 and H28 cells. Taken together, it was found that the defect of poly(dA) in the HGF promoter was present in various types of cancer, including lung, stomach, colorectal, pancreas and mesothelioma. The present study proposes the negative regulation mechanisms by methylation and SNP in intron 1 of HGF for HGF expression in cancer cells with short poly(dA).

  2. RECOGNIZING REFERENCES TO PHYSICAL PLACES INSIDE SHORT TEXTS BY USING PATTERNS AS A SEQUENCE OF GRAMMATICAL CATEGORIES

    Directory of Open Access Journals (Sweden)

    A. Romero

    2017-09-01

    Full Text Available Collecting data by crowdsourcing is an explored trend to support database population and update. This kind of data is unstructured and comes from text, in particular text in social networks. Geographic database is a particular case of database that can be populated by crowdsourcing which can be done when people report some urban event in a social network by writing a short message. An event can describe an accident or a non-functioning device in the urban area. The authorities then need to read and to interpret the message to provide some help for injured people or to fix a problem in a device installed in the urban area like a light or a problem on road. Our main interest is located on working with short messages organized in a collection. Most of the messages do not have geographical coordinates. The messages can then be classified by text patterns describing a location. In fact, people use a text pattern to describe an urban location. Our work tries to identify patterns inside a short text and to indicate when it describes a location. When a pattern is identified our approach look to describe the place where the event is located. The source messages used are tweets reporting events from several Mexican cities.

  3. Recognizing References to Physical Places Inside Short Texts by Using Patterns as a Sequence of Grammatical Categories

    Science.gov (United States)

    Romero, A.; Sol, D.

    2017-09-01

    Collecting data by crowdsourcing is an explored trend to support database population and update. This kind of data is unstructured and comes from text, in particular text in social networks. Geographic database is a particular case of database that can be populated by crowdsourcing which can be done when people report some urban event in a social network by writing a short message. An event can describe an accident or a non-functioning device in the urban area. The authorities then need to read and to interpret the message to provide some help for injured people or to fix a problem in a device installed in the urban area like a light or a problem on road. Our main interest is located on working with short messages organized in a collection. Most of the messages do not have geographical coordinates. The messages can then be classified by text patterns describing a location. In fact, people use a text pattern to describe an urban location. Our work tries to identify patterns inside a short text and to indicate when it describes a location. When a pattern is identified our approach look to describe the place where the event is located. The source messages used are tweets reporting events from several Mexican cities.

  4. Polyclonal cell activity of a repeat peptide derived from the sequence of an 85-kilodalton surface protein of Trypanosoma cruzi trypomastigotes.

    Science.gov (United States)

    Pestel, J; Defoort, J P; Gras-Masse, H; Afchain, D; Capron, A; Tartar, A; Ouaissi, A

    1992-01-01

    Some in vitro and in vivo biological activities of an octadecapeptide derived from an 85-kDa surface protein of Trypanosoma cruzi trypomastigote were studied. The peptide coupled to a carrier protein induced the proliferative response of lymph node cells from mice immunized with various antigens. Moreover, sera from mice immunized with the coupled peptide were found to contain antibodies against a number of self and nonself antigens: fibronectin, bovine serum albumin, myosin, tetanus toxoid, ovalbumin, keyhole limpet hemocyanin, and DNA. These results are discussed in the context of Chagas' disease immunopathology. PMID:1730508

  5. 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

  6. ArrayPitope: Automated Analysis of Amino Acid Substitutions for Peptide Microarray-Based Antibody Epitope Mapping

    DEFF Research Database (Denmark)

    Hansen, Christian Skjødt; Østerbye, Thomas; Marcatili, Paolo

    2017-01-01

    -reactivity. B cell epitopes are typically classified as either linear epitopes, i.e. short consecutive segments from the protein sequence or conformational epitopes adapted through native protein folding. Recent advances in high-density peptide microarrays enable high-throughput, high-resolution identification...

  7. A synthetic peptide derived from the animo acid sequence of canine parvovirus structural proteins which defines a B cell epitope and elicits antiviral antibody in BALB c mice.

    NARCIS (Netherlands)

    G.F. Rimmelzwaan (Guus); J. Carlson; F.G.C.M. Uytdehaag (Fons); A.D.M.E. Osterhaus (Albert)

    1990-01-01

    textabstractSynthetic peptides, recombinant fusion proteins and mouse monoclonal antibodies were used to delineate a B cell epitope of the VP'2 structural protein of canine parvovirus (CPV). Although this epitope is not preferentially recognized in the normal antibody response to CPV, virus-specific

  8. Peptide sequencing and characterization of post-translational modifications by enhanced ion-charging and liquid chromatography electron-transfer dissociation tandem mass spectrometry

    DEFF Research Database (Denmark)

    Kjeldsen, Frank; Giessing, Anders; Ingrell, Christian R

    2007-01-01

    We have tested the effect of m-nitrobenzyl alcohol (m-NBA) as a method to increase the average charge state of protonated gas-phase molecular ions generated by ESI from tryptic peptides and phosphopeptides. Various concentrations of m-NBA were added to the mobile phases of a liquid chromatography...

  9. Expression of multiple transgenes from a single construct using viral 2A peptides in Drosophila.

    Directory of Open Access Journals (Sweden)

    Richard W Daniels

    Full Text Available Expression of multiple reporter or effector transgenes in the same cell from a single construct is increasingly necessary in various experimental paradigms. The discovery of short, virus-derived peptide sequences that mediate a ribosome-skipping event enables generation of multiple separate peptide products from one mRNA. Here we describe methods and vectors to facilitate easy production of polycistronic-like sequences utilizing these 2A peptides tailored for expression in Drosophila both in vitro and in vivo. We tested the separation efficiency of different viral 2A peptides in cultured Drosophila cells and in vivo and found that the 2A peptides from porcine teschovirus-1 (P2A and Thosea asigna virus (T2A worked best. To demonstrate the utility of this approach, we used the P2A peptide to co-express the red fluorescent protein tdTomato and the genetically-encoded calcium indicator GCaMP5G in larval motorneurons. This technique enabled ratiometric calcium imaging with motion correction allowing us to record synaptic activity at the neuromuscular junction in an intact larval preparation through the cuticle. The tools presented here should greatly facilitate the generation of 2A peptide-mediated expression of multiple transgenes in Drosophila.

  10. Structure of N-Terminal Sequence Asp-Ala-Glu-Phe-Arg-His-Asp-Ser of Aβ-Peptide with Phospholipase A2 from Venom of Andaman Cobra Sub-Species Naja naja sagittifera at 2.0 Å Resolution

    Directory of Open Access Journals (Sweden)

    Zeenat Mirza

    2014-03-01

    Full Text Available Alzheimer’s disease (AD is one of the most significant social and health burdens of the present century. Plaques formed by extracellular deposits of amyloid β (Aβ are the prime player of AD’s neuropathology. Studies have implicated the varied role of phospholipase A2 (PLA2 in brain where it contributes to neuronal growth and inflammatory response. Overall contour and chemical nature of the substrate-binding channel in the low molecular weight PLA2s are similar. This study involves the reductionist fragment-based approach to understand the structure adopted by N-terminal fragment of Alzheimer’s Aβ peptide in its complex with PLA2. In the current communication, we report the structure determined by X-ray crystallography of N-terminal sequence Asp-Ala-Glu-Phe-Arg-His-Asp-Ser (DAEFRHDS of Aβ-peptide with a Group I PLA2 purified from venom of Andaman Cobra sub-species Naja naja sagittifera at 2.0 Å resolution (Protein Data Bank (PDB Code: 3JQ5. This is probably the first attempt to structurally establish interaction between amyloid-β peptide fragment and hydrophobic substrate binding site of PLA2 involving H bond and van der Waals interactions. We speculate that higher affinity between Aβ and PLA2 has the therapeutic potential of decreasing the Aβ–Aβ interaction, thereby reducing the amyloid aggregation and plaque formation in AD.

  11. Amino-terminal domain of the v-fms oncogene product includes a functional signal peptide that directs synthesis of a transforming glycoprotein in the absence of feline leukemia virus gag sequences

    International Nuclear Information System (INIS)

    Wheeler, E.F.; Roussel, M.F.; Hampe, A.; Walker, M.H.; Fried, V.A.; Look, A.T.; Rettenmier, C.W.; Sherr, C.J.

    1986-01-01

    The nucleotide sequence of a 5' segment of the human genomic c-fms proto-oncogene suggested that recombination between feline leukemia virus and feline c-fms sequences might have occurred in a region encoding the 5' untranslated portion of c-fms mRNA. The polyprotein precursor gP180/sup gag-fms/ encoded by the McDonough strain of feline sarcoma virus was therefore predicted to contain 34 v-fms-coded amino acids derived from sequences of the c-fms gene that are not ordinarily translated from the proto-oncogene mRNA. The (gP180/sup gag-fms/) polyprotein was cotranslationally cleaved near the gag-fms junction to remove its gag gene-coded portion. Determination of the amino-terminal sequence of the resulting v-fms-coded glycoprotein, gp120/sup v-fms/, showed that the site of proteolysis corresponded to a predicted signal peptidase cleavage site within the c-fms gene product. Together, these analyses suggested that the linked gag sequences may not be necessary for expression of a biologically active v-fms gene product. The gag-fms sequences of feline sarcoma virus strain McDonough and the v-fms sequences alone were inserted into a murine retroviral vector containing a neomycin resistance gene. The authors conclude that a cryptic hydrophobic signal peptide sequence in v-fms was unmasked by gag deletion, thereby allowing the correct orientation and transport of the v-fms was unmasked by gag deletion, thereby allowing the correct orientation and transport of the v-fms gene product within membranous organelles. It seems likely that the proteolytic cleavage of gP180/gag-fms/ is mediated by signal peptidase and that the amino termini of gp140/sup v-fms/ and the c-fms gene product are identical

  12. Amino-terminal domain of the v-fms oncogene product includes a functional signal peptide that directs synthesis of a transforming glycoprotein in the absence of feline leukemia virus gag sequences

    Energy Technology Data Exchange (ETDEWEB)

    Wheeler, E.F.; Roussel, M.F.; Hampe, A.; Walker, M.H.; Fried, V.A.; Look, A.T.; Rettenmier, C.W.; Sherr, C.J.

    1986-08-01

    The nucleotide sequence of a 5' segment of the human genomic c-fms proto-oncogene suggested that recombination between feline leukemia virus and feline c-fms sequences might have occurred in a region encoding the 5' untranslated portion of c-fms mRNA. The polyprotein precursor gP180/sup gag-fms/ encoded by the McDonough strain of feline sarcoma virus was therefore predicted to contain 34 v-fms-coded amino acids derived from sequences of the c-fms gene that are not ordinarily translated from the proto-oncogene mRNA. The (gP180/sup gag-fms/) polyprotein was cotranslationally cleaved near the gag-fms junction to remove its gag gene-coded portion. Determination of the amino-terminal sequence of the resulting v-fms-coded glycoprotein, gp120/sup v-fms/, showed that the site of proteolysis corresponded to a predicted signal peptidase cleavage site within the c-fms gene product. Together, these analyses suggested that the linked gag sequences may not be necessary for expression of a biologically active v-fms gene product. The gag-fms sequences of feline sarcoma virus strain McDonough and the v-fms sequences alone were inserted into a murine retroviral vector containing a neomycin resistance gene. The authors conclude that a cryptic hydrophobic signal peptide sequence in v-fms was unmasked by gag deletion, thereby allowing the correct orientation and transport of the v-fms was unmasked by gag deletion, thereby allowing the correct orientation and transport of the v-fms gene product within membranous organelles. It seems likely that the proteolytic cleavage of gP180/gag-fms/ is mediated by signal peptidase and that the amino termini of gp140/sup v-fms/ and the c-fms gene product are identical.

  13. Survey of clustered regularly interspaced short palindromic repeats and their associated Cas proteins (CRISPR/Cas) systems in multiple sequenced strains of Klebsiella pneumoniae.

    Science.gov (United States)

    Ostria-Hernández, Martha Lorena; Sánchez-Vallejo, Carlos Javier; Ibarra, J Antonio; Castro-Escarpulli, Graciela

    2015-08-04

    In recent years the emergence of multidrug resistant Klebsiella pneumoniae strains has been an increasingly common event. This opportunistic species is one of the five main bacterial pathogens that cause hospital infections worldwide and multidrug resistance has been associated with the presence of high molecular weight plasmids. Plasmids are generally acquired through horizontal transfer and therefore is possible that systems that prevent the entry of foreign genetic material are inactive or absent. One of these systems is CRISPR/Cas. However, little is known regarding the clustered regularly interspaced short palindromic repeats and their associated Cas proteins (CRISPR/Cas) system in K. pneumoniae. The adaptive immune system CRISPR/Cas has been shown to limit the entry of foreign genetic elements into bacterial organisms and in some bacteria it has been shown to be involved in regulation of virulence genes. Thus in this work we used bioinformatics tools to determine the presence or absence of CRISPR/Cas systems in available K. pneumoniae genomes. The complete CRISPR/Cas system was identified in two out of the eight complete K. pneumoniae genomes sequences and in four out of the 44 available draft genomes sequences. The cas genes in these strains comprises eight cas genes similar to those found in Escherichia coli, suggesting they belong to the type I-E group, although their arrangement is slightly different. As for the CRISPR sequences, the average lengths of the direct repeats and spacers were 29 and 33 bp, respectively. BLAST searches demonstrated that 38 of the 116 spacer sequences (33%) are significantly similar to either plasmid, phage or genome sequences, while the remaining 78 sequences (67%) showed no significant similarity to other sequences. The region where the CRISPR/Cas systems were located is the same in all the Klebsiella genomes containing it, it has a syntenic architecture, and is located among genes encoding for proteins likely involved in

  14. Human Antimicrobial Peptides and Proteins

    Directory of Open Access Journals (Sweden)

    Guangshun Wang

    2014-05-01

    Full Text Available As the key components of innate immunity, human host defense antimicrobial peptides and proteins (AMPs play a critical role in warding off invading microbial pathogens. In addition, AMPs can possess other biological functions such as apoptosis, wound healing, and immune modulation. This article provides an overview on the identification, activity, 3D structure, and mechanism of action of human AMPs selected from the antimicrobial peptide database. Over 100 such peptides have been identified from a variety of tissues and epithelial surfaces, including skin, eyes, ears, mouths, gut, immune, nervous and urinary systems. These peptides vary from 10 to 150 amino acids with a net charge between −3 and +20 and a hydrophobic content below 60%. The sequence diversity enables human AMPs to adopt various 3D structures and to attack pathogens by different mechanisms. While α-defensin HD-6 can self-assemble on the bacterial surface into nanonets to entangle bacteria, both HNP-1 and β-defensin hBD-3 are able to block cell wall biosynthesis by binding to lipid II. Lysozyme is well-characterized to cleave bacterial cell wall polysaccharides but can also kill bacteria by a non-catalytic mechanism. The two hydrophobic domains in the long amphipathic α-helix of human cathelicidin LL-37 lays the basis for binding and disrupting the curved anionic bacterial membrane surfaces by forming pores or via the carpet model. Furthermore, dermcidin may serve as ion channel by forming a long helix-bundle structure. In addition, the C-type lectin RegIIIα can initially recognize bacterial peptidoglycans followed by pore formation in the membrane. Finally, histatin 5 and GAPDH(2-32 can enter microbial cells to exert their effects. It appears that granulysin enters cells and kills intracellular pathogens with the aid of pore-forming perforin. This arsenal of human defense proteins not only keeps us healthy but also inspires the development of a new generation of personalized

  15. 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.

  16. Rapid discovery of peptide capture candidates with demonstrated specificity for structurally similar toxins

    Science.gov (United States)

    Sarkes, Deborah A.; Hurley, Margaret M.; Coppock, Matthew B.; Farrell, Mikella E.; Pellegrino, Paul M.; Stratis-Cullum, Dimitra N.

    2016-05-01

    Peptides have emerged as viable alternatives to antibodies for molecular-based sensing due to their similarity in recognition ability despite their relative structural simplicity. Various methods for peptide capture reagent discovery exist, including phage display, yeast display, and bacterial display. One of the primary advantages of peptide discovery by bacterial display technology is the speed to candidate peptide capture agent, due to both rapid growth of bacteria and direct utilization of the sorted cells displaying each individual peptide for the subsequent round of biopanning. We have previously isolated peptide affinity reagents towards protective antigen of Bacillus anthracis using a commercially available automated magnetic sorting platform with improved enrichment as compared to manual magnetic sorting. In this work, we focus on adapting our automated biopanning method to a more challenging sort, to demonstrate the specificity possible with peptide capture agents. This was achieved using non-toxic, recombinant variants of ricin and abrin, RiVax and abrax, respectively, which are structurally similar Type II ribosomal inactivating proteins with significant sequence homology. After only two rounds of biopanning, enrichment of peptide capture candidates binding abrax but not RiVax was achieved as demonstrated by Fluorescence Activated Cell Sorting (FACS) studies. Further sorting optimization included negative sorting against RiVax, proper selection of autoMACS programs for specific sorting rounds, and using freshly made buffer and freshly thawed protein target for each round of biopanning for continued enrichment over all four rounds. Most of the resulting candidates from biopanning for abrax binding peptides were able to bind abrax but not RiVax, demonstrating that short peptide sequences can be highly specific even at this early discovery stage.

  17. 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.

  18. 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.

  19. Effects of short-term glucocorticoid deprivation on growth hormone (GH) response to GH-releasing peptide-6: Studies in normal men and in patients with adrenal insufficiency

    OpenAIRE

    Pinto, Ana Claudia de Assis Rocha [UNIFESP; Dias-da-Silva, Magnus Régios [UNIFESP; Martins, Manoel R. [UNIFESP; Brunner, Elisa [UNIFESP; Lengyel, Ana Maria Judith [UNIFESP

    2000-01-01

    There are no data in the literature about the effects of glucocorticoid deprivation on GH-releasing peptide-g (GHRP-6)-induced GH release. the aims of this study were to evaluate GH responsiveness to GHRP-6 1) after metyrapone administration in normal men, and 2) in patients with chronic hypocortisolism after glucocorticoid withdrawal for 72 h. in normal subjects, metyrapone ingestion did not alter significantly GH responsiveness to GHRP-6 [n = 8; peak, 39.3 +/- 7.1 mu g/L; area under the cur...

  20. Semisolid meal enriched in oat bran decreases plasma glucose and insulin levels, but does not change gastrointestinal peptide responses or short-term appetite in healthy subjects

    DEFF Research Database (Denmark)

    Juvonen, Kristiina R.; Salmenkallio-Marttila, Marjatta; Lyly, Marika

    2011-01-01

    types and amounts of DF exert are still poorly understood. METHODS AND RESULTS: We investigated the effects of wheat and oat brans alone and as combination in semisolid food matrix on postprandial appetite profile and gastrointestinal (GI) hormonal responses. Twenty healthy, normal-weight subjects (5...... including 5 g wheat bran DF + 5 g oat bran DF. Blood samples were drawn before and 15, 30, 45, 60, 90, 120 and 180 min after the test meals to determine plasma glucose, ghrelin, peptide YY (PYY) and serum insulin concentrations. Subjective profiles of appetite were assessed using visual analogue scales (VAS...

  1. Towards Liquid Chromatography Time-Scale Peptide Sequencing and Characterization of Post-Translational Modifications in the Negative-Ion Mode Using Electron Detachment Dissociation Tandem Mass Spectrometry

    DEFF Research Database (Denmark)

    Kjeldsen, Frank; Hørning, Ole B; Jensen, Søren S

    2008-01-01

    Electron detachment dissociation (EDD) of peptide poly-anions is gentle towards post-translational modifications (PTMs) and produces predictable and interpretable fragment ion types (a., x ions). However, EDD is considered an inefficient fragmentation technique and has not yet been implemented...... coverage and extended PTM characterization the new regime of EDD in combination with other ion-electron fragmentation techniques in the positive-ion mode is a step towards a more comprehensive strategy of analysis in proteome research....

  2. Molecular characterization of long direct repeat (LDR) sequences expressing a stable mRNA encoding for a 35-amino-acid cell-killing peptide and a cis-encoded small antisense RNA in Escherichia coli.

    Science.gov (United States)

    Kawano, Mitsuoki; Oshima, Taku; Kasai, Hiroaki; Mori, Hirotada

    2002-07-01

    Genome sequence analyses of Escherichia coli K-12 revealed four copies of long repetitive elements. These sequences are designated as long direct repeat (LDR) sequences. Three of the repeats (LDR-A, -B, -C), each approximately 500 bp in length, are located as tandem repeats at 27.4 min on the genetic map. Another copy (LDR-D), 450 bp in length and nearly identical to LDR-A, -B and -C, is located at 79.7 min, a position that is directly opposite the position of LDR-A, -B and -C. In this study, we demonstrate that LDR-D encodes a 35-amino-acid peptide, LdrD, the overexpression of which causes rapid cell killing and nucleoid condensation of the host cell. Northern blot and primer extension analysis showed constitutive transcription of a stable mRNA (approximately 370 nucleotides) encoding LdrD and an unstable cis-encoded antisense RNA (approximately 60 nucleotides), which functions as a trans-acting regulator of ldrD translation. We propose that LDR encodes a toxin-antitoxin module. LDR-homologous sequences are not pre-sent on any known plasmids but are conserved in Salmonella and other enterobacterial species.

  3. A non-accelerating foreshock sequence followed by a short period of quiescence for a large inland earthquake

    Science.gov (United States)

    Doi, I.; Kawakata, H.

    2012-12-01

    Laboratory experiments [e.g. Scholz, 1968; Lockner et al., 1992] and field observations [e.g. Dodge et al., 1996; Helmstetter and Sornette, 2003; Bouchon et al., 2011] have elucidated part of foreshock behavior and mechanism, but we cannot identify foreshocks while they are occurring. Recently, in Japan, a dense seismic network, Hi-net (High Sensitivity Seismograph Network), provides continuous waveform records for regional seismic events. The data from this network enable us to analyze small foreshocks which occur on long period time scales prior to a major event. We have an opportunity to grasp the more detailed pattern of foreshock generation. Using continuous waveforms recorded at a seismic station located in close proximity to the epicenter of the 2008 Iwate-Miyagi inland earthquake, we conducted a detailed investigation of its foreshocks. In addition to the two officially recognized foreshocks, calculation of cross-correlation coefficients between the continuous waveform record and one of the previously recognized foreshocks revealed that 20 micro foreshocks occurred within the same general area. Our analysis also shows that all of these foreshocks occurred within the same general area relative to the main event. Over the two week period leading up to the Iwate-Miyagi earthquake, such foreshocks only occurred during the last 45 minutes, specifically over a 35 minute period followed by a 10 minute period of quiescence just before the mainshock. We found no evidence of acceleration of this foreshock sequence. Rock fracturing experiments using a constant loading rate or creep tests have consistently shown that the occurrence rate of small fracturing events (acoustic emissions; AEs) increases before the main rupture [Scholz, 1968]. This accelerative pattern of preceding events was recognized in case of the 1999 Izmit earthquake [Bouchon et al., 2011]. Large earthquakes however need not be accompanied by acceleration of foreshocks if a given fault's host rock

  4. 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

  5. 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...

  6. Identification of PrP sequences essential for the interaction between the PrP polymers and Aβ peptide in a yeast-based assay

    OpenAIRE

    Rubel, Aleksandr A; Ryzhova, Tatyana A; Antonets, Kirill S; Chernoff, Yury O; Galkin, Alexey P

    2013-01-01

    Alzheimer disease is associated with the accumulation of oligomeric amyloid β peptide (Aβ), accompanied by synaptic dysfunction and neuronal death. Polymeric form of prion protein (PrP), PrPSc, is implicated in transmissible spongiform encephalopathies (TSEs). Recently, it was shown that the monomeric cellular form of PrP (PrPC), located on the neuron surface, binds Aβ oligomers (and possibly other β-rich conformers) via the PrP23–27 and PrP90–110 segments, acting as Aβ receptor. On the other...

  7. Establishment of a root proteome reference map for the model legume Medicago truncatula using the expressed sequence tag database for peptide mass fingerprinting

    DEFF Research Database (Denmark)

    Mathesius, U; Keijzers, Guido; Natera, S H

    2001-01-01

    legume for the study of nodulation-related genes and proteins. Over 2,500 root proteins could be displayed reproducibly across an isoelectric focussing range of 4-7. We analysed 485 proteins by peptide mass fingerprinting, and 179 of those were identified by matching against the current M. truncatula....... This proteome map will be updated continuously (http://semele.anu.edu.au/2d/2d.html) and will be a powerful tool for investigating the molecular mechanisms of root symbioses in legumes....

  8. 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....

  9. Acylation of Therapeutic Peptides

    DEFF Research Database (Denmark)

    Trier, Sofie; Henriksen, Jonas Rosager; Jensen, Simon Bjerregaard

    ) , which promotes intestinal growth and is used to treat bowel disorders such as inflammatory bowel diseases and short bowel syndrome, and the 32 amino acid salmon calcitonin (sCT), which lowers blood calcium and is employed in the treatment of post-menopausal osteoporosis and hypercalcemia. The two...... peptides are similar in size and structure, but oppositely charged at physiological pH. Both peptides were acylated with linear acyl chains of systematically increasing length, where sCT was furthermore acylated at two different positions on the peptide backbone. For GLP-2, we found that increasing acyl...... remained optimal overall. The results indicate that rational acylation of GLP-2 can increase its in vitro intestinal absorption, alone or in combination with permeation enhancers, and are consistent with the initial project hypothesis. For sCT, an unpredicted effect of acylation largely superseded...

  10. 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.

  11. Femtomolar Ln(III) affinity in peptide-based ligands containing unnatural chelating amino acids.

    Science.gov (United States)

    Niedźwiecka, Agnieszka; Cisnetti, Federico; Lebrun, Colette; Delangle, Pascale

    2012-05-07

    The incorporation of unnatural chelating amino acids in short peptide sequences leads to lanthanide-binding peptides with a higher stability than sequences built exclusively from natural residues. In particular, the hexadentate peptide P(22), which incorporates two unnatural amino acids Ada(2) with aminodiacetate chelating arms, showed picomolar affinity for Tb(3+). To design peptides with higher denticity, expected to show higher affinity for Ln(3+), we synthesized the novel unnatural amino acid Ed3a(2) which carries an ethylenediamine triacetate side-chain and affords a pentadentate coordination site. The synthesis of the derivative Fmoc-Ed3a(2)(tBu)(3)-OH, with appropriate protecting groups for direct use in the solid phase peptide synthesis (Fmoc strategy), is described. The two high denticity peptides P(HD2) (Ac-Trp-Ed3a(2)-Pro-Gly-Ada(2)-Gly-NH(2)) and P(HD5) (Ac-Trp-Ada(2)-Pro-Gly-Ed3a(2)-Gly-NH(2)) led to octadentate Tb(3+) complexes with femtomolar stability in water. The position of the high denticity amino acid Ed3a(2) in the hexapeptide sequence appears to be critical for the control of the metal complex speciation. Whereas P(HD5) promotes the formation of polymetallic species in excess of Ln(3+), P(HD2) forms exclusively the mononuclear complex. The octadentate coordination of Tb(3+) by both P(HD) leads to total dehydration of the metal ion in the mononuclear complexes with long luminescence lifetimes (>2 ms). Hence, we demonstrated that unnatural amino acids carrying polyaminocarboxylate side-chains are interesting building blocks to design high affinity Ln-binding peptides. In particular the novel peptide P(HD2) forms a unique octadentate Tb(3+) complex with femtomolar stability in water and an improvement of the luminescence properties with respect to the trisaquo TbP(22) complex by a factor of 4.

  12. Protection against the Metabolic Syndrome by Guar Gum-Derived Short-Chain Fatty Acids Depends on Peroxisome Proliferator-Activated Receptor. and Glucagon-Like Peptide-1

    NARCIS (Netherlands)

    den Besten, Gijs; Gerding, Albert; van Dijk, Theo H.; Ciapaite, Jolita; Bleeker, Aycha; van Eunen, Karen; Havinga, Rick; Groen, Albert K.; Reijngoud, Dirk-Jan; Bakker, Barbara M.

    2015-01-01

    The dietary fiber guar gum has beneficial effects on obesity, hyperglycemia and hypercholesterolemia in both humans and rodents. The major products of colonic fermentation of dietary fiber, the short-chain fatty acids (SCFAs), have been suggested to play an important role. Recently, we showed that

  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. Dante-Z sequence as selective impulsion in high field mono and multidimensional NMR. Application to the study of proteins, peptides and their interactions

    International Nuclear Information System (INIS)

    Roumestand, C.; Toma, F.

    1992-01-01

    DANTE-Z is a simple and efficient way for NMR spectral selection. We present here different applications of DANTE-Z in high-resolution NMR of peptides and proteins. We have been using proton selective excitation by DANTE-Z to perform 1D-correlated (homo- or heteronuclear) experiments corresponding to one line of either 2D or 3D experiments. Following the same scheme, we could also edit planes of 3D experiments by concatenating 1D-correlated experiments with conventional 2D experiments. In the heteronuclear case (i.e. 1 H, 31 P), we could also edit planes of a 4D experiment by the simultaneous selection of 1 H and the X nucleus. Owing to the favourable excitation profile of DANTE-Z, we used it successfully for topological excitations (spectral width from 150 Hz up to 1500 Hz) in 'semi-soft'-2D experiments and 'soft'-2D experiment. These applications are illustrated by the results obtained at 600 MHz on a protein and a phosphonamide peptide

  15. Vaccatides: Antifungal Glutamine-Rich Hevein-Like Peptides from Vaccaria hispanica

    Directory of Open Access Journals (Sweden)

    Ka H. Wong

    2017-06-01

    Full Text Available Hevein and hevein-like peptides are disulfide-constrained chitin-binding cysteine-rich peptides. They are divided into three subfamilies, 6C-, 8C-, and 10C-hevein-like peptides, based on the number of cysteine residues. In addition, hevein-like peptides can exist in two forms, short and long. The long C-terminal form found in hevein and 10C-hevein-like peptides contain a C-terminal protein cargo. In contrast, the short form without a protein cargo is found in all three subfamilies. Here, we report the discovery and characterization of two novel glutamine-rich and protein cargo-free 8C-hevein-like peptides, vaccatides vH1 and vH2, from Vaccaria hispanica of the Caryophyllaceae family. Proteomic analyses showed that the vaccatides are 40–41 amino acids in length and contain a chitin-binding domain. NMR determination revealed that vaccatide vH2 displays a highly compact structure with a N-terminal cystine knot and an addition C-terminal disulfide bond. Stability studies showed that this compact structure renders vaccatide vH2 resistant to thermal, chemical and proteolytic degradation. The chitin-binding vH2 was shown to inhibit the mycelium growth of four phyto-pathogenic fungal strains with IC50 values in the micromolar range. Our findings show that vaccatides represent a new family of 8C-hevein-like peptides, which are protein cargo-free and glutamine-rich, characteristics that differentiate them from the prototypic hevein and the 10C-hevein-like peptides. In summary, this study enriches the existing library of hevein-like peptides and provides insight into their molecular diversity in sequence, structure and biosynthesis. Additionally, their highly disulfide-constrained structure could be used as a scaffold for developing metabolically and orally active peptidyl therapeutics.

  16. Characterization of Erwinia amylovora strains from different host plants using repetitive-sequences PCR analysis, and restriction fragment length polymorphism and short-sequence DNA repeats of plasmid pEA29.

    Science.gov (United States)

    Barionovi, D; Giorgi, S; Stoeger, A R; Ruppitsch, W; Scortichini, M

    2006-05-01

    The three main aims of the study were the assessment of the genetic relationship between a deviating Erwinia amylovora strain isolated from Amelanchier sp. (Maloideae) grown in Canada and other strains from Maloideae and Rosoideae, the investigation of the variability of the PstI fragment of the pEA29 plasmid using restriction fragment length polymorphism (RFLP) analysis and the determination of the number of short-sequence DNA repeats (SSR) by DNA sequence analysis in representative strains. Ninety-three strains obtained from 12 plant genera and different geographical locations were examined by repetitive-sequences PCR using Enterobacterial Repetitive Intergenic Consensus, BOX and Repetitive Extragenic Palindromic primer sets. Upon the unweighted pair group method with arithmetic mean analysis, a deviating strain from Amelanchier sp. was analysed using amplified ribosomal DNA restriction analysis (ARDRA) analysis and the sequencing of the 16S rDNA gene. This strain showed 99% similarity to other E. amylovora strains in the 16S gene and the same banding pattern with ARDRA. The RFLP analysis of pEA29 plasmid using MspI and Sau3A restriction enzymes showed a higher variability than that previously observed and no clear-cut grouping of the strains was possible. The number of SSR units reiterated two to 12 times. The strains obtained from pear orchards showing for the first time symptoms of fire blight had a low number of SSR units. The strains from Maloideae exhibit a wider genetic variability than previously thought. The RFLP analysis of a fragment of the pEA29 plasmid would not seem a reliable method for typing E. amylovora strains. A low number of SSR units was observed with first epidemics of fire blight. The current detection techniques are mainly based on the genetic similarities observed within the strains from the cultivated tree-fruit crops. For a more reliable detection of the fire blight pathogen also in wild and ornamentals Rosaceous plants the genetic

  17. Efficient expression of a soluble lipid transfer protein (LTP) of Platanus orientalis using short peptide tags and structural comparison with the natural form.

    Science.gov (United States)

    Salari, Farhad; Vahedi, Fatemeh; Chamani, Jamshidkhan; Varasteh, Abdolreza; Ketabdar, Hanieh; Sankian, Mojtaba

    2015-01-01

    Successful recombinant allergen-based immunotherapy has drawn a great deal of attention to use recombinant allergens for new therapeutic and/or diagnostic strategies. The Escherichia coli expression system is frequently used to produce recombinant allergens; however, protein expression in E. coli often results in inclusion bodies. Here, we focused on the expression of two recombinant soluble forms of Pla or 3 using solubility-enhancing peptide tags, human immune deficiency virus type 1 transactivator of transcription core domain and poly-arginine-lysine: rTAT-Pla or 3 and rPoly-Arg-Lys-Pla or 3. Structural characteristics and IgE reactivity of purified recombinant proteins were compared with natural Pla or 3 (nPla or 3) isolated from Platanus orientalis using circular dichroism spectra, fluorescence spectroscopy, and immunoblotting. Likewise, intrinsic viscosity and Stokes radius of the natural and recombinant Pla or 3 allergens were determined to analyze structural compactness in aqueous media. The results indicate high-level solubility and efficient expression of the fusion proteins (rTAT-Pla or 3 and rPoly-Arg-Lys-Pla or 3) compared with the wild-type recombinant. Furthermore, the similar structural characteristics and IgE-binding activities of the fusion proteins to nPla or 3 provide a promising tool for allergy diagnosis and treatment. © 2014 International Union of Biochemistry and Molecular Biology, Inc.

  18. The glucagon-like peptide-1 receptor in the ventromedial hypothalamus reduces short-term food intake in male mice by regulating nutrient sensor activity.

    Science.gov (United States)

    Burmeister, Melissa A; Brown, Jacob D; Ayala, Jennifer E; Stoffers, Doris A; Sandoval, Darleen A; Seeley, Randy J; Ayala, Julio E

    2017-12-01

    Pharmacological activation of the glucagon-like peptide-1 receptor (GLP-1R) in the ventromedial hypothalamus (VMH) reduces food intake. Here, we assessed whether suppression of food intake by GLP-1R agonists (GLP-1RA) in this region is dependent on AMP-activated protein kinase (AMPK) and mammalian target of rapamycin (mTOR). We found that pharmacological inhibition of glycolysis, and thus activation of AMPK, in the VMH attenuates the anorectic effect of the GLP-1R agonist exendin-4 (Ex4), indicating that glucose metabolism and inhibition of AMPK are both required for this effect. Furthermore, we found that Ex4-mediated anorexia in the VMH involved mTOR but not acetyl-CoA carboxylase, two downstream targets of AMPK. We support this by showing that Ex4 activates mTOR signaling in the VMH and Chinese hamster ovary (CHO)-K1 cells. In contrast to the clear acute pharmacological impact of the these receptors on food intake, knockdown of the VMH Glp1r conferred no changes in energy balance in either chow- or high-fat-diet-fed mice, and the acute anorectic and glucose tolerance effects of peripherally dosed GLP-1RA were preserved. These results show that the VMH GLP-1R regulates food intake by engaging key nutrient sensors but is dispensable for the effects of GLP-1RA on nutrient homeostasis. Copyright © 2017 the American Physiological Society.

  19. Ultrasmall Peptides Self-Assemble into Diverse Nanostructures: Morphological Evaluation and Potential Implications

    Directory of Open Access Journals (Sweden)

    Charlotte A.E. Hauser

    2011-09-01

    Full Text Available In this study, we perform a morphological evaluation of the diverse nanostructures formed by varying concentration and amino acid sequence of a unique class of ultrasmall self-assembling peptides. We modified these peptides by replacing the aliphatic amino acid at the C-aliphatic terminus with different aromatic amino acids. We tracked the effect of introducing aromatic residues on self-assembly and morphology of resulting nanostructures. Whereas aliphatic peptides formed long, helical fibers that entangle into meshes and entrap >99.9% water, the modified peptides contrastingly formed short, straight fibers with a flat morphology. No helical fibers were observed for the modified peptides. For the aliphatic peptides at low concentrations, different supramolecular assemblies such as hollow nanospheres and membrane blebs were found. Since the ultrasmall peptides are made of simple, aliphatic amino acids, considered to have existed in the primordial soup, study of these supramolecular assemblies could be relevant to understanding chemical evolution leading to the origin of life on Earth. In particular, we propose a variety of potential applications in bioengineering and nanotechnology for the diverse self-assembled nanostructures.

  20. ¹¹¹In-Bn-DTPA-nimotuzumab with/without modification with nuclear translocation sequence (NLS) peptides: an Auger electron-emitting radioimmunotherapeutic agent for EGFR-positive and trastuzumab (Herceptin)-resistant breast cancer.

    Science.gov (United States)

    Fasih, Aisha; Fonge, Humphrey; Cai, Zhongli; Leyton, Jeffrey V; Tikhomirov, Ilia; Done, Susan J; Reilly, Raymond M

    2012-08-01

    Increased expression of epidermal growth factor receptors (EGFR) in breast cancer (BC) is often associated with trastuzumab (Herceptin)-resistant forms of the disease and represents an attractive target for novel therapies. Nimotuzumab is a humanized IgG(1) monoclonal antibody that is in clinical trials for treatment of EGFR-overexpressing malignancies. We show here that nimotuzumab derivatized with benzylisothiocyanate diethylenetriaminepentaacetic acid for labelling with the subcellular range Auger electron-emitter, (111)In and modified with nuclear translocation sequence (NLS) peptides ((111)In-NLS-Bn-DTPA-nimotuzumab) was bound, internalized and transported to the nucleus of EGFR-positive BC cells. Emission of Auger electrons in close proximity to the nucleus caused multiple DNA double-strand breaks which diminished the clonogenic survival (CS) of MDA-MB-468 cells that have high EGFR density (2.4 × 10(6) receptors/cell) to less than 3 %. (111)In-Bn-DTPA-nimotuzumab without NLS peptide modification was sevenfold less effective for killing MDA-MB-468 cells. (111)In-Bn-DTPA-nimotuzumab with/without NLS peptide modification were equivalently cytotoxic to MDA-MB-231 and TrR1 BC cells that have moderate EGFR density (5.4 × 10(5) or 4.2 × 10(5) receptors/cell, respectively) reducing their CS by twofold. MDA-MB-231 cells have intrinsic trastuzumab resistance due to low HER2 density, whereas TrR1 cells have acquired resistance despite HER2 overexpression. Biodistribution and microSPECT/CT imaging revealed that (111)In-NLS-Bn-DTPA-nimotuzumab exhibited more rapid elimination from the blood and lower tumour uptake than (111)In-Bn-DTPA-nimotuzumab. Tumour uptake of the radioimmunoconjugates in mice with MDA-MB-468 xenografts was high (8-16 % injected dose/g) and was blocked by administration of an excess of unlabelled nimotuzumab, demonstrating EGFR specificity. We conclude that (111)In-Bn-DTPA-nimotuzumab with/without NLS peptide modification are promising Auger

  1. Structure of genes for dermaseptins B, antimicrobial peptides from frog skin. Exon 1-encoded prepropeptide is conserved in genes for peptides of highly different structures and activities.

    Science.gov (United States)

    Vouille, V; Amiche, M; Nicolas, P

    1997-09-01

    We cloned the genes of two members of the dermaseptin family, broad-spectrum antimicrobial peptides isolated from the skin of the arboreal frog Phyllomedusa bicolor. The dermaseptin gene Drg2 has a 2-exon coding structure interrupted by a small 137-bp intron, wherein exon 1 encoded a 22-residue hydrophobic signal peptide and the first three amino acids of the acidic propiece; exon 2 contained the 18 additional acidic residues of the propiece plus a typical prohormone processing signal Lys-Arg and a 32-residue dermaseptin progenitor sequence. The dermaseptin genes Drg2 and Drg1g2 have conserved sequences at both untranslated ends and in the first and second coding exons. In contrast, Drg1g2 comprises a third coding exon for a short version of the acidic propiece and a second dermaseptin progenitor sequence. Structural conservation between the two genes suggests that Drg1g2 arose recently from an ancestral Drg2-like gene through amplification of part of the second coding exon and 3'-untranslated region. Analysis of the cDNAs coding precursors for several frog skin peptides of highly different structures and activities demonstrates that the signal peptides and part of the acidic propieces are encoded by conserved nucleotides encompassed by the first coding exon of the dermaseptin genes. The organization of the genes that belong to this family, with the signal peptide and the progenitor sequence on separate exons, permits strikingly different peptides to be directed into the secretory pathway. The recruitment of such a homologous 'secretory' exon by otherwise non-homologous genes may have been an early event in the evolution of amphibian.

  2. Identification of Four-Jointed Box 1 (FJX1-Specific Peptides for Immunotherapy of Nasopharyngeal Carcinoma.

    Directory of Open Access Journals (Sweden)

    San Jiun Chai

    Full Text Available Nasopharyngeal carcinoma (NPC is highly prevalent in South East Asia and China. The poor outcome is due to late presentation, recurrence, distant metastasis and limited therapeutic options. For improved treatment outcome, immunotherapeutic approaches focusing on dendritic and autologous cytotoxic T-cell based therapies have been developed, but cost and infrastructure remain barriers for implementing these in low-resource settings. As our prior observations had found that four-jointed box 1 (FJX1, a tumor antigen, is overexpressed in NPCs, we investigated if short 9-20 amino acid sequence specific peptides matching to FJX1 requiring only intramuscular immunization to train host immune systems would be a better treatment option for this disease. Thus, we designed 8 FJX1-specific peptides and implemented an assay system to first, assess the binding of these peptides to HLA-A2 molecules on T2 cells. After, ELISPOT assays were used to determine the peptides immunogenicity and ability to induce potential cytotoxicity activity towards cancer cells. Also, T-cell proliferation assay was used to evaluate the potential of MHC class II peptides to stimulate the expansion of isolated T-cells. Our results demonstrate that these peptides are immunogenic and peptide stimulated T-cells were able to induce peptide-specific cytolytic activity specifically against FJX1-expressing cancer cells. In addition, we demonstrated that the MHC class II peptides were capable of inducing T-cell proliferation. Our results suggest that these peptides are capable of inducing specific cytotoxic cytokines secretion against FJX1-expressing cancer cells and serve as a potential vaccine-based therapy for NPC patients.

  3. Comparison of trapping profiles between d-peptides and glutathione in the identification of reactive metabolites

    Directory of Open Access Journals (Sweden)

    Jaana E. Laine

    2015-01-01

    Full Text Available Qualitative trapping profile of reactive metabolites arising from six structurally different compounds was tested with three different d-peptide isomers (Peptide 1, gly–tyr–pro–cys–pro–his-pro; Peptide 2, gly–tyr–pro–ala–pro–his–pro; Peptide 3, gly–tyr–arg–pro–cys–pro–his–lys–pro and glutathione (GSH using mouse and human liver microsomes as the biocatalyst. The test compounds were classified either as clinically “safe” (amlodipine, caffeine, ibuprofen, or clinically as “risky” (clozapine, nimesulide, ticlopidine; i.e., associated with severe clinical toxicity outcomes. Our working hypothesis was as follows: could the use of short different amino acid sequence containing d-peptides in adduct detection confer any add-on value to that obtained with GSH? All “risky” agents’ resulted in the formation of several GSH adducts in the incubation mixture and with at least one peptide adduct with both microsomal preparations. Amlodipine did not form any adducts with any of the trapping agents. No GSH and peptide 2 and 3 adducts were found with caffeine, but with peptide 1 one adduct with human liver microsomes was detected. Ibuprofen produced one Peptide 1-adduct with human and mouse liver microsomes but not with GSH. In conclusion, GSH still remains the gold trapping standard for reactive metabolites. However, targeted d-peptides could provide additional information about protein binding potential of electrophilic agents, but their clinical significance needs to be clarified using a wider spectrum of chemicals together with other safety estimates.

  4. Identification of Four-Jointed Box 1 (FJX1)-Specific Peptides for Immunotherapy of Nasopharyngeal Carcinoma

    Science.gov (United States)

    Chai, San Jiun; Yap, Yoke Yeow; Foo, Yoke Ching; Yap, Lee Fah; Ponniah, Sathibalan; Teo, Soo Hwang; Cheong, Sok Ching; Patel, Vyomesh; Lim, Kue Peng

    2015-01-01

    Nasopharyngeal carcinoma (NPC) is highly prevalent in South East Asia and China. The poor outcome is due to late presentation, recurrence, distant metastasis and limited therapeutic options. For improved treatment outcome, immunotherapeutic approaches focusing on dendritic and autologous cytotoxic T-cell based therapies have been developed, but cost and infrastructure remain barriers for implementing these in low-resource settings. As our prior observations had found that four-jointed box 1 (FJX1), a tumor antigen, is overexpressed in NPCs, we investigated if short 9–20 amino acid sequence specific peptides matching to FJX1 requiring only intramuscular immunization to train host immune systems would be a better treatment option for this disease. Thus, we designed 8 FJX1-specific peptides and implemented an assay system to first, assess the binding of these peptides to HLA-A2 molecules on T2 cells. After, ELISPOT assays were used to determine the peptides immunogenicity and ability to induce potential cytotoxicity activity towards cancer cells. Also, T-cell proliferation assay was used to evaluate the potential of MHC class II peptides to stimulate the expansion of isolated T-cells. Our results demonstrate that these peptides are immunogenic and peptide stimulated T-cells were able to induce peptide-specific cytolytic activity specifically against FJX1-expressing cancer cells. In addition, we demonstrated that the MHC class II peptides were capable of inducing T-cell proliferation. Our results suggest that these peptides are capable of inducing specific cytotoxic cytokines secretion against FJX1-expressing cancer cells and serve as a potential vaccine-based therapy for NPC patients. PMID:26536470

  5. Chimeric opioid peptides: tools for identifying opioid receptor types.

    OpenAIRE

    Xie, G X; Miyajima, A; Yokota, T; Arai, K; Goldstein, A

    1990-01-01

    We synthesized several chimeric peptides in which the N-terminal nine residues of dynorphin-32, a peptide selective for the kappa opioid receptor, were replaced by opioid peptides selective for other opioid receptor types. Each chimeric peptide retained the high affinity and type selectivity characteristic of its N-terminal sequence. The common C-terminal two-thirds of the chimeric peptides served as an epitope recognized by the same monoclonal antibody. When bound to receptors on a cell surf...

  6. Detection of optic nerve atrophy following a single episode of unilateral optic neuritis by MRI using a fat-saturated short-echo fast FLAIR sequence

    International Nuclear Information System (INIS)

    Hickman, S.J.; Brex, P.A.; Silver, N.C.; Barker, G.J.; Miller, D.H.; Brierley, C.M.H.; Compston, D.A.S.; Scolding, N.J.; Moseley, I.F.; Plant, G.T.

    2001-01-01

    We describe an MRI technique for quantifying optic nerve atrophy resulting from a single episode of unilateral optic neuritis. We imaged 17 patients, with a median time since onset of optic neuritis of 21 months (range 3-81 months), using a coronal-oblique fat-saturated short-echo fast fluid-attenuated inversion-recovery (sTE fFLAIR) sequence. The mean cross-sectional area of the intraorbital portion of the optic nerves was calculated by a blinded observer from five consecutive 3 mm slices from the orbital apex forwards using a semiautomated contouring technique and compared with data from 16 controls. The mean optic nerve area was 11.2mm 2 in the affected eye of the patients, 12.9mm 2 in the contralateral eye (P = 0.006 compared to the affected eye) and 12.8mm 2 in controls (P = 0.03 compared to the affected eyes). There was a significant negative correlation between disease duration and the size of the affected optic nerve (r = -0.59, P = 0.012). The measurement coefficient of variation was 4.8 %. The sTE fFLAIR sequence enables measurement of optic nerve area with sufficient reproducibility to show optic nerve atrophy following a single episode of unilateral optic neuritis. The correlation of increasing optic nerve atrophy with disease duration would be consistent with ongoing axonal loss in a persistently demyelinated lesion, or Wallerian degeneration following axonal damage during the acute inflammatory phase. (orig.)

  7. The application of strand invasion phenomenon, directed by peptide nucleic acid (PNA) and single-stranded DNA binding protein (SSB) for the recognition of specific sequences of human endogenous retroviral HERV-W family.

    Science.gov (United States)

    Machnik, Grzegorz; Bułdak, Łukasz; Ruczyński, Jarosław; Gąsior, Tomasz; Huzarska, Małgorzata; Belowski, Dariusz; Alenowicz, Magdalena; Mucha, Piotr; Rekowski, Piotr; Okopień, Bogusław

    2017-05-01

    The HERV-W family of human endogenous retroviruses represents a group of numerous sequences that show close similarity in genetic composition. It has been documented that some members of HERV-W-derived expression products are supposed to play significant role in humans' pathology, such as multiple sclerosis or schizophrenia. Other members of the family are necessary to orchestrate physiological processes (eg, ERVWE1 coding syncytin-1 that is engaged in syncytiotrophoblast formation). Therefore, an assay that would allow the recognition of particular form of HERV-W members is highly desirable. A peptide nucleic acid (PNA)-mediated technique for the discrimination between multiple sclerosis-associated retrovirus and ERVWE1 sequence has been developed. The assay uses a PNA probe that, being fully complementary to the ERVWE1 but not to multiple sclerosis-associated retrovirus (MSRV) template, shows high selective potential. Single-stranded DNA binding protein facilitates the PNA-mediated, sequence-specific formation of strand invasion complex and, consequently, local DNA unwinding. The target DNA may be then excluded from further analysis in any downstream process such as single-stranded DNA-specific exonuclease action. Finally, the reaction conditions have been optimized, and several PNA probes that are targeted toward distinct loci along whole HERV-W env sequences have been evaluated. We believe that PNA/single-stranded DNA binding protein-based application has the potential to selectively discriminate particular HERV-W molecules as they are at least suspected to play pathogenic role in a broad range of medical conditions, from psycho-neurologic disorders (multiple sclerosis and schizophrenia) and cancers (breast cancer) to that of an auto-immunologic background (psoriasis and lupus erythematosus). Copyright © 2016 John Wiley & Sons, Ltd.

  8. Magnetic resonance imaging of pelvic entheses - a systematic comparison between short tau inversion recovery (STIR) and T1-weighted, contrast-enhanced, fat-saturated sequences

    International Nuclear Information System (INIS)

    Klang, Eyal; Aharoni, Dvora; Rimon, Uri; Eshed, Iris; Hermann, Kay-Geert; Herman, Amir; Shazar, Nachshon

    2014-01-01

    To assess the contribution of contrast material in detecting and evaluating enthesitis of pelvic entheses by MRI. Sixty-seven hip or pelvic 1.5-T MRIs (30:37 male:female, mean age: 53 years) were retrospectively evaluated for the presence of hamstring and gluteus medius (GM) enthesitis by two readers (a resident and an experienced radiologist). Short tau inversion recovery (STIR) and T1-weighted pre- and post-contrast (T1+Gd) images were evaluated by each reader at two sessions. A consensus reading of two senior radiologists was regarded as the gold standard. Clinical data was retrieved from patients' referral form and medical files. Cohen's kappa was used for intra- and inter-observer agreement calculation. Diagnostic properties were calculated against the gold standard reading. A total of 228 entheses were evaluated. Gold standard analysis diagnosed 83 (36 %) enthesitis lesions. Intra-reader reliability for the experienced reader was significantly (p = 0.0001) higher in the T1+Gd images compared to the STIR images (hamstring: k = 0.84/0.45, GM: k = 0.84/0.47). Sensitivity and specificity increased from 0.74/0.8 to 0.87/0.9 in the STIR images and T1+Gd sequences. Intra-reader reliability for the inexperienced reader was lower (p > 0.05). Evidence showing that contrast material improves the reliability, sensitivity, and specificity of detecting enthesitis supports its use in this setting. (orig.)

  9. 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.

  10. 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.

  11. 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

  12. The isolation and amino acid sequence of an adrenocorticotrophin from the pars distalis and a corticotrophin-like intermediate-lobe peptide from the neurointermediate lobe of the pituitary of the dogfish Squalus acanthias

    Science.gov (United States)

    Lowry, Philip J.; Bennett, Hugh P. J.; McMartin, Colin; Scott, Alexander P.

    1974-01-01

    An adrenocorticotrophic hormone (ACTH) was isolated from extracts of the pars distalis of the pituitary of the dogfish Squalus acanthias by gel filtration and ion-exchange chromatography. It had 15% of the potency of human ACTH in promoting cortico-steroidogenesis in isolated rat adrenal cells. Sequence analysis revealed it to be a nonatria-contapeptide with the following primary structure: Ser-Tyr-Ser-Met-Glu-His-Phe-Arg-Trp-Gly-Lys-Pro-Met-Gly-Arg-Lys-Arg-Arg-Pro-Ile-Lys-Val-Tyr-Pro-Asn-Ser-Phe-Glu-Asp-Glu-Ser-Val-Glu-Asn-Met-Gly-Pro-Glu-Leu. The N-terminal tridecapeptide sequence was identical with the proposed structure of dogfish α-melanocyte-stimulating hormone (α-MSH). On comparison with human ACTH eleven amino acid differences were seen, nine of which are in the 20–39 region of the molecule which is not essential for the steroidogenic activity of ACTH. A peptide identical with the 18–39 portion of this new ACTH was similarly isolated from the neurointermediate lobe of the pituitary where considerable amounts of dogfish α-MSH were found. This supported our view that ACTH as well as having a distinct biological role of its own is also the precursor of α-MSH. PMID:4375977

  13. 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.

  14. Interpreting peptide mass spectra by VEMS

    DEFF Research Database (Denmark)

    Mathiesen, Rune; Lundsgaard, M.; Welinder, Karen G.

    2003-01-01

    the calculated and the experimental mass spectrum of the called peptide. The program package includes four accessory programs. VEMStrans creates protein databases in FASTA format from EST or cDNA sequence files. VEMSdata creates a virtual peptide database from FASTA files. VEMSdist displays the distribution......Most existing Mass Spectra (MS) analysis programs are automatic and provide limited opportunity for editing during the interpretation. Furthermore, they rely entirely on publicly available databases for interpretation. VEMS (Virtual Expert Mass Spectrometrist) is a program for interactive analysis...... of peptide MS/MS spectra imported in text file format. Peaks are annotated, the monoisotopic peaks retained, and the b-and y-ion series identified in an interactive manner. The called peptide sequence is searched against a local protein database for sequence identity and peptide mass. The report compares...

  15. Antimicrobial Peptides: An Introduction.

    Science.gov (United States)

    Haney, Evan F; Mansour, Sarah C; Hancock, Robert E W

    2017-01-01

    The "golden era" of antibiotic discovery has long passed, but the need for new antibiotics has never been greater due to the emerging threat of antibiotic resistance. This urgency to develop new antibiotics has motivated researchers to find new methods to combat pathogenic microorganisms resulting in a surge of research focused around antimicrobial peptides (AMPs; also termed host defense peptides) and their potential as therapeutics. During the past few decades, more than 2000 AMPs have been identified from a diverse range of organisms (animals, fungi, plants, and bacteria). While these AMPs share a number of common features and a limited number of structural motifs; their sequences, activities, and targets differ considerably. In addition to their antimicrobial effects, AMPs can also exhibit immunomodulatory, anti-biofilm, and anticancer activities. These diverse functions have spurred tremendous interest in research aimed at understanding the activity of AMPs, and various protocols have been described to assess different aspects of AMP function including screening and evaluating the activities of natural and synthetic AMPs, measuring interactions with membranes, optimizing peptide function, and scaling up peptide production. Here, we provide a general overview of AMPs and introduce some of the methodologies that have been used to advance AMP research.

  16. Effect of short-term vs. long-term elevation of dietary protein intake on responsiveness of rat thick ascending limbs to peptide hormones.

    Science.gov (United States)

    Goldstein, David L; Plaga, Kimberly

    2002-10-01

    We compared the renal responses of rats on three diet regimens. Rats received either 8% protein food (low-protein, LP) for 10 weeks following weaning, 8% protein for 9 weeks followed by 1 week on 30% protein (short-term high-protein, SHP), or 30% protein for 10 weeks (high-protein, HP). Kidneys from HP rats were enlarged by approximately 50%, or 20% when corrected for body mass. Most of this hypertrophy resulted from enlargement of the inner stripe of the outer medulla, site of the thick ascending limbs (TAL), and TAL from HP rats were larger in diameter. SHP rats had TAL diameters similar to HP rats, but changes in renal mass or height of renal zones did not reach statistical significance. The activity of adenylyl cyclase (AC) in TAL, measured from the accumulation of cAMP in isolated tubules, increased with dose of both arginine vasopressin (AVP) and glucagon in all rats. However, HP rats had significantly higher hormone-induced AC activity than LP or SHP rats, which were not different from each other. Our results suggest that tubule hypertrophy may precede up-regulation of hormone-sensitive AC activity during the progression of renal response to elevated dietary protein.

  17. 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...

  18. 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.

  19. Whole body sodium MRI at 3T using an asymmetric birdcage resonator and short echo time sequence: first images of a male volunteer

    Science.gov (United States)

    Wetterling, Friedrich; Corteville, Dominique M.; Kalayciyan, Raffi; Rennings, Andreas; Konstandin, Simon; Nagel, Armin M.; Stark, Helmut; Schad, Lothar R.

    2012-07-01

    Sodium magnetic resonance imaging (23Na MRI) is a non-invasive technique which allows spatial resolution of the tissue sodium concentration (TSC) in the human body. TSC measurements could potentially serve to monitor early treatment success of chemotherapy on patients who suffer from whole body metastases. Yet, the acquisition of whole body sodium (23Na) images has been hampered so far by the lack of large resonators and the extremely low signal-to-noise ratio (SNR) achieved with existing resonator systems. In this study, a 23Na resonator was constructed for whole body 23Na MRI at 3T comprising of a 16-leg, asymmetrical birdcage structure with 34 cm height, 47.5 cm width and 50 cm length. The resonator was driven in quadrature mode and could be used either as a transceiver resonator or, since active decoupling was included, as a transmit-only resonator in conjunction with a receive-only (RO) surface resonator. The relative B1-field profile was simulated and measured on phantoms, and 3D whole body 23Na MRI data of a healthy male volunteer were acquired in five segments with a nominal isotropic resolution of (6 × 6 × 6) mm3 and a 10 min acquisition time per scan. The measured SNR values in the 23Na-MR images varied from 9 ± 2 in calf muscle, 15 ± 2 in brain tissue, 23 ± 2 in the prostate and up to 42 ± 5 in the vertebral discs. Arms, legs, knees and hands could also be resolved with applied resonator and short time-to-echo (TE) (0.5 ms) radial sequence. Up to fivefold SNR improvement was achieved through combining the birdcage with local RO surface coil. In conclusion, 23Na MRI of the entire human body provides sub-cm spatial resolution, which allows resolution of all major human body parts with a scan time of less than 60 min.

  20. High clinical and morphologic response using 90Y-DOTA-octreotate sequenced with 177Lu-DOTA-octreotate induction peptide receptor chemoradionuclide therapy (PRCRT) for bulky neuroendocrine tumours.

    Science.gov (United States)

    Kong, Grace; Callahan, Jason; Hofman, Michael S; Pattison, David A; Akhurst, Tim; Michael, Michael; Eu, Peter; Hicks, Rodney J

    2017-03-01

    Bulky disease is an adverse prognostic factor for 177 Lu-DOTA-octreotate ( 177 Lu-DOTATATE) peptide receptor radionuclide therapy (PRRT). 90 Y-DOTA-octreotate ( 90 Y-DOTATATE) has theoretical advantages in this setting but may less effectively treat co-existent smaller deposits and have higher toxicity than 177 Lu-DOTATATE. The aim of this study was to assess the efficacy and safety of using these agents sequentially. We reviewed patients (pts) with at least one lesion of a transaxial diameter >4 cm who completed 1-2 cycles of 90 Y-DOTATATE followed by 2-3 cycles of 177 Lu-DOTATATE, with treatment empirically adapted to disease size and burden in individual patients. Data collected included morphological and molecular imaging response, toxicity, and progression-free and overall survival. Twenty-six pts (17 men; aged 27-74 years) received a median cumulative activity of 6.5 GBq 90 Y-DOTATATE, and 21 GBq 177 Lu-DOTATATE. All but one received radiosensitising chemotherapy. Adverse prognostic factors included ENETS grade 2 or 3 in 58 %, and FDG-avid disease in 73 %. Nineteen pts treated for progressive disease had stabilisation (37 %) or regression on CT (42 % partial response, 21 % minor response), with a mean 59 % (8-99 %) reduction in disease burden. All seven pts treated for uncontrolled symptoms reported improvement during PRRT with 4/7 having complete symptom resolution at 3 months. Eight patients had grade 3/4 lymphopaenia, and two patients grade 3/4 thrombocytopaenia without significant hepatic or renal toxicity. Median survival was not reached after a median follow-up of 35 months. Median progression-free survival was 33 months. PRCRT with 90 Y -DOTATATE followed by 177 Lu-DOTATATE in individualised regimens achieved high clinical and morphological response in patients with bulky tumours. Despite lack of a control arm, the efficacy of this treatment approach appears higher than reported results with either agent used alone or other approved

  1. De Novo Assembly of Human Herpes Virus Type 1 (HHV-1) Genome, Mining of Non-Canonical Structures and Detection of Novel Drug-Resistance Mutations Using Short- and Long-Read Next Generation Sequencing Technologies.

    Science.gov (United States)

    Karamitros, Timokratis; Harrison, Ian; Piorkowska, Renata; Katzourakis, Aris; Magiorkinis, Gkikas; Mbisa, Jean Lutamyo

    2016-01-01

    Human herpesvirus type 1 (HHV-1) has a large double-stranded DNA genome of approximately 152 kbp that is structurally complex and GC-rich. This makes the assembly of HHV-1 whole genomes from short-read sequencing data technically challenging. To improve the assembly of HHV-1 genomes we have employed a hybrid genome assembly protocol using data from two sequencing technologies: the short-read Roche 454 and the long-read Oxford Nanopore MinION sequencers. We sequenced 18 HHV-1 cell culture-isolated clinical specimens collected from immunocompromised patients undergoing antiviral therapy. The susceptibility of the samples to several antivirals was determined by plaque reduction assay. Hybrid genome assembly resulted in a decrease in the number of contigs in 6 out of 7 samples and an increase in N(G)50 and N(G)75 of all 7 samples sequenced by both technologies. The approach also enhanced the detection of non-canonical contigs including a rearrangement between the unique (UL) and repeat (T/IRL) sequence regions of one sample that was not detectable by assembly of 454 reads alone. We detected several known and novel resistance-associated mutations in UL23 and UL30 genes. Genome-wide genetic variability ranged from genomes will be useful in determining genetic determinants of drug resistance, virulence, pathogenesis and viral evolution. The numerous, complex repeat regions of the HHV-1 genome currently remain a barrier towards this goal.

  2. 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.

  3. Identification of sixteen peptides reflecting heat and/or storage induced processes by profiling of commercial milk samples.

    Science.gov (United States)

    Ebner, Jennifer; Baum, Florian; Pischetsrieder, Monika

    2016-09-16

    Peptide profiles of different drinking milk samples were examined to study how the peptide fingerprint of milk reflects processing conditions. The combination of a simple and fast method for peptide extraction using stage tips and MALDI-TOF-MS enabled the fast and easy generation and relative quantification of peptide fingerprints for high-temperature short-time (HTST), extended shelf life (ESL) and ultra-high temperature (UHT) milk of the same dairies. The relative quantity of 16 peptides changed as a function of increasing heat load. Additional heating experiments showed that among those, the intensity of peptide β-casein 196-209 (m/z 1460.9Da) was most heavily influenced by heat treatment indicating a putative marker peptide for milk processing conditions. Storage experiments with HTST- and UHT milk revealed that the differences between different types of milk samples were not only caused by the heating process. Relevant was also the proteolytic activity of enzymes during storage, which were differently influenced by the heat treatment. These results indicate that the peptide profile may be suitable to monitor processing as well as storage conditions of milk. In the present study, peptide profiling of different types of milk was carried out by MALDI-TOF-MS after stage-tip extraction and relative quantification using an internal reference peptide. Although MALDI-TOF-MS covers only part of the peptidome, the method is easy and quick and is, therefore, suited for routine analysis to address several aspects of food authenticity. Using this method, 16 native peptides were detected in milk that could be modulated by different industrial processes. Subsequent heating and storage experiments with pasteurized and UHT milk confirmed that these peptides are indeed related to the production or storage conditions of the respective products. Furthermore, the heating experiments revealed one peptide, namely the β-casein-derived sequence β-casein 196-209, which underwent

  4. Synthesis and Pharmacology of α/β(3)-Peptides Based on the Melanocortin Agonist Ac-His-dPhe-Arg-Trp-NH2 Sequence.

    Science.gov (United States)

    Singh, Anamika; Tala, Srinivasa R; Flores, Viktor; Freeman, Katie; Haskell-Luevano, Carrie

    2015-05-14

    The melanocortin-3 and -4 receptors are expressed in the brain and play key roles in regulating feeding behavior, metabolism, and energy homeostasis. In the present study, incorporation of β(3)-amino acids into a melanocortin tetrapeptide template was investigated. Four linear α/β(3)-hybrid tetrapeptides were designed with the modifications at the Phe, Arg, and Trp residues in the agonist sequence Ac-His-dPhe-Arg-Trp-NH2. The most potent mouse melanocortin-4 receptor (mMC4R) agonist, Ac-His-dPhe-Arg-β(3)hTrp-NH2 (8) showed 35-fold selectivity versus the mMC3R. The study presented here has identified a new template with heterogeneous backbone for designing potent and selective melanocortin receptor ligands.

  5. Synthesis and Pharmacology of α/β3-Peptides Based on the Melanocortin Agonist Ac-His-dPhe-Arg-Trp-NH2 Sequence

    Science.gov (United States)

    2015-01-01

    The melanocortin-3 and -4 receptors are expressed in the brain and play key roles in regulating feeding behavior, metabolism, and energy homeostasis. In the present study, incorporation of β3-amino acids into a melanocortin tetrapeptide template was investigated. Four linear α/β3-hybrid tetrapeptides were designed with the modifications at the Phe, Arg, and Trp residues in the agonist sequence Ac-His-dPhe-Arg-Trp-NH2. The most potent mouse melanocortin-4 receptor (mMC4R) agonist, Ac-His-dPhe-Arg-β3hTrp-NH2 (8) showed 35-fold selectivity versus the mMC3R. The study presented here has identified a new template with heterogeneous backbone for designing potent and selective melanocortin receptor ligands. PMID:26005535

  6. Protection against the Metabolic Syndrome by Guar Gum-Derived Short-Chain Fatty Acids Depends on Peroxisome Proliferator-Activated Receptor γ and Glucagon-Like Peptide-1.

    Science.gov (United States)

    den Besten, Gijs; Gerding, Albert; van Dijk, Theo H; Ciapaite, Jolita; Bleeker, Aycha; van Eunen, Karen; Havinga, Rick; Groen, Albert K; Reijngoud, Dirk-Jan; Bakker, Barbara M

    2015-01-01

    The dietary fiber guar gum has beneficial effects on obesity, hyperglycemia and hypercholesterolemia in both humans and rodents. The major products of colonic fermentation of dietary fiber, the short-chain fatty acids (SCFAs), have been suggested to play an important role. Recently, we showed that SCFAs protect against the metabolic syndrome via a signaling cascade that involves peroxisome proliferator-activated receptor (PPAR) γ repression and AMP-activated protein kinase (AMPK) activation. In this study we investigated the molecular mechanism via which the dietary fiber guar gum protects against the metabolic syndrome. C57Bl/6J mice were fed a high-fat diet supplemented with 0% or 10% of the fiber guar gum for 12 weeks and effects on lipid and glucose metabolism were studied. We demonstrate that, like SCFAs, also guar gum protects against high-fat diet-induced metabolic abnormalities by PPARγ repression, subsequently increasing mitochondrial uncoupling protein 2 expression and AMP/ATP ratio, leading to the activation of AMPK and culminating in enhanced oxidative metabolism in both liver and adipose tissue. Moreover, guar gum markedly increased peripheral glucose clearance, possibly mediated by the SCFA-induced colonic hormone glucagon-like peptide-1. Overall, this study provides novel molecular insights into the beneficial effects of guar gum on the metabolic syndrome and strengthens the potential role of guar gum as a dietary-fiber intervention.

  7. Protection against the Metabolic Syndrome by Guar Gum-Derived Short-Chain Fatty Acids Depends on Peroxisome Proliferator-Activated Receptor γ and Glucagon-Like Peptide-1.

    Directory of Open Access Journals (Sweden)

    Gijs den Besten

    Full Text Available The dietary fiber guar gum has beneficial effects on obesity, hyperglycemia and hypercholesterolemia in both humans and rodents. The major products of colonic fermentation of dietary fiber, the short-chain fatty acids (SCFAs, have been suggested to play an important role. Recently, we showed that SCFAs protect against the metabolic syndrome via a signaling cascade that involves peroxisome proliferator-activated receptor (PPAR γ repression and AMP-activated protein kinase (AMPK activation. In this study we investigated the molecular mechanism via which the dietary fiber guar gum protects against the metabolic syndrome. C57Bl/6J mice were fed a high-fat diet supplemented with 0% or 10% of the fiber guar gum for 12 weeks and effects on lipid and glucose metabolism were studied. We demonstrate that, like SCFAs, also guar gum protects against high-fat diet-induced metabolic abnormalities by PPARγ repression, subsequently increasing mitochondrial uncoupling protein 2 expression and AMP/ATP ratio, leading to the activation of AMPK and culminating in enhanced oxidative metabolism in both liver and adipose tissue. Moreover, guar gum markedly increased peripheral glucose clearance, possibly mediated by the SCFA-induced colonic hormone glucagon-like peptide-1. Overall, this study provides novel molecular insights into the beneficial effects of guar gum on the metabolic syndrome and strengthens the potential role of guar gum as a dietary-fiber intervention.

  8. Peptides in headlock – a novel high-affinity and versatile peptide-binding nanobody for proteomics and microscopy

    Science.gov (United States)

    Braun, Michael B.; Traenkle, Bjoern; Koch, Philipp A.; Emele, Felix; Weiss, Frederik; Poetz, Oliver; Stehle, Thilo; Rothbauer, Ulrich

    2016-01-01

    Nanobodies are highly valuable tools for numerous bioanalytical and biotechnical applications. Here, we report the characterization of a nanobody that binds a short peptide epitope with extraordinary affinity. Structural analysis reveals an unusual binding mode where the extended peptide becomes part of a β-sheet structure in the nanobody. This interaction relies on sequence-independent backbone interactions augmented by a small number of specificity-determining side chain contacts. Once bound, the peptide is fastened by two nanobody side chains that clamp it in a headlock fashion. Exploiting this unusual binding mode, we generated a novel nanobody-derived capture and detection system. Matrix-coupled nanobody enables the fast and efficient isolation of epitope-tagged proteins from prokaryotic and eukaryotic expression systems. Additionally, the fluorescently labeled nanobody visualizes subcellular structures in different cellular compartments. The high-affinity-binding and modifiable peptide tag of this system renders it a versatile and robust tool to combine biochemical analysis with microscopic studies. PMID:26791954

  9. Peptides in headlock--a novel high-affinity and versatile peptide-binding nanobody for proteomics and microscopy.

    Science.gov (United States)

    Braun, Michael B; Traenkle, Bjoern; Koch, Philipp A; Emele, Felix; Weiss, Frederik; Poetz, Oliver; Stehle, Thilo; Rothbauer, Ulrich

    2016-01-21

    Nanobodies are highly valuable tools for numerous bioanalytical and biotechnical applications. Here, we report the characterization of a nanobody that binds a short peptide epitope with extraordinary affinity. Structural analysis reveals an unusual binding mode where the extended peptide becomes part of a β-sheet structure in the nanobody. This interaction relies on sequence-independent backbone interactions augmented by a small number of specificity-determining side chain contacts. Once bound, the peptide is fastened by two nanobody side chains that clamp it in a headlock fashion. Exploiting this unusual binding mode, we generated a novel nanobody-derived capture and detection system. Matrix-coupled nanobody enables the fast and efficient isolation of epitope-tagged proteins from prokaryotic and eukaryotic expression systems. Additionally, the fluorescently labeled nanobody visualizes subcellular structures in different cellular compartments. The high-affinity-binding and modifiable peptide tag of this system renders it a versatile and robust tool to combine biochemical analysis with microscopic studies.

  10. Cardioprotective peptides from marine sources.

    Science.gov (United States)

    Harnedy, Padraigín A; FitzGerald, Richard J

    2013-05-01

    Elevated blood pressure or hypertension is one of the fastest growing health problems worldwide. Although the etiology of essential hypertension has a genetic component, dietary factors play an important role. With the high costs and adverse side-effects associated with synthetic antihypertensive drugs and the awareness of the link between diet and health there has been increased focus on identification of food components that may contribute to cardiovascular health. In recent years special interest has been paid to the cardioprotective activity of peptides derived from food proteins including marine proteins. These peptides are latent within the sequence of the parent protein and only become active when released by proteolytic digestion during gastrointestinal digestion or through food processing. Current data on antihypertensive activity of marine-derived protein hydrolysates/peptides in animal and human studies is reviewed herein. Furthermore, products containing protein hydrolysates/peptides from marine origin with antihypertensive effects are discussed.

  11. Sequence of the radioactive tryptic peptide obtained after inactivating the F1-ATPase of the thermophilic bacterium PS3 with 5'-p-fluorosulfonylbenzoyl[3H]adenosine at 65 degrees C

    International Nuclear Information System (INIS)

    Bullough, D.A.; Yoshida, M.; Allison, W.S.

    1986-01-01

    Following a lag of about 30 min, the F1-ATPase from the thermophilic bacterium, PS3 (TF1), was inactivated slowly by 0.8 mM 5'-p-fluorosulfonylbenzoyladenosine (FSBA) at 23 degrees C and pH 7.0. When the enzyme was treated with 0.2 mM FSBA at pH 7.0 and 23 degrees C for 15 min and gel-filtered, no enzyme activity was lost. However, the lag in inactivation was abolished when the enzyme was subsequently incubated with 2.0 mM FSBA at 23 degrees C in the pH range from 6.8 to 10.0. The pH-inactivation profile obtained under these conditions revealed a pK alpha of about 9.3 which was associated with the inactivation. When pretreated TF1 was inactivated at 23 degrees C with [3H]FSBA by about 90%, greater than 20 mol of [3H]SBA was incorporated per mole of enzyme. TF1 was inactivated rapidly by 0.8 mM FSBA at pH 6.4 and 65 degrees C, and no lag was observed. Following inactivation of TF1 with 0.8 mM [3H]FSBA at 65 degrees C and pH 6.4, about 10 mol of [3H]SBA was incorporated per mole of enzyme. When a tryptic digest of the labeled enzyme was fractionated by reversed-phase high-performance liquid chromatography, a single major radioactive peptide was isolated. When subjected to automatic Edman degradation, this peptide was shown to have the amino acid sequence: A-L-A-P-E-I-V-G-E-E-H-X-Q-V-A-R, where X indicates that a phenylthiohydantoin derivative was not detected in cycle 12. However, from the DNA sequence of the gene encoding the subunit of TF1 (Y. Kagawa, M. Ishizuka, T. Saishu, and S. Nakao (1985)), this position has been shown to be occupied by tyrosine. This tyrosine is homologous with beta-Tyr-368 of the bovine mitochondrial F1-ATPase (MF1) the modification of which is responsible for the inactivation MF1 by FSBA

  12. The preparation and characterization of peptide's lung cancer imaging agent

    International Nuclear Information System (INIS)

    Liu Jianfeng; Chu Liping; Wang Yan; Wang Yueying; Liu Jinjian; Wu Hongying

    2010-01-01

    Objective: To screen in vivo lung cancer specific binding seven peptides by T7 phage display peptide library, so as to prepare peptide's lung cancer early diagnostic agent. Methods: Use phage display in vivo technology, the 7-peptide phage that binding the lung cancer specifically was obtained, then the DNA sequence was measured and the seven peptide was synthesized. After labeled by 125 I, the seven peptide was injected into mice via vein and the distribution was observed. Results: One peptide was obtained by four rounds screening, and the peptide can bind lung cancer tissue specifically. Two hours after injection get the best imaging of lung cancer, metabolism of peptide in mice is fast, the distribution in vivo is decrease six hours and almost disappear 20 hours after injection. Conclusion: The peptide can image and diagnose lung cancer better. (authors)

  13. Minimalistic peptide supramolecular co-assembly: expanding the conformational space for nanotechnology.

    Science.gov (United States)

    Makam, Pandeeswar; Gazit, Ehud

    2018-05-21

    Molecular self-assembly is a ubiquitous process in nature and central to bottom-up nanotechnology. In particular, the organization of peptide building blocks into ordered supramolecular structures has gained much interest due to the unique properties of the products, including biocompatibility, chemical and structural diversity, robustness and ease of large-scale synthesis. In addition, peptides, as short as dipeptides, contain all the molecular information needed to spontaneously form well-ordered structures at both the nano- and the micro-scale. Therefore, peptide supramolecular assembly has been effectively utilized to produce novel materials with tailored properties for various applications in the fields of material science, engineering, medicine, and biology. To further expand the conformational space of peptide assemblies in terms of structural and functional complexity, multicomponent (two or more) peptide supramolecular co-assembly has recently evolved as a promising extended approach, similar to the structural diversity of natural sequence-defined biopolymers (proteins) as well as of synthetic covalent co-polymers. The use of this methodology was recently demonstrated in various applications, such as nanostructure physical dimension control, the creation of non-canonical complex topologies, mechanical strength modulation, the design of light harvesting soft materials, fabrication of electrically conducting devices, induced fluorescence, enzymatic catalysis and tissue engineering. In light of these significant advancements in the field of peptide supramolecular co-assembly in the last few years, in this tutorial review, we provide an updated overview and future prospects of this emerging subject.

  14. Facilitating protein solubility by use of peptide extensions

    Science.gov (United States)

    Freimuth, Paul I; Zhang, Yian-Biao; Howitt, Jason

    2013-09-17

    Expression vectors for expression of a protein or polypeptide of interest as a fusion product composed of the protein or polypeptide of interest fused at one terminus to a solubility enhancing peptide extension are provided. Sequences encoding the peptide extensions are provided. The invention further comprises antibodies which bind specifically to one or more of the solubility enhancing peptide extensions.

  15. 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

  16. A Tetrameric Peptide Derived from Bovine Lactoferricin Exhibits Specific Cytotoxic Effects against Oral Squamous-Cell Carcinoma Cell Lines

    Directory of Open Access Journals (Sweden)

    Víctor A. Solarte

    2015-01-01

    Full Text Available Several short linear peptides derived from cyclic bovine lactoferricin were synthesized and tested for their cytotoxic effect against the oral cavity squamous-cell carcinoma (OSCC cell lines CAL27 and SCC15. As a control, an immortalized and nontumorigenic cell line, Het-1A, was used. Linear peptides based on the RRWQWR core sequence showed a moderate cytotoxic effect and specificity towards tumorigenic cells. A tetrameric peptide, LfcinB(20–254, containing the RRWQWR motif, exhibited greater cytotoxic activity (>90% in both OSCC cell lines compared to the linear lactoferricin peptide or the lactoferrin protein. Additionally, this tetrameric peptide showed the highest specificity towards tumorigenic cells among the tested peptides. Interestingly, this effect was very fast, with cell shrinkage, severe damage to cell membrane permeability, and lysis within one hour of treatment. Our results are consistent with a necrotic effect rather than an apoptotic one and suggest that this tetrameric peptide could be considered as a new candidate for the therapeutic treatment of OSCC.

  17. A Tetrameric Peptide Derived from Bovine Lactoferricin Exhibits Specific Cytotoxic Effects against Oral Squamous-Cell Carcinoma Cell Lines.

    Science.gov (United States)

    Solarte, Víctor A; Rosas, Jaiver E; Rivera, Zuly J; Arango-Rodríguez, Martha L; García, Javier E; Vernot, Jean-Paul

    2015-01-01

    Several short linear peptides derived from cyclic bovine lactoferricin were synthesized and tested for their cytotoxic effect against the oral cavity squamous-cell carcinoma (OSCC) cell lines CAL27 and SCC15. As a control, an immortalized and nontumorigenic cell line, Het-1A, was used. Linear peptides based on the RRWQWR core sequence showed a moderate cytotoxic effect and specificity towards tumorigenic cells. A tetrameric peptide, LfcinB(20-25)4, containing the RRWQWR motif, exhibited greater cytotoxic activity (>90%) in both OSCC cell lines compared to the linear lactoferricin peptide or the lactoferrin protein. Additionally, this tetrameric peptide showed the highest specificity towards tumorigenic cells among the tested peptides. Interestingly, this effect was very fast, with cell shrinkage, severe damage to cell membrane permeability, and lysis within one hour of treatment. Our results are consistent with a necrotic effect rather than an apoptotic one and suggest that this tetrameric peptide could be considered as a new candidate for the therapeutic treatment of OSCC.

  18. 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

  19. Development of short and highly potent self-assembling elastin-derived pentapeptide repeats containing aromatic amino acid residues.

    Science.gov (United States)

    Taniguchi, Suguru; Watanabe, Noriko; Nose, Takeru; Maeda, Iori

    2016-01-01

    Tropoelastin is the primary component of elastin, which forms the elastic fibers that make up connective tissues. The hydrophobic domains of tropoelastin are thought to mediate the self-assembly of elastin into fibers, and the temperature-mediated self-assembly (coacervation) of one such repetitive peptide sequence (VPGVG) has been utilized in various bio-applications. To elucidate a mechanism for coacervation activity enhancement and to develop more potent coacervatable elastin-derived peptides, we synthesized two series of peptide analogs containing an aromatic amino acid, Trp or Tyr, in addition to Phe-containing analogs and tested their functional characteristics. Thus, position 1 of the hydrophobic pentapeptide repeat of elastin (X(1)P(2)G(3)V(4)G(5)) was substituted by Trp or Tyr. Eventually, we acquired a novel, short Trp-containing elastin-derived peptide analog (WPGVG)3 with potent coacervation ability. From the results obtained during this process, we determined the importance of aromaticity and hydrophobicity for the coacervation potency of elastin-derived peptide analogs. Generally, however, the production of long-chain synthetic polypeptides in quantities sufficient for commercial use remain cost-prohibitive. Therefore, the identification of (WPGVG)3, which is a 15-mer short peptide consisting simply of five natural amino acids and shows temperature-dependent self-assembly activity, might serve as a foundation for the development of various kinds of biomaterials. Copyright © 2015 European Peptide Society and John Wiley & Sons, Ltd.

  20. A cardioactive peptide from the southern armyworm, Spodoptera eridania.

    Science.gov (United States)

    Furuya, K; Hackett, M; Cirelli, M A; Schegg, K M; Wang, H; Shabanowitz, J; Hunt, D F; Schooley, D A

    1999-01-01

    A cardioactive peptide was isolated from extracts of whole heads of the southern armyworm, Spodoptera eridania. This peptide has the sequence ENFAVGCTPGYQRTADGRCKPTF (Mr = 2516.8), determined from both Edman sequencing and tandem mass spectrometry in combination with off-line micropreparative capillary liquid chromatography. This peptide, termed Spoer-CAP23, has excitatory effects on a semi-isolated heart from larval Manduca sexta, causing an inotropic effect at low concentrations of peptide and chronotropic and inotropic effects at high doses. The threshold concentration for stimulatory effects of the synthetic peptide on the semi-isolated heart was about 1 nM, suggesting a physiological role as a neuropeptide.

  1. Short Communication Phylogenetic Characterization of HIV Type 1 CRF01_AE V3 Envelope Sequences in Pregnant Women in Northern Vietnam

    Science.gov (United States)

    Caridha, Rozina; Ha, Tran Thi Thanh; Gaseitsiwe, Simani; Hung, Pham Viet; Anh, Nguyen Mai; Bao, Nguyen Huy; Khang, Dinh Duy; Hien, Nguyen Tran; Cam, Phung Dac; Chiodi, Francesca

    2012-01-01

    Abstract Characterization of HIV-1 strains is important for surveillance of the HIV-1 epidemic. In Vietnam HIV-1-infected pregnant women often fail to receive the care they are entitled to. Here, we analyzed phylogenetically HIV-1 env sequences from 37 HIV-1-infected pregnant women from Ha Noi (n=22) and Hai Phong (n=15), where they delivered in 2005–2007. All carried CRF01_AE in the gp120 V3 region. In 21 women CRF01_AE was also found in the reverse transcriptase gene. We compared their env gp120 V3 sequences phylogenetically in a maximum likelihood tree to those of 198 other CRF01_AE sequences in Vietnam and 229 from neighboring countries, predominantly Thailand, from the HIV-1 database. Altogether 464 sequences were analyzed. All but one of the maternal sequences colocalized with sequences from northern Vietnam. The maternal sequences had evolved the least when compared to sequences collected in Ha Noi in 2002, as shown by analysis of synonymous and nonsynonymous changes, than to other Vietnamese sequences collected earlier and/or elsewhere. Since the HIV-1 epidemic in women in Vietnam may still be underestimated, characterization of HIV-1 in pregnant women is important to observe how HIV-1 has evolved and follow its molecular epidemiology. PMID:21936713

  2. Distribution of Campylobacter jejuni isolates from turkey farms and different stages at slaughter using pulsed-field gel electrophoresis and flaA-short variable region sequencing.

    Science.gov (United States)

    Perko-Mäkelä, P; Alter, T; Isohanni, P; Zimmermann, S; Lyhs, U

    2011-09-01

    The aim of this study was to assess the diversity of thermotolerant Campylobacter spp. isolated from turkey flocks at six rearing farms 1-2 weeks prior to slaughter (360 faecal swab samples) and from 11 different stages at the slaughterhouse (636 caecal, environmental, neck skin and meat samples). A total of 121 Campylobacter isolates were identified to species level using a multiplex PCR assay and were typed by pulsed-field gel electrophoresis (PFGE) and flaA-short variable region (SVR) sequencing. All Campylobacter isolates were identified as Campylobacter jejuni. PFGE analysis with KpnI restriction enzyme resulted in 11 PFGE types (I-XI) and flaA SVR typing yielded in nine flaA-SVR alleles. The Campylobacter-positive turkey flocks A, C and E were colonized by a limited number of Campylobacter clones at the farm and slaughter. The present study confirms the traceability of flock-specific strains (PFGE types I, V and IX; flaA types 21, 36 and 161) from the farm along the entire processing line to meat cuts. It seems that stress factors such as high temperature of the defeathering water (54-56 °C), drying of the carcass skin during air chilling (24 h at 2 °C), and oxygen in the air could not eliminate Campylobacter completely. Campylobacter-negative flocks became contaminated during processing by the same subtypes of Campylobacter introduced into the slaughter house by preceeding positive flocks even if they were slaughtered on subsequent days. Proper and efficient cleaning and disinfection of slaughter and processing premises are needed to avoid cross-contamination, especially in countries with a low prevalence of Campylobacter spp. The majority of flaA SVR alleles displayed a distinct association with a specific PFGE type. However, a linear relationship for all strains among both typing methods could not be established. To specify genetic relatedness of strains, a combination of different genotyping methods, is needed. © 2011 Blackwell Verlag GmbH.

  3. A synthetic peptide shows retro- and anterograde neuronal transport before disrupting the chemosensation of plant-pathogenic nematodes.

    Directory of Open Access Journals (Sweden)

    Dong Wang

    2011-03-01

    Full Text Available Cyst nematodes are a group of plant pathogens each with a defined host range that cause major losses to crops including potato, soybean and sugar beet. The infective mobile stage hatches from dormant eggs and moves a short distance through the soil to plant roots, which it then invades. A novel strategy for control has recently been proposed in which the plant is able to secrete a peptide which disorientates the infective stage and prevents invasion of the pathogen. This study provides indirect evidence to support the mechanism by which one such peptide disrupts chemosensory function in nematodes. The peptide is a disulphide-constrained 7-mer with the amino acid sequence CTTMHPRLC that binds to nicotinic acetylcholine receptors. A fluorescently tagged version of this peptide with both epifluorescent and confocal microscopy was used to demonstrate that retrograde transport occurs from an aqueous environment along bare-ending primary cilia of chemoreceptive sensilla. The peptide is transported to the cell bodies of these neurons and on to a limited number of other neurons to which they connect. It appears to be localised in both neuronal processes and organelles adjacent to nuclei of some neurons suggesting it could be transported through the Golgi apparatus. The peptide takes 2.5 h to reach the neuronal cell bodies. Comparative studies established that similar but less abundant uptake occurs for Caenorhabditis elegans along its well studied dye-filling chemoreceptive neurons. Incubation in peptide solution or root-exudate from transgenic plants that secrete the peptide disrupted normal orientation of infective cyst nematodes to host root diffusate. The peptide probably undergoes transport along the dye-filling non-cholinergic chemoreceptive neurons to their synapses where it is taken up by the interneurons to which they connect. Coordinated responses to chemoreception are disrupted when the sub-set of cholinergic interneurons secrete the peptide

  4. A synthetic peptide shows retro- and anterograde neuronal transport before disrupting the chemosensation of plant-pathogenic nematodes.

    Science.gov (United States)

    Wang, Dong; Jones, Laura M; Urwin, Peter E; Atkinson, Howard J

    2011-03-07

    Cyst nematodes are a group of plant pathogens each with a defined host range that cause major losses to crops including potato, soybean and sugar beet. The infective mobile stage hatches from dormant eggs and moves a short distance through the soil to plant roots, which it then invades. A novel strategy for control has recently been proposed in which the plant is able to secrete a peptide which disorientates the infective stage and prevents invasion of the pathogen. This study provides indirect evidence to support the mechanism by which one such peptide disrupts chemosensory function in nematodes. The peptide is a disulphide-constrained 7-mer with the amino acid sequence CTTMHPRLC that binds to nicotinic acetylcholine receptors. A fluorescently tagged version of this peptide with both epifluorescent and confocal microscopy was used to demonstrate that retrograde transport occurs from an aqueous environment along bare-ending primary cilia of chemoreceptive sensilla. The peptide is transported to the cell bodies of these neurons and on to a limited number of other neurons to which they connect. It appears to be localised in both neuronal processes and organelles adjacent to nuclei of some neurons suggesting it could be transported through the Golgi apparatus. The peptide takes 2.5 h to reach the neuronal cell bodies. Comparative studies established that similar but less abundant uptake occurs for Caenorhabditis elegans along its well studied dye-filling chemoreceptive neurons. Incubation in peptide solution or root-exudate from transgenic plants that secrete the peptide disrupted normal orientation of infective cyst nematodes to host root diffusate. The peptide probably undergoes transport along the dye-filling non-cholinergic chemoreceptive neurons to their synapses where it is taken up by the interneurons to which they connect. Coordinated responses to chemoreception are disrupted when the sub-set of cholinergic interneurons secrete the peptide at synapses that

  5. Post-staining electroblotting for efficient and reliable peptide blotting.

    Science.gov (United States)

    Lee, Der-Yen; Chang, Geen-Dong

    2015-01-01

    Post-staining electroblotting has been previously described to transfer Coomassie blue-stained proteins from polyacrylamide gel onto polyvinylidene difluoride (PVDF) membranes. Actually, stained peptides can also be efficiently and reliably transferred. Because of selective staining procedures for peptides and increased retention of stained peptides on the membrane, even peptides with molecular masses less than 2 kDa such as bacitracin and granuliberin R are transferred with satisfactory results. For comparison, post-staining electroblotting is about 16-fold more sensitive than the conventional electroblotting for visualization of insulin on the membrane. Therefore, the peptide blots become practicable and more accessible to further applications, e.g., blot overlay detection or immunoblotting analysis. In addition, the efficiency of peptide transfer is favorable for N-terminal sequence analysis. With this method, peptide blotting can be normalized for further analysis such as blot overlay assay, immunoblotting, and N-terminal sequencing for identification of peptide in crude or partially purified samples.

  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. Predicting short-term mortality in advanced decompensated heart failure - role of the updated acute decompensated heart failure/N-terminal pro-B-type natriuretic Peptide risk score.

    Science.gov (United States)

    Scrutinio, Domenico; Ammirati, Enrico; Passantino, Andrea; Guida, Pietro; D'Angelo, Luciana; Oliva, Fabrizio; Ciccone, Marco Matteo; Iacoviello, Massimo; Dentamaro, Ilaria; Santoro, Daniela; Lagioia, Rocco; Sarzi Braga, Simona; Guzzetti, Daniela; Frigerio, Maria

    2015-01-01

    The first few months after admission are the most vulnerable period in patients with acute decompensated heart failure (ADHF). We assessed the association of the updated ADHF/N-terminal pro-B-type natriuretic peptide (NT-proBNP) risk score with 90-day and in-hospital mortality in 701 patients admitted with advanced ADHF, defined as severe symptoms of worsening HF, severely depressed left ventricular ejection fraction, and the need for i.v. diuretic and/or inotropic drugs. A total of 15.7% of the patients died within 90 days of admission and 5.2% underwent ventricular assist device (VAD) implantation or urgent heart transplantation (UHT). The C-statistic of the ADHF/NT-proBNP risk score for 90-day mortality was 0.810 (95% CI: 0.769-0.852). Predicted and observed mortality rates were in close agreement. When the composite outcome of death/VAD/UHT at 90 days was considered, the C-statistic decreased to 0.741. During hospitalization, 7.6% of the patients died. The C-statistic for in-hospital mortality was 0.815 (95% CI: 0.761-0.868) and Hosmer-Lemeshow χ(2)=3.71 (P=0.716). The updated ADHF/NT-proBNP risk score outperformed the Acute Decompensated Heart Failure National Registry, the Organized Program to Initiate Lifesaving Treatment in Patients Hospitalized for Heart Failure, and the American Heart Association Get with the Guidelines Program predictive models. Updated ADHF/NT-proBNP risk score is a valuable tool for predicting short-term mortality in severe ADHF, outperforming existing inpatient predictive models.

  8. [New strategy for RNA vectorization in mammalian cells. Use of a peptide vector].

    Science.gov (United States)

    Vidal, P; Morris, M C; Chaloin, L; Heitz, F; Divita, G

    1997-04-01

    A major barrier for gene delivery is the low permeability of nucleic acids to cellular membranes. The development of antisenses and gene therapy has focused mainly on improving methods of oligonucleotide or gene delivery to the cell. In this report we described a new strategy for RNA cell delivery, based on a short single peptide. This peptide vector is derived from both the fusion domain of the gp41 protein of HIV and the nuclear localization sequence of the SV40 large T antigen. This peptide vector localizes rapidly to the cytoplasm then to the nucleus of human fibroblasts (HS-68) within a few minutes and exhibits a high affinity for a single-stranded mRNA encoding the p66 subunit of the HIV-1 reverse transcriptase (in a 100 nM range). The peptide/RNA complex formation involves mainly electrostatic interactions between the basic residues of the peptide and the charges on the phosphate group of the RNA. In the presence of the peptide-vector fluorescently-labelled mRNA is delivered into the cytoplasm of mammalian cells (HS68 human fibroblasts) in less than 1 h with a relatively high efficiency (80%). This new concept based on a peptide-derived vector offers several advantages compared to other compounds commonly used in gene delivery. This vector is highly soluble and exhibits no cytotoxicity at the concentrations used for optimal gene delivery. This result clearly supports the fact that this peptide vector is a powerful tool and that it can be used widely, as much for laboratory research as for new applications and development in gene and/or antisense therapy.

  9. Chimeric opioid peptides: Tools for identifying opioid receptor types

    International Nuclear Information System (INIS)

    Xie, G.; Miyajima, A.; Yokota, T.; Arai, K.; Goldstein, A.

    1990-01-01

    The authors synthesized several chimeric [125J-labelled] peptides in which the N-terminal nine residues of dynorphin-32, a peptide selective for the κ opioid receptor, were replaced by opioid peptides selective for other opioid receptor types. Each chimeric peptide retained the high affinity and type selectivity characteristic of its N-terminal sequence. The common C-terminal two-thirds of the chimeric peptides served as an epitope recognized by the same monoclonal antibody. When bound to receptors on a cell surface or membrane preparation, these peptides could still bind specifically to the monoclonal antibody. These chimeric peptides should be useful for isolating μ, δ, and κ opioid receptors and for identifying opioid receptors on transfected cells in expression cloning procedures. The general approach using chimeric peptides should be applicable to other peptide receptors

  10. THE USE OF DEDICATED PEPTIDE LIBRARIES PERMITS THE DISCOVERY OF HIGH-AFFINITY BINDING PEPTIDES

    NARCIS (Netherlands)

    DEKOSTER, HS; AMONS, R; BENCKHUIJSEN, WE; FEIJLBRIEF, M; SCHELLEKENS, GA; DRIJFHOUT, JW

    1995-01-01

    The motif for peptide binding to monoclonal antibody mAb A16, which is known to be directed against glycoprotein D of Herpes simplex virus type 1, was determined using two dedicated peptide libraries. As a starting point for this study we used an A-16 binding lead sequence, which had previously been

  11. Escherichia coli Peptide Binding Protein OppA Has a Preference for Positively Charged Peptides

    NARCIS (Netherlands)

    Klepsch, M. M.; Kovermann, M.; Löw, C.; Balbach, J.; Permentier, H. P.; Fusetti, F.; de Gier, J. W.; Gier, Jan-Willem de; Slotboom, D. J.; Berntsson, R. P. -A.

    2011-01-01

    The Escherichia coli peptide binding protein OppA is an essential component of the oligopeptide transporter Opp. Based on studies on its orthologue from Salmonella typhimurium, it has been proposed that OppA binds peptides between two and five amino acids long, with no apparent sequence selectivity.

  12. Generating quantitative models describing the sequence specificity of biological processes with the stabilized matrix method

    Directory of Open Access Journals (Sweden)

    Sette Alessandro

    2005-05-01

    Full Text Available Abstract Background Many processes in molecular biology involve the recognition of short sequences of nucleic-or amino acids, such as the binding of immunogenic peptides to major histocompatibility complex (MHC molecules. From experimental data, a model of the sequence specificity of these processes can be constructed, such as a sequence motif, a scoring matrix or an artificial neural network. The purpose of these models is two-fold. First, they can provide a summary of experimental results, allowing for a deeper understanding of the mechanisms involved in sequence recognition. Second, such models can be used to predict the experimental outcome for yet untested sequences. In the past we reported the development of a method to generate such models called the Stabilized Matrix Method (SMM. This method has been successfully applied to predicting peptide binding to MHC molecules, peptide transport by the transporter associated with antigen presentation (TAP and proteasomal cleavage of protein sequences. Results Herein we report the implementation of the SMM algorithm as a publicly available software package. Specific features determining the type of problems the method is most appropriate for are discussed. Advantageous features of the package are: (1 the output generated is easy to interpret, (2 input and output are both quantitative, (3 specific computational strategies to handle experimental noise are built in, (4 the algorithm is designed to effectively handle bounded experimental data, (5 experimental data from randomized peptide libraries and conventional peptides can easily be combined, and (6 it is possible to incorporate pair interactions between positions of a sequence. Conclusion Making the SMM method publicly available enables bioinformaticians and experimental biologists to easily access it, to compare its performance to other prediction methods, and to extend it to other applications.

  13. Localization of an O-glycosylated site in the recombinant barley alpha-amylase 1 produced in yeast and correction of the amino acid sequence using matrix-assisted laser desorption/ionization mass spectrometry of peptide mixtures

    DEFF Research Database (Denmark)

    Andersen, Jens S.; Søgaard, M; Svensson, B

    1994-01-01

    Matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) of peptide mixtures was used to characterize recombinant barley alpha-amylase 1, produced in yeast. Three peptide mixtures were generated by cleavage with CNBr, digestion with endoproteinase Lys-C and Asp-N, respectively...

  14. Characterization of foot-and-mouth disease virus's viral peptides with LC-ESI-MS

    International Nuclear Information System (INIS)

    Pzdemir, Z.O.; Bulut, E.K.; Mustafeva, Z.; Karahan, M.

    2010-01-01

    Peptides and proteins play a central role in numerous biological and physiological processes in living organisms. Viral capsid peptides are part of the viruses' outer shell of genetic materials. Viruses are recognized by immune system via capsid peptides. Depending on this property of capsid peptides, prototypes synthetic peptide-based vaccine can be developed. In this work, we synthesized three different viral peptide sequences of foot-and-mouth disease virus with microwave enhanced solid phase synthesis method. These peptides were characterized by using liquid chromatography electro spray interface mass spectrometry (LC-ESI-MS) with electro spray ionization. We briefly describe the essential facts for peptide characterization. (author)

  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. 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.

  17. Analysis of a new strain of Euphorbia mosaic virus with distinct replication specificity unveils a lineage of begomoviruses with short Rep sequences in the DNA-B intergenic region

    Directory of Open Access Journals (Sweden)

    Argüello-Astorga Gerardo R

    2010-10-01

    Full Text Available Abstract Background Euphorbia mosaic virus (EuMV is a member of the SLCV clade, a lineage of New World begomoviruses that display distinctive features in their replication-associated protein (Rep and virion-strand replication origin. The first entirely characterized EuMV isolate is native from Yucatan Peninsula, Mexico; subsequently, EuMV was detected in weeds and pepper plants from another region of Mexico, and partial DNA-A sequences revealed significant differences in their putative replication specificity determinants with respect to EuMV-YP. This study was aimed to investigate the replication compatibility between two EuMV isolates from the same country. Results A new isolate of EuMV was obtained from pepper plants collected at Jalisco, Mexico. Full-length clones of both genomic components of EuMV-Jal were biolistically inoculated into plants of three different species, which developed symptoms indistinguishable from those induced by EuMV-YP. Pseudorecombination experiments with EuMV-Jal and EuMV-YP genomic components demonstrated that these viruses do not form infectious reassortants in Nicotiana benthamiana, presumably because of Rep-iteron incompatibility. Sequence analysis of the EuMV-Jal DNA-B intergenic region (IR led to the unexpected discovery of a 35-nt-long sequence that is identical to a segment of the rep gene in the cognate viral DNA-A. Similar short rep sequences ranging from 35- to 51-nt in length were identified in all EuMV isolates and in three distinct viruses from South America related to EuMV. These short rep sequences in the DNA-B IR are positioned downstream to a ~160-nt non-coding domain highly similar to the CP promoter of begomoviruses belonging to the SLCV clade. Conclusions EuMV strains are not compatible in replication, indicating that this begomovirus species probably is not a replicating lineage in nature. The genomic analysis of EuMV-Jal led to the discovery of a subgroup of SLCV clade viruses that contain in

  18. 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.

  19. Structural Characterization of Peptide Antibodies

    DEFF Research Database (Denmark)

    Chailyan, Anna; Marcatili, Paolo

    2015-01-01

    The role of proteins as very effective immunogens for the generation of antibodies is indisputable. Nevertheless, cases in which protein usage for antibody production is not feasible or convenient compelled the creation of a powerful alternative consisting of synthetic peptides. Synthetic peptides...... can be modified to obtain desired properties or conformation, tagged for purification, isotopically labeled for protein quantitation or conjugated to immunogens for antibody production. The antibodies that bind to these peptides represent an invaluable tool for biological research and discovery....... To better understand the underlying mechanisms of antibody-antigen interaction here we present a pipeline developed by us to structurally classify immunoglobulin antigen binding sites and to infer key sequence residues and other variables that have a prominent role in each structural class....

  20. Human Vitronectin-Derived Peptide Covalently Grafted onto Titanium Surface Improves Osteogenic Activity: A Pilot In Vivo Study on Rabbits.

    Science.gov (United States)

    Cacchioli, Antonio; Ravanetti, Francesca; Bagno, Andrea; Dettin, Monica; Gabbi, Carlo

    2009-10-01

    Peptide and protein exploitation for the biochemical functionalization of biomaterial surfaces allowed fabricating biomimetic devices able to evoke and promote specific and advantageous cell functions in vitro and in vivo. In particular, cell adhesion improvement to support the osseointegration of implantable devices has been thoroughly investigated. This study was aimed at checking the biological activity of the (351-359) human vitronectin precursor (HVP) sequence, mapped on the human vitronectin protein; the peptide was covalently linked to the surface of titanium cylinders, surgically inserted in the femurs of New Zealand white rabbits and analyzed at short experimental time points (4, 9, and 16 days after surgery). To assess the osteogenic activity of the peptide, three vital fluorochromic bone markers were used (calcein green, xylenol orange, and calcein blue) to stain the areas of newly grown bone. Static and dynamic histomorphometric parameters were measured at the bone-implant interface and at different distances from the surface. The biological role of the (351-359)HVP sequence was checked by comparing peptide-grafted samples and controls, analyzing how and how much its effects change with time across the bone regions surrounding the implant surface. The results obtained reveal a major activity of the investigated peptide 4 days after surgery, within the bone region closest to the implant surface, and larger bone to implant contact 9 and 16 days after surgery. Thus, improved primary fixation of endosseous devices can be foreseen, resulting in an increased osteointegration.

  1. Self-assembling peptide semiconductors

    Science.gov (United States)

    Tao, Kai; Makam, Pandeeswar; Aizen, Ruth; Gazit, Ehud

    2017-01-01

    Semiconductors are central to the modern electronics and optics industries. Conventional semiconductive materials bear inherent limitations, especially in emerging fields such as interfacing with biological systems and bottom-up fabrication. A promising candidate for bioinspired and durable nanoscale semiconductors is the family of self-assembled nanostructures comprising short peptides. The highly ordered and directional intermolecular π-π interactions and hydrogen-bonding network allow the formation of quantum confined structures within the peptide self-assemblies, thus decreasing the band gaps of the superstructures into semiconductor regions. As a result of the diverse architectures and ease of modification of peptide self-assemblies, their semiconductivity can be readily tuned, doped, and functionalized. Therefore, this family of electroactive supramolecular materials may bridge the gap between the inorganic semiconductor world and biological systems. PMID:29146781

  2. Novel Zn2+-chelating peptides selected from a fimbria-displayed random peptide library

    DEFF Research Database (Denmark)

    Kjærgaard, Kristian; Schembri, Mark; Klemm, Per

    2001-01-01

    The display of peptide sequences on the surface of bacteria is a technology that offers exciting applications in biotechnology and medical research. Type 1 fimbriae are surface organelles of Escherichia coli which mediate D-mannose-sensitive binding to different host surfaces by virtue of the Fim......H adhesin. FimH is a component of the fimbrial organelle that can accommodate and display a diverse range of peptide sequences on the E. coli cell surface. In this study we have constructed a random peptide library in FimH. The library, consisting of similar to 40 million individual clones, was screened...

  3. 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.

  4. 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.

  5. 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.

  6. SANDPUMA: ensemble predictions of nonribosomal peptide chemistry reveal biosynthetic diversity across Actinobacteria

    NARCIS (Netherlands)

    Chevrette, Marc G.; Aicheler, Fabian; Kohlbacher, Oliver; Currie, Cameron R.; Medema, M.H.

    2017-01-01

    Nonribosomally synthesized peptides (NRPs) are natural products with widespread applications in medicine and biotechnology. Many algorithms have been developed to predict the substrate specificities of nonribosomal peptide synthetase adenylation (A) domains from DNA sequences, which enables

  7. Artificial neural network study on organ-targeting peptides

    Science.gov (United States)

    Jung, Eunkyoung; Kim, Junhyoung; Choi, Seung-Hoon; Kim, Minkyoung; Rhee, Hokyoung; Shin, Jae-Min; Choi, Kihang; Kang, Sang-Kee; Lee, Nam Kyung; Choi, Yun-Jaie; Jung, Dong Hyun

    2010-01-01

    We report a new approach to studying organ targeting of peptides on the basis of peptide sequence information. The positive control data sets consist of organ-targeting peptide sequences identified by the peroral phage-display technique for four organs, and the negative control data are prepared from random sequences. The capacity of our models to make appropriate predictions is validated by statistical indicators including sensitivity, specificity, enrichment curve, and the area under the receiver operating characteristic (ROC) curve (the ROC score). VHSE descriptor produces statistically significant training models and the models with simple neural network architectures show slightly greater predictive power than those with complex ones. The training and test set statistics indicate that our models could discriminate between organ-targeting and random sequences. We anticipate that our models will be applicable to the selection of organ-targeting peptides for generating peptide drugs or peptidomimetics.

  8. Breast MRI at very short TE (minTE). Image analysis of minTE sequences on non-fat-saturated, subtracted T1-weighted images

    International Nuclear Information System (INIS)

    Wenkel, Evelyn; Janka, Rolf; Kaemmerer, Nadine; Uder, Michael; Hammon, Matthias; Brand, Michael; Hartmann, Arndt

    2017-01-01

    The aim was to evaluate a minimum echo time (minTE) protocol for breast magnetic resonance imaging (MRI) in patients with breast lesions compared to a standard TE (nTE) time protocol. Breasts of 144 women were examined with a 1.5 Tesla MRI scanner. Additionally to the standard gradient-echo sequence with nTE (4.8 ms), a variant with minimum TE (1.2 ms) was used in an interleaved fashion which leads to a better temporal resolution and should reduce the scan time by approximately 50%. Lesion sizes were measured and the signal-to-noise ratio (SNR) as well as the contrast-to-noise ratio (CNR) were calculated. Subjective confidence was evaluated using a 3-point scale before looking at the nTE sequences (1 = very sure that I can identify a lesion and classify it, 2 = quite sure that I can identify a lesion and classify it, 3 = definitely want to see nTE for final assessment) and the subjective image quality of all examinations was evaluated using a four-grade scale (1 = sharp, 2 = slight blur, 3 = moderate blur and 4 = severe blur/not evaluable) for lesion and skin sharpness. Lesion morphology and contrast enhancement were also evaluated. With minTE sequences, no lesion was rated with ''definitely want to see nTE sequences for final assessment''. The difference of the longitudinal and transverse diameter did not differ significantly (p>0.05). With minTE, lesions and skin were rated to be significantly more blurry (p<0.01 for lesions and p<0.05 for skin). There was no difference between both sequences with respect to SNR, CNR, lesion morphology, contrast enhancement and detection of multifocal disease. Dynamic breast MRI with a minTE protocol is feasible without a major loss of information (SNR, CNR, lesion morphology, contrast enhancement and lesion sizes) and the temporal resolution can be increased by a factor of 2 using minTE sequences.

  9. Breast MRI at very short TE (minTE). Image analysis of minTE sequences on non-fat-saturated, subtracted T1-weighted images

    Energy Technology Data Exchange (ETDEWEB)

    Wenkel, Evelyn; Janka, Rolf; Kaemmerer, Nadine; Uder, Michael; Hammon, Matthias; Brand, Michael [Univ. Hospital Erlangen (Germany). Dept. of Radiology; Geppert, Christian [Siemens Healthcare GmbH, Erlangen (Germany); Hartmann, Arndt [Univ. Hospital Erlangen (Germany). Dept. of Pathology

    2017-02-15

    The aim was to evaluate a minimum echo time (minTE) protocol for breast magnetic resonance imaging (MRI) in patients with breast lesions compared to a standard TE (nTE) time protocol. Breasts of 144 women were examined with a 1.5 Tesla MRI scanner. Additionally to the standard gradient-echo sequence with nTE (4.8 ms), a variant with minimum TE (1.2 ms) was used in an interleaved fashion which leads to a better temporal resolution and should reduce the scan time by approximately 50%. Lesion sizes were measured and the signal-to-noise ratio (SNR) as well as the contrast-to-noise ratio (CNR) were calculated. Subjective confidence was evaluated using a 3-point scale before looking at the nTE sequences (1 = very sure that I can identify a lesion and classify it, 2 = quite sure that I can identify a lesion and classify it, 3 = definitely want to see nTE for final assessment) and the subjective image quality of all examinations was evaluated using a four-grade scale (1 = sharp, 2 = slight blur, 3 = moderate blur and 4 = severe blur/not evaluable) for lesion and skin sharpness. Lesion morphology and contrast enhancement were also evaluated. With minTE sequences, no lesion was rated with ''definitely want to see nTE sequences for final assessment''. The difference of the longitudinal and transverse diameter did not differ significantly (p>0.05). With minTE, lesions and skin were rated to be significantly more blurry (p<0.01 for lesions and p<0.05 for skin). There was no difference between both sequences with respect to SNR, CNR, lesion morphology, contrast enhancement and detection of multifocal disease. Dynamic breast MRI with a minTE protocol is feasible without a major loss of information (SNR, CNR, lesion morphology, contrast enhancement and lesion sizes) and the temporal resolution can be increased by a factor of 2 using minTE sequences.

  10. Development of novel ligands for peptide GPCRs.

    Science.gov (United States)

    Moran, Brian M; McKillop, Aine M; O'Harte, Finbarr Pm

    2016-12-01

    Incretin based glucagon-like peptide-1 receptor (GLP-1R) agonists which target a G-protein coupled receptor (GPCR) are currently used in the treatment of type 2 diabetes. This review focuses on GPCRs from pancreatic β-cells, including GLP-1, glucose-dependent insulinotropic polypeptide (GIP), glucagon, somatostatin, pancreatic polypeptide (PP), cholecystokinin (CCK), peptide YY (PYY), oxyntomodulin (OXM) and ghrelin receptors. In addition, fatty acids GPCRs are thought to have an increasing role in regulating peptide secretions namely short fatty acids GPCR (GPR41, GPR43), medium chain fatty acid GPCR (GPR84), long chain fatty acid GPCR (GPR40, GPR120) and cannabinoid-like GPCR (GPR55, GPR119). Several pre-clinical and clinical trials are currently ongoing in peptide GPCR based therapies, including dual and triple agonist peptides which activate two or more GPCRs simultaneously. Copyright © 2016 Elsevier Ltd. All rights reserved.

  11. An efficient method for isolating antibody fragments against small peptides by antibody phage display

    DEFF Research Database (Denmark)

    Duan, Zhi; Siegumfeldt, Henrik

    2010-01-01

    We generated monoclonal scFv (single chain variable fragment) antibodies from an antibody phage display library towards three small synthetic peptides derived from the sequence of s1-casein. Key difficulties for selection of scFv-phages against small peptides were addressed. Small peptides do....... The scFvs were sequenced and characterized, and specificity was characterized by ELISA. The methods developed in this study are universally applicable for antibody phage display to efficiently produce antibody fragments against small peptides....

  12. A short note on the paper of Liu et al. (2012). A relative Lempel-Ziv complexity: Application to comparing biological sequences. Chemical Physics Letters, volume 530, 19 March 2012, pages 107-112

    Science.gov (United States)

    Arit, Turkan; Keskin, Burak; Firuzan, Esin; Cavas, Cagin Kandemir; Liu, Liwei; Cavas, Levent

    2018-04-01

    The report entitled "L. Liu, D. Li, F. Bai, A relative Lempel-Ziv complexity: Application to comparing biological sequences, Chem. Phys. Lett. 530 (2012) 107-112" mentions on the powerful construction of phylogenetic trees based on Lempel-Ziv algorithm. On the other hand, the method explained in the paper does not give promising result on the data set on invasive Caulerpa taxifolia in the Mediterranean Sea. The phylogenetic trees are obtained by the proposed method of the aforementioned paper in this short note.

  13. Peptide imprinted receptors for the determination of the small cell lung cancer associated biomarker progastrin releasing peptide

    DEFF Research Database (Denmark)

    Qader, A. A.; Urraca, J.; Torsetnes, S. B.

    2014-01-01

    Peptide imprinted polymers were developed for detection of progastrin releasing peptide (ProGRP); a low abundant blood based biomarker for small cell lung cancer. The polymers targeted the proteotypic nona-peptide sequence NLLGLIEAK and were used for selective enrichment of the proteotypic peptide...... prior to LCMS based quantification. Peptide imprinted polymers with the best affinity characteristics were first identified from a 96-polymer combinatorial library. The effects of functional monomers, crosslinker, porogen, and template on adsorption capacity and selectivity for NLLGLIEAK were...

  14. 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...

  15. Double quick, double click reversible peptide "stapling".

    Science.gov (United States)

    Grison, Claire M; Burslem, George M; Miles, Jennifer A; Pilsl, Ludwig K A; Yeo, David J; Imani, Zeynab; Warriner, Stuart L; Webb, Michael E; Wilson, Andrew J

    2017-07-01

    The development of constrained peptides for inhibition of protein-protein interactions is an emerging strategy in chemical biology and drug discovery. This manuscript introduces a versatile, rapid and reversible approach to constrain peptides in a bioactive helical conformation using BID and RNase S peptides as models. Dibromomaleimide is used to constrain BID and RNase S peptide sequence variants bearing cysteine (Cys) or homocysteine ( h Cys) amino acids spaced at i and i + 4 positions by double substitution. The constraint can be readily removed by displacement of the maleimide using excess thiol. This new constraining methodology results in enhanced α-helical conformation (BID and RNase S peptide) as demonstrated by circular dichroism and molecular dynamics simulations, resistance to proteolysis (BID) as demonstrated by trypsin proteolysis experiments and retained or enhanced potency of inhibition for Bcl-2 family protein-protein interactions (BID), or greater capability to restore the hydrolytic activity of the RNAse S protein (RNase S peptide). Finally, use of a dibromomaleimide functionalized with an alkyne permits further divergent functionalization through alkyne-azide cycloaddition chemistry on the constrained peptide with fluorescein, oligoethylene glycol or biotin groups to facilitate biophysical and cellular analyses. Hence this methodology may extend the scope and accessibility of peptide stapling.

  16. 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

  17. Towards Identify Selective Antibacterial Peptides Based on Abstracts Meaning

    Directory of Open Access Journals (Sweden)

    Liliana I. Barbosa-Santillán

    2016-01-01

    Full Text Available We present an Identify Selective Antibacterial Peptides (ISAP approach based on abstracts meaning. Laboratories and researchers have significantly increased the report of their discoveries related to antibacterial peptides in primary publications. It is important to find antibacterial peptides that have been reported in primary publications because they can produce antibiotics of different generations that attack and destroy the bacteria. Unfortunately, researchers used heterogeneous forms of natural language to describe their discoveries (sometimes without the sequence of the peptides. Thus, we propose that learning the words meaning instead of the antibacterial peptides sequence is possible to identify and predict antibacterial peptides reported in the PubMed engine. The ISAP approach consists of two stages: training and discovering. ISAP founds that the 35% of the abstracts sample had antibacterial peptides and we tested in the updated Antimicrobial Peptide Database 2 (APD2. ISAP predicted that 45% of the abstracts had antibacterial peptides. That is, ISAP found that 810 antibacterial peptides were not classified like that, so they are not reported in APD2. As a result, this new search tool would complement the APD2 with a set of peptides that are candidates to be antibacterial. Finally, 20% of the abstracts were not semantic related to APD2.

  18. Radio-immunoassays for glucagon-like peptides 1 and 2 (GLP-1 and GLP-2)

    DEFF Research Database (Denmark)

    Orskov, C; Holst, J J

    1987-01-01

    Gene-sequencing studies have shown that the glucagon precursor contains two additional glucagon-like sequences, the so-called glucagon-like peptides 1 and 2 (GLP-1 and GLP-2). We developed radio-immunoassays against synthetic peptides corresponding to these sequences. Antisera were raised in rabb...

  19. Hydroxyapatite-binding peptides for bone growth and inhibition

    Science.gov (United States)

    Bertozzi, Carolyn R [Berkeley, CA; Song, Jie [Shrewsbury, MA; Lee, Seung-Wuk [Walnut Creek, CA

    2011-09-20

    Hydroxyapatite (HA)-binding peptides are selected using combinatorial phage library display. Pseudo-repetitive consensus amino acid sequences possessing periodic hydroxyl side chains in every two or three amino acid sequences are obtained. These sequences resemble the (Gly-Pro-Hyp).sub.x repeat of human type I collagen, a major component of extracellular matrices of natural bone. A consistent presence of basic amino acid residues is also observed. The peptides are synthesized by the solid-phase synthetic method and then used for template-driven HA-mineralization. Microscopy reveal that the peptides template the growth of polycrystalline HA crystals .about.40 nm in size.

  20. Virtual screening using combinatorial cyclic peptide libraries reveals protein interfaces readily targetable by cyclic peptides.

    Science.gov (United States)

    Duffy, Fergal J; O'Donovan, Darragh; Devocelle, Marc; Moran, Niamh; O'Connell, David J; Shields, Denis C

    2015-03-23

    Protein-protein and protein-peptide interactions are responsible for the vast majority of biological functions in vivo, but targeting these interactions with small molecules has historically been difficult. What is required are efficient combined computational and experimental screening methods to choose among a number of potential protein interfaces worthy of targeting lead macrocyclic compounds for further investigation. To achieve this, we have generated combinatorial 3D virtual libraries of short disulfide-bonded peptides and compared them to pharmacophore models of important protein-protein and protein-peptide structures, including short linear motifs (SLiMs), protein-binding peptides, and turn structures at protein-protein interfaces, built from 3D models available in the Protein Data Bank. We prepared a total of 372 reference pharmacophores, which were matched against 108,659 multiconformer cyclic peptides. After normalization to exclude nonspecific cyclic peptides, the top hits notably are enriched for mimetics of turn structures, including a turn at the interaction surface of human α thrombin, and also feature several protein-binding peptides. The top cyclic peptide hits also cover the critical "hot spot" interaction sites predicted from the interaction crystal structure. We have validated our method by testing cyclic peptides predicted to inhibit thrombin, a key protein in the blood coagulation pathway of important therapeutic interest, identifying a cyclic peptide inhibitor with lead-like activity. We conclude that protein interfaces most readily targetable by cyclic peptides and related macrocyclic drugs may be identified computationally among a set of candidate interfaces, accelerating the choice of interfaces against which lead compounds may be screened.

  1. Growth hormone-releasing peptides.

    Science.gov (United States)

    Ghigo, E; Arvat, E; Muccioli, G; Camanni, F

    1997-05-01

    Growth hormone-releasing peptides (GHRPs) are synthetic, non-natural peptides endowed with potent stimulatory effects on somatotrope secretion in animals and humans. They have no structural homology with GHRH and act via specific receptors present either at the pituitary or the hypothalamic level both in animals and in humans. The GHRP receptor has recently been cloned and, interestingly, it does not show sequence homology with other G-protein-coupled receptors known so far. This evidence strongly suggests the existence of a natural GHRP-like ligand which, however, has not yet been found. The mechanisms underlying the GHRP effect are still unclear. At present, several data favor the hypothesis that GHRPs could act by counteracting somatostatinergic activity both at the pituitary and the hypothalamic level and/or, at least partially, via a GHRH-mediated mechanism. However, the possibility that GHRPs act via an unknown hypothalamic factor (U factor) is still open. GHRP-6 was the first hexapeptide to be extensively studied in humans. More recently, a heptapeptide, GHRP-1, and two other hexapeptides, GHRP-2 and Hexarelin, have been synthesized and are now available for human studies. Moreover, non-peptidyl GHRP mimetics have been developed which act via GHRP receptors and their effects have been clearly demonstrated in animals and in humans in vivo. Among non-peptidyl GHRPs, MK-0677 seems the most interesting molecule. The GH-releasing activity of GHRPs is marked and dose-related after intravenous, subcutaneous, intranasal and even oral administration. The effect of GHRPs is reproducible and undergoes partial desensitization, more during continuous infusion, less during intermittent administration: in fact, prolonged administration of GHRPs increases IGF-1 levels both in animals and in humans. The GH-releasing effect of GHRPs does not depend on sex but undergoes age-related variations. It increases from birth to puberty, persists at a similar level in adulthood and

  2. Two RNAs or DNAs May Artificially Fuse Together at a Short Homologous Sequence (SHS) during Reverse Transcription or Polymerase Chain Reactions, and Thus Reporting an SHS-Containing Chimeric RNA Requires Extra Caution

    Science.gov (United States)

    Xie, Bingkun; Yang, Wei; Ouyang, Yongchang; Chen, Lichan; Jiang, Hesheng; Liao, Yuying; Liao, D. Joshua

    2016-01-01

    Tens of thousands of chimeric RNAs have been reported. Most of them contain a short homologous sequence (SHS) at the joining site of the two partner genes but are not associated with a fusion gene. We hypothesize that many of these chimeras may be technical artifacts derived from SHS-caused mis-priming in reverse transcription (RT) or polymerase chain reactions (PCR). We cloned six chimeric complementary DNAs (cDNAs) formed by human mitochondrial (mt) 16S rRNA sequences at an SHS, which were similar to several expression sequence tags (ESTs).These chimeras, which could not be detected with cDNA protection assay, were likely formed because some regions of the 16S rRNA are reversely complementary to another region to form an SHS, which allows the downstream sequence to loop back and anneal at the SHS to prime the synthesis of its complementary strand, yielding a palindromic sequence that can form a hairpin-like structure.We identified a 16S rRNA that ended at the 4th nucleotide(nt) of the mt-tRNA-leu was dominant and thus should be the wild type. We also cloned a mouse Bcl2-Nek9 chimeric cDNA that contained a 5-nt unmatchable sequence between the two partners, contained two copies of the reverse primer in the same direction but did not contain the forward primer, making it unclear how this Bcl2-Nek9 was formed and amplified. Moreover, a cDNA was amplified because one primer has 4 nts matched to the template, suggesting that there may be many more artificial cDNAs than we have realized, because the nuclear and mt genomes have many more 4-nt than 5-nt or longer homologues. Altogether, the chimeric cDNAs we cloned are good examples suggesting that many cDNAs may be artifacts due to SHS-caused mis-priming and thus greater caution should be taken when new sequence is obtained from a technique involving DNA polymerization. PMID:27148738

  3. Expression of the cationic antimicrobial peptide lactoferricin fused with the anionic peptide in Escherichia coli.

    Science.gov (United States)

    Kim, Ha-Kun; Chun, Dae-Sik; Kim, Joon-Sik; Yun, Cheol-Ho; Lee, Ju-Hoon; Hong, Soon-Kwang; Kang, Dae-Kyung

    2006-09-01

    Direct expression of lactoferricin, an antimicrobial peptide, is lethal to Escherichia coli. For the efficient production of lactoferricin in E. coli, we developed an expression system in which the gene for the lysine- and arginine-rich cationic lactoferricin was fused to an anionic peptide gene to neutralize the basic property of lactoferricin, and successfully overexpressed the concatemeric fusion gene in E. coli. The lactoferricin gene was linked to a modified magainin intervening sequence gene by a recombinational polymerase chain reaction, thus producing an acidic peptide-lactoferricin fusion gene. The monomeric acidic peptide-lactoferricin fusion gene was multimerized and expressed in E. coli BL21(DE3) upon induction with isopropyl-beta-D-thiogalactopyranoside. The expression levels of the fusion peptide reached the maximum at the tetramer, while further increases in the copy number of the fusion gene substantially reduced the peptide expression level. The fusion peptides were isolated and cleaved to generate the separate lactoferricin and acidic peptide. About 60 mg of pure recombinant lactoferricin was obtained from 1 L of E. coli culture. The purified recombinant lactoferricin was found to have a molecular weight similar to that of chemically synthesized lactoferricin. The recombinant lactoferricin showed antimicrobial activity and disrupted bacterial membrane permeability, as the native lactoferricin peptide does.

  4. Halogenation dictates the architecture of amyloid peptide nanostructures.

    Science.gov (United States)

    Pizzi, Andrea; Pigliacelli, Claudia; Gori, Alessandro; Nonappa; Ikkala, Olli; Demitri, Nicola; Terraneo, Giancarlo; Castelletto, Valeria; Hamley, Ian W; Baldelli Bombelli, Francesca; Metrangolo, Pierangelo

    2017-07-20

    Amyloid peptides yield a plethora of interesting nanostructures though difficult to control. Here we report that depending on the number, position, and nature of the halogen atoms introduced into either one or both phenylalanine benzene rings of the amyloid β peptide-derived core-sequence KLVFF, four different architectures were obtained in a controlled manner. Our findings demonstrate that halogenation may develop as a general strategy to engineer amyloidal peptide self-assembly and obtain new amyloidal nanostructures.

  5. Optimizing the identification of citrullinated peptides by mass spectrometry

    DEFF Research Database (Denmark)

    Bennike, Tue; Lauridsen, Kasper B.; Olesen, Michael Kruse

    2013-01-01

    Citrullinated proteins have been associated with several diseases and citrullination can most likely function as a target for novel diagnostic agents and unravel disease etiologies. The correct identification of citrullinated proteins is therefore of most importance. Mass spectrometry (MS) driven...... of trypsin, digestion was performed on synthetic peptide sets containing either arginine or citrulline. The peptide sequences originated from disease-associated in vivo citrullinated proteins; some reported as being C-terminal tryptic citrullinated peptides. Furthermore, the proteolytic activity was verified...

  6. Recent progress in fluorine-18 labelled peptide radiopharmaceuticals

    Energy Technology Data Exchange (ETDEWEB)

    Okarvi, S.M. [Cyclotron and Radiopharmaceuticals Department, King Faisal Specialist Hospital and Research Centre, Riyadh (Saudi Arabia)

    2001-07-01

    The application of biologically active peptides labelled with positron-emitting nuclides has emerged as a useful and interesting field in nuclear medicine. Small synthetic receptor-binding peptides are currently the preferred agents over proteins and antibodies for diagnostic imaging of various tumours. Due to the smaller size of peptides, both higher target-to-background ratios and rapid blood clearance can often be achieved with radiolabelled peptides. Hence, short-lived positron emission tomography (PET) isotopes are potential candidates for labelling peptides. Among a number of positron-emitting nuclides, fluorine-18 appears to be the best candidate for labelling bioactive peptides by virtue of its favourable physical and nuclear characteristics. The major disadvantage of labelling peptides with {sup 18}F is the laborious and time-consuming preparation of the {sup 18}F labelling agents. In recent years, various techniques have been developed which allow efficient labelling of peptides with {sup 18}F without affecting their receptor-binding properties. Moreover, the development of a variety of prosthetic groups has facilitated the efficient and site-specific labelling of peptides with {sup 18}F. The {sup 18}F-labelled peptides hold enormous clinical potential owing to their ability to quantitatively detect and characterise a wide variety of human diseases when using PET. Recently, a number of {sup 18}F-labelled bioactive peptides have shown great promise as diagnostic imaging agents. This review presents the recent developments in {sup 18}F-labelled biologically active peptides used in PET. (orig.)

  7. Recent progress in fluorine-18 labelled peptide radiopharmaceuticals

    International Nuclear Information System (INIS)

    Okarvi, S.M.

    2001-01-01

    The application of biologically active peptides labelled with positron-emitting nuclides has emerged as a useful and interesting field in nuclear medicine. Small synthetic receptor-binding peptides are currently the preferred agents over proteins and antibodies for diagnostic imaging of various tumours. Due to the smaller size of peptides, both higher target-to-background ratios and rapid blood clearance can often be achieved with radiolabelled peptides. Hence, short-lived positron emission tomography (PET) isotopes are potential candidates for labelling peptides. Among a number of positron-emitting nuclides, fluorine-18 appears to be the best candidate for labelling bioactive peptides by virtue of its favourable physical and nuclear characteristics. The major disadvantage of labelling peptides with 18 F is the laborious and time-consuming preparation of the 18 F labelling agents. In recent years, various techniques have been developed which allow efficient labelling of peptides with 18 F without affecting their receptor-binding properties. Moreover, the development of a variety of prosthetic groups has facilitated the efficient and site-specific labelling of peptides with 18 F. The 18 F-labelled peptides hold enormous clinical potential owing to their ability to quantitatively detect and characterise a wide variety of human diseases when using PET. Recently, a number of 18 F-labelled bioactive peptides have shown great promise as diagnostic imaging agents. This review presents the recent developments in 18 F-labelled biologically active peptides used in PET. (orig.)

  8. Novel ZnO-binding peptides obtained by the screening of a phage display peptide library

    Energy Technology Data Exchange (ETDEWEB)

    Golec, Piotr [Institute of Biochemistry and Biophysics, Polish Academy of Sciences, Laboratory of Molecular Biology (affiliated with the University of Gdansk) (Poland); Karczewska-Golec, Joanna [University of Gdansk and Medical University of Gdansk, Laboratory of Molecular Bacteriology, Intercollegiate Faculty of Biotechnology (Poland); Los, Marcin; Wegrzyn, Grzegorz, E-mail: wegrzyn@biotech.univ.gda.pl [University of Gdansk, Department of Molecular Biology (Poland)

    2012-11-15

    Zinc oxide (ZnO) is a semiconductor compound with a potential for wide use in various applications, including biomaterials and biosensors, particularly as nanoparticles (the size range of ZnO nanoparticles is from 2 to 100 nm, with an average of about 35 nm). Here, we report isolation of novel ZnO-binding peptides, by screening of a phage display library. Interestingly, amino acid sequences of the ZnO-binding peptides reported in this paper and those described previously are significantly different. This suggests that there is a high variability in sequences of peptides which can bind particular inorganic molecules, indicating that different approaches may lead to discovery of different peptides of generally the same activity (e.g., binding of ZnO) but having various detailed properties, perhaps crucial under specific conditions of different applications.

  9. Penetration of the signal sequence of Escherichia coli PhoE protein into phospholipid model membranes leads to lipid-specific changes in signal peptide structure and alterations of lipid organization

    International Nuclear Information System (INIS)

    Batenburg, A.M.; Demel, R.A.; Verkleij, A.J.; de Kruijff, B.

    1988-01-01

    In order to obtain more insight in the initial steps of the process of protein translocation across membranes, biophysical investigations were undertaken on the lipid specificity and structural consequences of penetration of the PhoE signal peptide into lipid model membranes and on the conformation of the signal peptide adopted upon interaction with the lipids. When the monolayer technique and differential scanning calorimetry are used, a stronger penetration is observed for negatively charged lipids, significantly influenced by the physical state of the lipid but not by temperature or acyl chain unsaturation as such. Although the interaction is principally electrostatic, as indicated also by the strong penetration of N-terminal fragments into negatively charged lipid monolayers, the effect of ionic strength suggests an additional hydrophobic component. Most interestingly with regard to the mechanism of protein translocation, the molecular area of the peptide in the monolayer also shows lipid specificity: the area in the presence of PC is consistent with a looped helical orientation, whereas in the presence of cardiolipin a time-dependent conformational change is observed, most likely leading from a looped to a stretched orientation with the N-terminus directed toward the water. This is in line also with the determined peptide-lipid stoichiometry. Preliminary 31 P NMR and electron microscopy data on the interaction with lipid bilayer systems indicate loss of bilayer structure

  10. The earthquake sequence of 21-22 February, 1983 at Ramnicu Sarat, Romania: source parameters retrieved by short period local data inversion

    International Nuclear Information System (INIS)

    Ardeleanu, L.; Radulian, M.; Sileny, J.; Panza, G. F.

    2002-01-01

    High-frequency seismograms from the Romanian telemetered local network are inverted to retrieve the seismic moment tensor of five weak earthquakes (2.9 ≤ M L ≤ 3.6) belonging to the seismic sequence of Ramnicu Sarat (Romania) of 21-22 February 1983. A grossly simplified 1-D approximation of the crust - the horizontally layered inelastic earth model - is used to construct the Green's function by modal summation. The deviation from the real velocity structure originates the error in forward modelling, which is roughly estimated from the differences in the moment tensor rate functions obtained from subsets of the complete station network. This error is transformed into estimates of confidence regions of the time function, moment tensor and its principal axes, and error bars of the scalar moment. The 1-D approximation of the crust results in a large uncertainty of the source time function, which is almost completely undetermined. The mechanism is more confident, especially the determination of the orientation of its deviatoric part. The most robust source parameter from the three investigated items is the scalar moment. (authors)

  11. Killing of Mycobacterium avium by lactoferricin peptides: improved activity of arginine- and D-amino-acid-containing molecules.

    Science.gov (United States)

    Silva, Tânia; Magalhães, Bárbara; Maia, Sílvia; Gomes, Paula; Nazmi, Kamran; Bolscher, Jan G M; Rodrigues, Pedro N; Bastos, Margarida; Gomes, Maria Salomé

    2014-06-01

    Mycobacterium avium causes respiratory disease in susceptible individuals, as well as disseminated infections in immunocompromised hosts, being an important cause of morbidity and mortality among these populations. Current therapies consist of a combination of antibiotics taken for at least 6 months, with no more than 60% overall clinical success. Furthermore, mycobacterial antibiotic resistance is increasing worldwide, urging the need to develop novel classes of antimicrobial drugs. One potential and interesting alternative strategy is the use of antimicrobial peptides (AMP). These are present in almost all living organisms as part of their immune system, acting as a first barrier against invading pathogens. In this context, we investigated the effect of several lactoferrin-derived AMP against M. avium. Short peptide sequences from both human and bovine lactoferricins, namely, hLFcin1-11 and LFcin17-30, as well as variants obtained by specific amino acid substitutions, were evaluated. All tested peptides significantly inhibited the axenic growth of M. avium, the bovine peptides being more active than the human. Arginine residues were found to be crucial for the display of antimycobacterial activity, whereas the all-d-amino-acid analogue of the bovine sequence displayed the highest mycobactericidal activity. These findings reveal the promising potential of lactoferricins against mycobacteria, thus opening the way for further research on their development and use as a new weapon against mycobacterial infections. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  12. Peptide Nucleic Acids Having 2,6-Diaminopurine Nucleobases

    DEFF Research Database (Denmark)

    2003-01-01

    A novel class of compounds, known as peptide nucleic acids, bind complementary DNA and RNA strands more strongly than a corresponding DNA strand, and exhibit increased sequence specificity and binding affinity. The peptide nucleic acids of the invention comprise ligands selected from a group...

  13. Peptide Nucleic Acids Having Amino Acid Side Chains

    DEFF Research Database (Denmark)

    1998-01-01

    A novel class of compounds, known as peptide nucleic acids, bind complementary DNA and RNA strands more strongly than the corresponding DNA or RNA strands, and exhibit increased sequence specificity and solubility. The peptide nucleic acids comprise ligands selected from a group consisting...

  14. A Simple PB/LIE Free Energy Function Accurately Predicts the Peptide Binding Specificity of the Tiam1 PDZ Domain.

    Science.gov (United States)

    Panel, Nicolas; Sun, Young Joo; Fuentes, Ernesto J; Simonson, Thomas

    2017-01-01

    PDZ domains generally bind short amino acid sequences at the C-terminus of target proteins, and short peptides can be used as inhibitors or model ligands. Here, we used experimental binding assays and molecular dynamics simulations to characterize 51 complexes involving the Tiam1 PDZ domain and to test the performance of a semi-empirical free energy function. The free energy function combined a Poisson-Boltzmann (PB) continuum electrostatic term, a van der Waals interaction energy, and a surface area term. Each term was empirically weighted, giving a Linear Interaction Energy or "PB/LIE" free energy. The model yielded a mean unsigned deviation of 0.43 kcal/mol and a Pearson correlation of 0.64 between experimental and computed free energies, which was superior to a Null model that assumes all complexes have the same affinity. Analyses of the models support several experimental observations that indicate the orientation of the α 2 helix is a critical determinant for peptide specificity. The models were also used to predict binding free energies for nine new variants, corresponding to point mutants of the Syndecan1 and Caspr4 peptides. The predictions did not reveal improved binding; however, they suggest that an unnatural amino acid could be used to increase protease resistance and peptide lifetimes in vivo . The overall performance of the model should allow its use in the design of new PDZ ligands in the future.

  15. A Simple PB/LIE Free Energy Function Accurately Predicts the Peptide Binding Specificity of the Tiam1 PDZ Domain

    Directory of Open Access Journals (Sweden)

    Nicolas Panel

    2017-09-01

    Full Text Available PDZ domains generally bind short amino acid sequences at the C-terminus of target proteins, and short peptides can be used as inhibitors or model ligands. Here, we used experimental binding assays and molecular dynamics simulations to characterize 51 complexes involving the Tiam1 PDZ domain and to test the performance of a semi-empirical free energy function. The free energy function combined a Poisson-Boltzmann (PB continuum electrostatic term, a van der Waals interaction energy, and a surface area term. Each term was empirically weighted, giving a Linear Interaction Energy or “PB/LIE” free energy. The model yielded a mean unsigned deviation of 0.43 kcal/mol and a Pearson correlation of 0.64 between experimental and computed free energies, which was superior to a Null model that assumes all complexes have the same affinity. Analyses of the models support several experimental observations that indicate the orientation of the α2 helix is a critical determinant for peptide specificity. The models were also used to predict binding free energies for nine new variants, corresponding to point mutants of the Syndecan1 and Caspr4 peptides. The predictions did not reveal improved binding; however, they suggest that an unnatural amino acid could be used to increase protease resistance and peptide lifetimes in vivo. The overall performance of the model should allow its use in the design of new PDZ ligands in the future.

  16. Modern treatment of short bowel syndrome

    DEFF Research Database (Denmark)

    Jeppesen, Palle B

    2013-01-01

    Recently, the US Food and Drug Administration and the European Medicines Agency approved the glucagon-like peptide 2 analogue, teduglutide, for the treatment of short bowel syndrome (SBS), and this review describes the physiological basis for its clinical use.......Recently, the US Food and Drug Administration and the European Medicines Agency approved the glucagon-like peptide 2 analogue, teduglutide, for the treatment of short bowel syndrome (SBS), and this review describes the physiological basis for its clinical use....

  17. 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.

  18. 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...

  19. Genome-wide analysis of short interspersed nuclear elements SINES revealed high sequence conservation, gene association and retrotranspositional activity in wheat.

    Science.gov (United States)

    Ben-David, Smadar; Yaakov, Beery; Kashkush, Khalil

    2013-10-01

    Short interspersed nuclear elements (SINEs) are non-autonomous non-LTR retroelements that are present in most eukaryotic species. While SINEs have been intensively investigated in humans and other animal systems, they are poorly studied in plants, especially in wheat (Triticum aestivum). We used quantitative PCR of various wheat species to determine the copy number of a wheat SINE family, termed Au SINE, combined with computer-assisted analyses of the publicly available 454 pyrosequencing database of T. aestivum. In addition, we utilized site-specific PCR on 57 Au SINE insertions, transposon methylation display and transposon display on newly formed wheat polyploids to assess retrotranspositional activity, epigenetic status and genetic rearrangements in Au SINE, respectively. We retrieved 3706 different insertions of Au SINE from the 454 pyrosequencing database of T. aestivum, and found that most of the elements are inserted in A/T-rich regions, while approximately 38% of the insertions are associated with transcribed regions, including known wheat genes. We observed typical retrotransposition of Au SINE in the second generation of a newly formed wheat allohexaploid, and massive hypermethylation in CCGG sites surrounding Au SINE in the third generation. Finally, we observed huge differences in the copy numbers in diploid Triticum and Aegilops species, and a significant increase in the copy numbers in natural wheat polyploids, but no significant increase in the copy number of Au SINE in the first four generations for two of three newly formed allopolyploid species used in this study. Our data indicate that SINEs may play a prominent role in the genomic evolution of wheat through stress-induced activation. © 2013 Ben-Gurion University The Plant Journal © 2013 John Wiley & Sons Ltd.

  20. Signal-BNF: A Bayesian Network Fusing Approach to Predict Signal Peptides

    Directory of Open Access Journals (Sweden)

    Zhi Zheng

    2012-01-01

    Full Text Available A signal peptide is a short peptide chain that directs the transport of a protein and has become the crucial vehicle in finding new drugs or reprogramming cells for gene therapy. As the avalanche of new protein sequences generated in the postgenomic era, the challenge of identifying new signal sequences has become even more urgent and critical in biomedical engineering. In this paper, we propose a novel predictor called Signal-BNF to predict the N-terminal signal peptide as well as its cleavage site based on Bayesian reasoning network. Signal-BNF is formed by fusing the results of different Bayesian classifiers which used different feature datasets as its input through weighted voting system. Experiment results show that Signal-BNF is superior to the popular online predictors such as Signal-3L and PrediSi. Signal-BNF is featured by high prediction accuracy that may serve as a useful tool for further investigating many unclear details regarding the molecular mechanism of the zip code protein-sorting system in cells.

  1. Roles of beta-turns in protein folding: from peptide models to protein engineering.

    Science.gov (United States)

    Marcelino, Anna Marie C; Gierasch, Lila M

    2008-05-01

    Reverse turns are a major class of protein secondary structure; they represent sites of chain reversal and thus sites where the globular character of a protein is created. It has been speculated for many years that turns may nucleate the formation of structure in protein folding, as their propensity to occur will favor the approximation of their flanking regions and their general tendency to be hydrophilic will favor their disposition at the solvent-accessible surface. Reverse turns are local features, and it is therefore not surprising that their structural properties have been extensively studied using peptide models. In this article, we review research on peptide models of turns to test the hypothesis that the propensities of turns to form in short peptides will relate to the roles of corresponding sequences in protein folding. Turns with significant stability as isolated entities should actively promote the folding of a protein, and by contrast, turn sequences that merely allow the chain to adopt conformations required for chain reversal are predicted to be passive in the folding mechanism. We discuss results of protein engineering studies of the roles of turn residues in folding mechanisms. Factors that correlate with the importance of turns in folding indeed include their intrinsic stability, as well as their topological context and their participation in hydrophobic networks within the protein's structure.

  2. Amyloid fibril formation from sequences of a natural beta-structured fibrous protein, the adenovirus fiber.

    Science.gov (United States)

    Papanikolopoulou, Katerina; Schoehn, Guy; Forge, Vincent; Forsyth, V Trevor; Riekel, Christian; Hernandez, Jean-François; Ruigrok, Rob W H; Mitraki, Anna

    2005-01-28

    Amyloid fibrils are fibrous beta-structures that derive from abnormal folding and assembly of peptides and proteins. Despite a wealth of structural studies on amyloids, the nature of the amyloid structure remains elusive; possible connections to natural, beta-structured fibrous motifs have been suggested. In this work we focus on understanding amyloid structure and formation from sequences of a natural, beta-structured fibrous protein. We show that short peptides (25 to 6 amino acids) corresponding to repetitive sequences from the adenovirus fiber shaft have an intrinsic capacity to form amyloid fibrils as judged by electron microscopy, Congo Red binding, infrared spectroscopy, and x-ray fiber diffraction. In the presence of the globular C-terminal domain of the protein that acts as a trimerization motif, the shaft sequences adopt a triple-stranded, beta-fibrous motif. We discuss the possible structure and arrangement of these sequences within the amyloid fibril, as compared with the one adopted within the native structure. A 6-amino acid peptide, corresponding to the last beta-strand of the shaft, was found to be sufficient to form amyloid fibrils. Structural analysis of these amyloid fibrils suggests that perpendicular stacking of beta-strand repeat units is an underlying common feature of amyloid formation.

  3. The preparation and identification of peptide imaging agent of lung cancer

    International Nuclear Information System (INIS)

    Chu Liping; Wang Yan; Wang Yueying; Liu Jinjian; Wu Hongying; Liu Jianfeng

    2010-01-01

    Objective: To screen in vivo lung cancer specific binding 7-peptide from T7 phage display random peptide library and prepare peptide imaging agent in early in early diagnosis of lung cancer. Methods: Used phage display in vivo technology to get the 7-peptide phage that can bind the lung cancer specifically, then sequenced and synthesized 7-peptide. After being labeled by 125 I, this 7-peptide was injected into mice via vein and the distribution in the mice tumor mold was observed. Results: One 7-peptide was obtained after four rounds of screening, and the peptide could bind lung cancer tissue specifically. Metabolism of this peptide in mice was fast and imaging of lung cancer was best two hours later after injection. The distribution in vivo decreased and almost disappeared after six hours. Conclusion: This 7-peptide could be used to image and diagnose of lung cancer effectively. (authors)

  4. Using a biomimetic membrane surface experiment to investigate the activity of the magnetite biomineralisation protein Mms6† †Electronic supplementary information (ESI) available: Including Mms6 protein and peptide sequences, additional QCM-D and SEM data and protein modelling. See DOI: 10.1039/c5ra16469a Click here for additional data file.

    Science.gov (United States)

    Bird, Scott M.; Rawlings, Andrea E.; Galloway, Johanna M.

    2016-01-01

    Magnetotactic bacteria are able to synthesise precise nanoparticles of the iron oxide magnetite within their cells. These particles are formed in dedicated organelles termed magnetosomes. These lipid membrane compartments use a range of biomineralisation proteins to nucleate and regulate the magnetite crystallisation process. A key component is the membrane protein Mms6, which binds to iron ions and helps to control the formation of the inorganic core. We have previously used Mms6 on gold surfaces patterned with a self-assembled monolayer to successfully produce arrays of magnetic nanoparticles. Here we use this surface system as a mimic of the interior face of the magnetosome membrane to study differences between intact Mms6 and the acid-rich C-terminal peptide subregion of the Mms6 protein. When immobilised on surfaces, the peptide is unable to reproduce the particle size or homogeneity control exhibited by the full Mms6 protein in our experimental setup. Moreover, the peptide is unable to support anchoring of a dense array of nanoparticles to the surface. This system also allows us to deconvolute particle binding from particle nucleation, and shows that Mms6 particle binding is less efficient when supplied with preformed magnetite nanoparticles when compared to particles precipitated from solution in the presence of the surface immobilised Mms6. This suggests that Mms6 binds to iron ions rather than to magnetite surfaces in our system, and is perhaps a nucleating agent rather than a controller of magnetite crystal growth. The comparison between the peptide and the protein under identical experimental conditions indicates that the full length sequence is required to support the full function of Mms6 on surfaces. PMID:27019707

  5. Antimicrobial peptides design by evolutionary multiobjective optimization.

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    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.

  6. Optimization of de novo transcriptome assembly from high-throughput short read sequencing data improves functional annotation for non-model organisms

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    Haznedaroglu Berat Z

    2012-07-01

    Full Text Available Abstract Background The k-mer hash length is a key factor affecting the output of de novo transcriptome assembly packages using de Bruijn graph algorithms. Assemblies constructed with varying single k-mer choices might result in the loss of unique contiguous sequences (contigs and relevant biological information. A common solution to this problem is the clustering of single k-mer assemblies. Even though annotation is one of the primary goals of a transcriptome assembly, the success of assembly strategies does not consider the impact of k-mer selection on the annotation output. This study provides an in-depth k-mer selection analysis that is focused on the degree of functional annotation achieved for a non-model organism where no reference genome information is available. Individual k-mers and clustered assemblies (CA were considered using three representative software packages. Pair-wise comparison analyses (between individual k-mers and CAs were produced to reveal missing Kyoto Encyclopedia of Genes and Genomes (KEGG ortholog identifiers (KOIs, and to determine a strategy that maximizes the recovery of biological information in a de novo transcriptome assembly. Results Analyses of single k-mer assemblies resulted in the generation of various quantities of contigs and functional annotations within the selection window of k-mers (k-19 to k-63. For each k-mer in this window, generated assemblies contained certain unique contigs and KOIs that were not present in the other k-mer assemblies. Producing a non-redundant CA of k-mers 19 to 63 resulted in a more complete functional annotation than any single k-mer assembly. However, a fraction of unique annotations remained (~0.19 to 0.27% of total KOIs in the assemblies of individual k-mers (k-19 to k-63 that were not present in the non-redundant CA. A workflow to recover these unique annotations is presented. Conclusions This study demonstrated that different k-mer choices result in various quantities

  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. 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....

  9. 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....

  10. Peptide Nucleic Acids (PNA)

    DEFF Research Database (Denmark)

    2002-01-01

    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....

  11. Peptide Nucleic Acid Synthons

    DEFF Research Database (Denmark)

    2004-01-01

    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....

  12. Antimicrobial Peptides in 2014

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    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.

  13. PIPE: a protein-protein interaction prediction engine based on the re-occurring short polypeptide sequences between known interacting protein pairs

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    Greenblatt Jack

    2006-07-01

    included in genome-wide yeast TAP tagging projects. Conclusion PIPE analysis can predict yeast protein-protein interactions. Also, PIPE analysis can be used to study the internal architecture of yeast protein complexes. The data also suggests that a finite set of short polypeptide signals seem to be responsible for the majority of the yeast protein-protein interactions.

  14. Peptides Interfering 3A Protein Dimerization Decrease FMDV Multiplication.

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    Mónica González-Magaldi

    Full Text Available Nonstructural protein 3A is involved in relevant functions in foot-and-mouth disease virus (FMDV replication. FMDV 3A can form homodimers and preservation of the two hydrophobic α-helices (α1 and α2 that stabilize the dimer interface is essential for virus replication. In this work, small peptides mimicking residues involved in the dimer interface were used to interfere with dimerization and thus gain insight on its biological function. The dimer interface peptides α1, α2 and that spanning the two hydrophobic α-helices, α12, impaired in vitro dimer formation of a peptide containing the two α-helices, this effect being higher with peptide α12. To assess the effect of dimer inhibition in cultured cells, the interfering peptides were N-terminally fused to a heptaarginine (R7 sequence to favor their intracellular translocation. Thus, when fused to R7, interference peptides (100 μM were able to inhibit dimerization of transiently expressed 3A, the higher inhibitions being found with peptides α1 and α12. The 3A dimerization impairment exerted by the peptides correlated with significant, specific reductions in the viral yield recovered from peptide-treated FMDV infected cells. In this case, α2 was the only peptide producing significant reductions at concentrations lower than 100 μM. Thus, dimer interface peptides constitute a tool to understand the structure-function relationship of this viral protein and point to 3A dimerization as a potential antiviral target.

  15. 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

  16. Constitutive expression of transgenes encoding derivatives of the synthetic antimicrobial peptide BP100: impact on rice host plant fitness

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    Nadal Anna

    2012-09-01

    Full Text Available Abstract Background The Biopeptide BP100 is a synthetic and strongly cationic α-helical undecapeptide with high, specific antibacterial activity against economically important plant-pathogenic bacteria, and very low toxicity. It was selected from a library of synthetic peptides, along with other peptides with activities against relevant bacterial and fungal species. Expression of the BP100 series of peptides in plants is of major interest to establish disease-resistant plants and facilitate molecular farming. Specific challenges were the small length, peptide degradation by plant proteases and toxicity to the host plant. Here we approached the expression of the BP100 peptide series in plants using BP100 as a proof-of-concept. Results Our design considered up to three tandemly arranged BP100 units and peptide accumulation in the endoplasmic reticulum (ER, analyzing five BP100 derivatives. The ER retention sequence did not reduce the antimicrobial activity of chemically synthesized BP100 derivatives, making this strategy possible. Transformation with sequences encoding BP100 derivatives (bp100der was over ten-fold less efficient than that of the hygromycin phosphotransferase (hptII transgene. The BP100 direct tandems did not show higher antimicrobial activity than BP100, and genetically modified (GM plants constitutively expressing them were not viable. In contrast, inverted repeats of BP100, whether or not elongated with a portion of a natural antimicrobial peptide (AMP, had higher antimicrobial activity, and fertile GM rice lines constitutively expressing bp100der were produced. These GM lines had increased resistance to the pathogens Dickeya chrysanthemi and Fusarium verticillioides, and tolerance to oxidative stress, with agronomic performance comparable to untransformed lines. Conclusions Constitutive expression of transgenes encoding short cationic α-helical synthetic peptides can have a strong negative impact on rice fitness. However, GM

  17. Design of a dual-function peptide probe as a binder of angiotensin II and an inducer of silver nanoparticle aggregation for use in label-free colorimetric assays.

    Science.gov (United States)

    Okochi, Mina; Kuboyama, Masashi; Tanaka, Masayoshi; Honda, Hiroyuki

    2015-09-01

    Label-free colorimetric assays using metallic nanoparticles have received much recent attention, for their application in simple and sensitive methods for detection of biomolecules. Short peptide probes that can bind to analyte biomolecules are attractive ligands in molecular nanotechnology; however, identification of biological recognition motifs is usually based on trial-and-error experiments. Herein, a peptide probe was screened for colorimetric detection of angiotensin II (Ang II) using a mechanism for non-crosslinking aggrega