WorldWideScience

Sample records for post-translational modification patterns

  1. Proteomic analysis of post-translational modifications

    DEFF Research Database (Denmark)

    Mann, Matthias; Jensen, Ole N

    2003-01-01

    Post-translational modifications modulate the activity of most eukaryote proteins. Analysis of these modifications presents formidable challenges but their determination generates indispensable insight into biological function. Strategies developed to characterize individual proteins are now...... systematically applied to protein populations. The combination of function- or structure-based purification of modified 'subproteomes', such as phosphorylated proteins or modified membrane proteins, with mass spectrometry is proving particularly successful. To map modification sites in molecular detail, novel...

  2. Post-Translational Modifications of TRP Channels

    Directory of Open Access Journals (Sweden)

    Olaf Voolstra

    2014-04-01

    Full Text Available Transient receptor potential (TRP channels constitute an ancient family of cation channels that have been found in many eukaryotic organisms from yeast to human. TRP channels exert a multitude of physiological functions ranging from Ca2+ homeostasis in the kidney to pain reception and vision. These channels are activated by a wide range of stimuli and undergo covalent post-translational modifications that affect and modulate their subcellular targeting, their biophysical properties, or channel gating. These modifications include N-linked glycosylation, protein phosphorylation, and covalent attachment of chemicals that reversibly bind to specific cysteine residues. The latter modification represents an unusual activation mechanism of ligand-gated ion channels that is in contrast to the lock-and-key paradigm of receptor activation by its agonists. In this review, we summarize the post-translational modifications identified on TRP channels and, when available, explain their physiological role.

  3. Post-translational modifications regulate signalling by Ror1

    Czech Academy of Sciences Publication Activity Database

    Kaucká, M.; Krejčí, Pavel; Plevová, K.; Pavlová, Š.; Procházková, Jiřina; Janovská, P.; Valnohová, J.; Kozubík, Alois; Pospíšilová, Š.; Bryja, Vítězslav

    2011-01-01

    Roč. 203, č. 3 (2011), s. 351-362 ISSN 1748-1708 Institutional support: RVO:68081707 Keywords : chronic lymphocytic leukaemia * glycosylation * post-translational modification Subject RIV: BO - Biophysics Impact factor: 3.090, year: 2011

  4. Top-down and Middle-down Protein Analysis Reveals that Intact and Clipped Human Histones Differ in Post-translational Modification Patterns

    DEFF Research Database (Denmark)

    Tvardovskiy, Andrey; Wrzesinski, Krzysztof; Sidoli, Simone

    2015-01-01

    Post-translational modifications (PTMs) of histone proteins play a fundamental role in regulation of DNA-templated processes. There is also growing evidence that proteolytic cleavage of histone N-terminal tails, known as histone clipping, influences nucleosome dynamics and functional properties...... hepatocytes and the hepatocellular carcinoma cell line HepG2/C3A when grown in spheroid (3D) culture, but not in a flat (2D) culture. Using tandem mass spectrometry we localized four different clipping sites in H3 and one clipping site in H2B. We show that in spheroid culture clipped H3 proteoforms are mainly...

  5. Cell signaling, post-translational protein modifications and NMR spectroscopy

    International Nuclear Information System (INIS)

    Theillet, Francois-Xavier; Smet-Nocca, Caroline; Liokatis, Stamatios; Thongwichian, Rossukon; Kosten, Jonas; Yoon, Mi-Kyung; Kriwacki, Richard W.; Landrieu, Isabelle; Lippens, Guy; Selenko, Philipp

    2012-01-01

    Post-translationally modified proteins make up the majority of the proteome and establish, to a large part, the impressive level of functional diversity in higher, multi-cellular organisms. Most eukaryotic post-translational protein modifications (PTMs) denote reversible, covalent additions of small chemical entities such as phosphate-, acyl-, alkyl- and glycosyl-groups onto selected subsets of modifiable amino acids. In turn, these modifications induce highly specific changes in the chemical environments of individual protein residues, which are readily detected by high-resolution NMR spectroscopy. In the following, we provide a concise compendium of NMR characteristics of the main types of eukaryotic PTMs: serine, threonine, tyrosine and histidine phosphorylation, lysine acetylation, lysine and arginine methylation, and serine, threonine O-glycosylation. We further delineate the previously uncharacterized NMR properties of lysine propionylation, butyrylation, succinylation, malonylation and crotonylation, which, altogether, define an initial reference frame for comprehensive PTM studies by high-resolution NMR spectroscopy.

  6. Targeting post-translational modifications of histones for cancer therapy.

    Science.gov (United States)

    Hsu, Y-C; Hsieh, Y-H; Liao, C-C; Chong, L-W; Lee, C-Y; Yu, Y-L; Chou, R-H

    2015-10-30

    Post-translational modifications (PTMs) on histones including acetylation, methylation, phosphorylation, citrullination, ubiquitination, ADP ribosylation, and sumoylation, play important roles in different biological events including chromatin dynamics, DNA replication, and transcriptional regulation. Aberrant histones PTMs leads to abnormal gene expression and uncontrolled cell proliferation, followed by development of cancers. Therefore, targeting the enzymes required for specific histone PTMs holds a lot of potential for cancer treatment. In this review article, we retrospect the latest studies in the regulations of acetylation, methylation, and phosphorylation of histones. We also summarize inhibitors/drugs that target these modifications for cancer treatment.

  7. The interplay of post-translational modification and gene therapy

    Directory of Open Access Journals (Sweden)

    Osamor VC

    2016-02-01

    Full Text Available Victor Chukwudi Osamor,1–3 Shalom N Chinedu,3,4 Dominic E Azuh,3,5 Emeka Joshua Iweala,3,4 Olubanke Olujoke Ogunlana3,4 1Covenant University Bioinformatics Research (CUBRe Unit, Department of Computer and Information Sciences, College of Science and Technology (CST, Covenant University, Ota, Ogun State, Nigeria; 2Institute of Informatics (Computational biology and Bioinformatics, Faculty of Mathematics, Informatics and Mechanics, University of Warsaw (Uniwersytet Warszawski, Warszawa, Poland; 3Covenant University Public Health and Well-being Research Group (CUPHWERG, Covenant University, 4Biochemistry and Molecular Biology Unit, Department of Biological Sciences, College of Science and Technology, Covenant University, Canaan Land, 5Department of Economics and Development Studies, Covenant University, Ota, Ogun State, Nigeria Abstract: Several proteins interact either to activate or repress the expression of other genes during transcription. Based on the impact of these activities, the proteins can be classified into readers, modifier writers, and modifier erasers depending on whether histone marks are read, added, or removed, respectively, from a specific amino acid. Transcription is controlled by dynamic epigenetic marks with serious health implications in certain complex diseases, whose understanding may be useful in gene therapy. This work highlights traditional and current advances in post-translational modifications with relevance to gene therapy delivery. We report that enhanced understanding of epigenetic machinery provides clues to functional implication of certain genes/gene products and may facilitate transition toward revision of our clinical treatment procedure with effective fortification of gene therapy delivery. Keywords: post-translational modification, gene therapy, epigenetics, histone, methylation

  8. Molecular dynamics simulation of phosphorylated KID post-translational modification.

    Directory of Open Access Journals (Sweden)

    Hai-Feng Chen

    2009-08-01

    Full Text Available Kinase-inducible domain (KID as transcriptional activator can stimulate target gene expression in signal transduction by associating with KID interacting domain (KIX. NMR spectra suggest that apo-KID is an unstructured protein. After post-translational modification by phosphorylation, KID undergoes a transition from disordered to well folded protein upon binding to KIX. However, the mechanism of folding coupled to binding is poorly understood.To get an insight into the mechanism, we have performed ten trajectories of explicit-solvent molecular dynamics (MD for both bound and apo phosphorylated KID (pKID. Ten MD simulations are sufficient to capture the average properties in the protein folding and unfolding.Room-temperature MD simulations suggest that pKID becomes more rigid and stable upon the KIX-binding. Kinetic analysis of high-temperature MD simulations shows that bound pKID and apo-pKID unfold via a three-state and a two-state process, respectively. Both kinetics and free energy landscape analyses indicate that bound pKID folds in the order of KIX access, initiation of pKID tertiary folding, folding of helix alpha(B, folding of helix alpha(A, completion of pKID tertiary folding, and finalization of pKID-KIX binding. Our data show that the folding pathways of apo-pKID are different from the bound state: the foldings of helices alpha(A and alpha(B are swapped. Here we also show that Asn139, Asp140 and Leu141 with large Phi-values are key residues in the folding of bound pKID. Our results are in good agreement with NMR experimental observations and provide significant insight into the general mechanisms of binding induced protein folding and other conformational adjustment in post-translational modification.

  9. Post-Translational Modifications of Histones in Human Sperm.

    Science.gov (United States)

    Krejčí, Jana; Stixová, Lenka; Pagáčová, Eva; Legartová, Soňa; Kozubek, Stanislav; Lochmanová, Gabriela; Zdráhal, Zbyněk; Sehnalová, Petra; Dabravolski, Siarhei; Hejátko, Jan; Bártová, Eva

    2015-10-01

    We examined the levels and distribution of post-translationally modified histones and protamines in human sperm. Using western blot immunoassay, immunofluorescence, mass spectrometry (MS), and FLIM-FRET approaches, we analyzed the status of histone modifications and the protamine P2. Among individual samples, we observed variability in the levels of H3K9me1, H3K9me2, H3K27me3, H3K36me3, and H3K79me1, but the level of acetylated (ac) histones H4 was relatively stable in the sperm head fractions, as demonstrated by western blot analysis. Sperm heads with lower levels of P2 exhibited lower levels of H3K9ac, H3K9me1, H3K27me3, H3K36me3, and H3K79me1. A very strong correlation was observed between the levels of P2 and H3K9me2. FLIM-FRET analysis additionally revealed that acetylated histones H4 are not only parts of sperm chromatin but also appear in a non-integrated form. Intriguingly, H4ac and H3K27me3 were detected in sperm tail fractions via western blot analysis. An appearance of specific histone H3 and H4 acetylation and H3 methylation in sperm tail fractions was also confirmed by both LC-MS/MS and MALDI-TOF MS analysis. Taken together, these data indicate that particular post-translational modifications of histones are uniquely distributed in human sperm, and this distribution varies among individuals and among the sperm of a single individual. © 2015 Wiley Periodicals, Inc.

  10. Tyrosine Sulfation as a Protein Post-Translational Modification

    Directory of Open Access Journals (Sweden)

    Yuh-Shyong Yang

    2015-01-01

    Full Text Available Integration of inorganic sulfate into biological molecules plays an important role in biological systems and is directly involved in the instigation of diseases. Protein tyrosine sulfation (PTS is a common post-translational modification that was first reported in the literature fifty years ago. However, the significance of PTS under physiological conditions and its link to diseases have just begun to be appreciated in recent years. PTS is catalyzed by tyrosylprotein sulfotransferase (TPST through transfer of an activated sulfate from 3'-phosphoadenosine-5'-phosphosulfate to tyrosine in a variety of proteins and peptides. Currently, only a small fraction of sulfated proteins is known and the understanding of the biological sulfation mechanisms is still in progress. In this review, we give an introductory and selective brief review of PTS and then summarize the basic biochemical information including the activity and the preparation of TPST, methods for the determination of PTS, and kinetics and reaction mechanism of TPST. This information is fundamental for the further exploration of the function of PTS that induces protein-protein interactions and the subsequent biochemical and physiological reactions.

  11. Proteomic analysis of post translational modifications in cyanobacteria.

    Science.gov (United States)

    Xiong, Qian; Chen, Zhuo; Ge, Feng

    2016-02-16

    Cyanobacteria are a diverse group of Gram-negative bacteria and the only prokaryotes capable of oxygenic photosynthesis. Recently, cyanobacteria have attracted great interest due to their crucial roles in global carbon and nitrogen cycles and their ability to produce clean and renewable biofuels. To survive in various environmental conditions, cyanobacteria have developed a complex signal transduction network to sense environmental signals and implement adaptive changes. The post-translational modifications (PTMs) systems play important regulatory roles in the signaling networks of cyanobacteria. The systematic investigation of PTMs could contribute to the comprehensive description of protein species and to elucidate potential biological roles of each protein species in cyanobacteria. Although the proteomic studies of PTMs carried out in cyanobacteria were limited, these data have provided clues to elucidate their sophisticated sensing mechanisms that contribute to their evolutionary and ecological success. This review aims to summarize the current status of PTM studies and recent publications regarding PTM proteomics in cyanobacteria, and discuss the novel developments and applications for the analysis of PTMs in cyanobacteria. Challenges, opportunities and future perspectives in the proteomics studies of PTMs in cyanobacteria are also discussed. Copyright © 2015 Elsevier B.V. All rights reserved.

  12. Regulation of the tumor suppressor PML by sequential post-translational modifications

    International Nuclear Information System (INIS)

    Schmitz, M. Lienhard; Grishina, Inna

    2012-01-01

    Post-translational modifications (PTMs) regulate multiple biological functions of the promyelocytic leukemia (PML) protein and also the fission, disassembly, and rebuilding of PML nuclear bodies (PML-NBs) during the cell cycle. Pathway-specific PML modification patterns ensure proper signal output from PML-NBs that suit the specific functional requirements. Here we comprehensively review the signaling pathways and enzymes that modify PML and also the oncogenic PML-RARα fusion protein. Many PTMs occur in a hierarchical and timely organized fashion. Phosphorylation or acetylation constitutes typical starting points for many PML modifying events, while degradative ubiquitination is an irreversible end point of the modification cascade. As this hierarchical organization of PTMs frequently turns phosphorylation events as primordial events, kinases or phosphatases regulating PML phosphorylation may be interesting drug targets to manipulate the downstream modifications and thus the stability and function of PML or PML-RARα.

  13. Muc1 based breast cancer vaccines: role of post translational modifications

    International Nuclear Information System (INIS)

    Begum, M.; Khurshid, R.; Nagra, S.A.

    2008-01-01

    Vaccine development is one of the most promising fields in cancer research. After autologous transplantation, due to low tumour burden, patients are more likely to respond immunologically to a cancer vaccine. MUC1 with its adhesive and anti adhesive functions, immunostimulatory and immunosuppressive activities, is therefore a good candidate for breast cancer vaccine. A structure-based insight into the immunogenicity of natural MUC1 glyco forms, of its sub-domains, motifs and post translational modification like glycosylation and myriostoylation may aid the design of tumour vaccines. Primary sequences of human MUC1 were retrieved from the SWISSPROT data bank. Protein pattern search: The primary sequence of Human MUC1 was searched at PROSITE (a dictionary of protein sites and patterns) database. Our study observes that post-translational modifications play an important role in presenting MUC1 as a candidate for breast cancer vaccine. It is found that the phosphorylation and glycosylation of important functional motifs of MUC1 may take part in the production of cytokines that may provide immunization. (author)

  14. Modification-specific proteomics: strategies for characterization of post-translational modifications using enrichment techniques

    DEFF Research Database (Denmark)

    Zhao, Yingming; Jensen, Ole N

    2009-01-01

    More than 300 different types of protein post-translational modifications (PTMs) have been described, many of which are known to have pivotal roles in cellular physiology and disease. Nevertheless, only a handful of PTMs have been extensively investigated at the proteome level. Knowledge of protein...... substrates and their PTM sites is key to dissection of PTM-mediated cellular processes. The past several years have seen a tremendous progress in developing MS-based proteomics technologies for global PTM analysis, including numerous studies of yeast and other microbes. Modification-specific enrichment...

  15. Spatial and Temporal Effects in Protein Post-translational Modification Distributions in the Developing Mouse Brain

    DEFF Research Database (Denmark)

    Edwards, Alistair V G; Edwards, Gregory J; Schwämmle, Veit

    2014-01-01

    Protein post-translational modification (PTM) is a powerful way to modify the behavior of cellular proteins and thereby cellular behavior. Multiple recent studies of evolutionary trends have shown that certain pairs of protein post-translational modifications tend to occur closer to each other than...... for observations of increasingly frequent and diverse protein modification in cell biology. In this study, we use mass spectrometry and proteomic strategies to present biological data showing spatiotemporal PTM co-localization across multiple PTM categories, which display changes over development of the brain...

  16. Alpha1-acid glycoprotein post-translational modifications: a comparative two dimensional electrophoresis based analysis

    Directory of Open Access Journals (Sweden)

    P. Roncada

    2010-04-01

    Full Text Available Alpha1-acid glycoprotein (AGP is an immunomodulatory protein expressed by hepatocytes in response to the systemic reaction that follows tissue damage caused by inflammation, infection or trauma. A proteomic approach based on two dimensional electrophoresis, immunoblotting and staining of 2DE gels with dyes specific for post-translational modifications (PTMs such as glycosylation and phosphorylation has been used to evaluate the differential interspecific protein expression of AGP purified from human, bovine and ovine sera. By means of these techniques, several isoforms have been identified in the investigated species: they have been found to change both with regard to the number of isoforms expressed under physiological condition and with regard to the quality of PTMs (i.e. different oligosaccharidic chains, presence/absence of phosphorilations. In particular, it is suggested that bovine serum AGP may have one of the most complex pattern of PTMs among serum proteins of mammals studied so far.

  17. Variable elimination in post-translational modification reaction networks with mass-action kinetics

    DEFF Research Database (Denmark)

    Feliu, Elisenda; Wiuf, Carsten

    2013-01-01

    We define a subclass of chemical reaction networks called post-translational modification systems. Important biological examples of such systems include MAPK cascades and two-component systems which are well-studied experimentally as well as theoretically. The steady states of such a system...

  18. Cysteine S-glycosylation, a new post-translational modification found in glycopeptide bacteriocins

    Czech Academy of Sciences Publication Activity Database

    Stepper, J.; Shastri, S.; Loo, T. S.; Preston, J. C.; Novák, Petr; Man, Petr; Moore, Ch. H.; Havlíček, Vladimír; Patchett, M. L.; Norris, G. E.

    2011-01-01

    Roč. 585, č. 4 (2011), s. 645-650 ISSN 0014-5793 Institutional research plan: CEZ:AV0Z50200510 Keywords : Post-translational modification * Glycosylation * Bacteriocin Subject RIV: CE - Biochemistry Impact factor: 3.538, year: 2011

  19. Elucidating Host-Pathogen Interactions Based on Post-Translational Modifications Using Proteomics Approaches

    DEFF Research Database (Denmark)

    Ravikumar, Vaishnavi; Jers, Carsten; Mijakovic, Ivan

    2015-01-01

    can be efficiently applied to gain an insight into the molecular mechanisms involved. The measurement of the proteome and post-translationally modified proteome dynamics using mass spectrometry, results in a wide array of information, such as significant changes in protein expression, protein...... display host specificity through a complex network of molecular interactions that aid their survival and propagation. Co-infection states further lead to complications by increasing the microbial burden and risk factors. Quantitative proteomics based approaches and post-translational modification analysis...... pathogen interactions....

  20. Profiling of integral membrane proteins and their post translational modifications using high-resolution mass spectrometry

    Science.gov (United States)

    Souda, Puneet; Ryan, Christopher M.; Cramer, William A.; Whitelegge, Julian

    2011-01-01

    Integral membrane proteins pose challenges to traditional proteomics approaches due to unique physicochemical properties including hydrophobic transmembrane domains that limit solubility in aqueous solvents. A well resolved intact protein molecular mass profile defines a protein’s native covalent state including post-translational modifications, and is thus a vital measurement toward full structure determination. Both soluble loop regions and transmembrane regions potentially contain post-translational modifications that must be characterized if the covalent primary structure of a membrane protein is to be defined. This goal has been achieved using electrospray-ionization mass spectrometry (ESI-MS) with low-resolution mass analyzers for intact protein profiling, and high-resolution instruments for top-down experiments, toward complete covalent primary structure information. In top-down, the intact protein profile is supplemented by gas-phase fragmentation of the intact protein, including its transmembrane regions, using collisionally activated and/or electroncapture dissociation (CAD/ECD) to yield sequence-dependent high-resolution MS information. Dedicated liquid chromatography systems with aqueous/organic solvent mixtures were developed allowing us to demonstrate that polytopic integral membrane proteins are amenable to ESI-MS analysis, including top-down measurements. Covalent post-translational modifications are localized regardless of their position in transmembrane domains. Top-down measurements provide a more detail oriented high-resolution description of post-transcriptional and post-translational diversity for enhanced understanding beyond genomic translation. PMID:21982782

  1. Lysine-Directed Post-translational Modifications of Tau Protein in Alzheimer's Disease and Related Tauopathies

    Directory of Open Access Journals (Sweden)

    Christiana Kontaxi

    2017-08-01

    Full Text Available Tau is a microtubule-associated protein responsible mainly for stabilizing the neuronal microtubule network in the brain. Under normal conditions, tau is highly soluble and adopts an “unfolded” conformation. However, it undergoes conformational changes resulting in a less soluble form with weakened microtubule stabilizing properties. Altered tau forms characteristic pathogenic inclusions in Alzheimer's disease and related tauopathies. Although, tau hyperphosphorylation is widely considered to be the major trigger of tau malfunction, tau undergoes several post-translational modifications at lysine residues including acetylation, methylation, ubiquitylation, SUMOylation, and glycation. We are only beginning to define the site-specific impact of each type of lysine modification on tau biology as well as the possible interplay between them, but, like phosphorylation, these modifications are likely to play critical roles in tau's normal and pathobiology. This review summarizes the latest findings focusing on lysine post-translational modifications that occur at both endogenous tau protein and pathological tau forms in AD and other tauopathies. In addition, it highlights the significance of a site-dependent approach of studying tau post-translational modifications under normal and pathological conditions.

  2. Golgi structure formation, function, and post-translational modifications in mammalian cells.

    Science.gov (United States)

    Huang, Shijiao; Wang, Yanzhuang

    2017-01-01

    The Golgi apparatus is a central membrane organelle for trafficking and post-translational modifications of proteins and lipids in cells. In mammalian cells, it is organized in the form of stacks of tightly aligned flattened cisternae, and dozens of stacks are often linked laterally into a ribbon-like structure located in the perinuclear region of the cell. Proper Golgi functionality requires an intact architecture, yet Golgi structure is dynamically regulated during the cell cycle and under disease conditions. In this review, we summarize our current understanding of the relationship between Golgi structure formation, function, and regulation, with focus on how post-translational modifications including phosphorylation and ubiquitination regulate Golgi structure and on how Golgi unstacking affects its functions, in particular, protein trafficking, glycosylation, and sorting in mammalian cells.

  3. Post-Translational Modifications of RelB NF-κB Subunit and Associated Functions

    Directory of Open Access Journals (Sweden)

    Véronique Baud

    2016-05-01

    Full Text Available The family of NF-κB transcription factors plays a key role in diverse biological processes, such as inflammatory and immune responses, cell survival and tumor development. Beyond the classical NF-κB activation pathway, a second NF-κB pathway has more recently been uncovered, the so-called alternative NF-κB activation pathway. It has been shown that this pathway mainly controls the activity of RelB, a member of the NF-κB family. Post-translational modifications, such as phosphorylation, acetylation, methylation, ubiquitination and SUMOylation, have recently emerged as a strategy for the fine-tuned regulation of NF-κB. Our review discusses recent progress in the understanding of RelB regulation by post-translational modifications and the associated functions in normal and pathological conditions.

  4. Managing the complexity of communication: regulation of gap junctions by post-translational modification

    DEFF Research Database (Denmark)

    Axelsen, Lene Nygaard; Callø, Kirstine; von Holstein-Rathlou, Niels-Henrik

    2013-01-01

    Gap junctions are comprised of connexins that form cell-to-cell channels which couple neighboring cells to accommodate the exchange of information. The need for communication does, however, change over time and therefore must be tightly controlled. Although the regulation of connexin protein...... probability, single channel conductance or selectivity. The most extensively investigated post translational modifications are phosphorylations, which have been documented in all mammalian connexins. Besides phosphorylations, some connexins are known to be ubiquitinated, SUMOylated, nitrosylated, hydroxylated...

  5. Computational and statistical methods for high-throughput analysis of post-translational modifications of proteins

    DEFF Research Database (Denmark)

    Schwämmle, Veit; Braga, Thiago Verano; Roepstorff, Peter

    2015-01-01

    The investigation of post-translational modifications (PTMs) represents one of the main research focuses for the study of protein function and cell signaling. Mass spectrometry instrumentation with increasing sensitivity improved protocols for PTM enrichment and recently established pipelines...... for high-throughput experiments allow large-scale identification and quantification of several PTM types. This review addresses the concurrently emerging challenges for the computational analysis of the resulting data and presents PTM-centered approaches for spectra identification, statistical analysis...

  6. Rho GTPases, their post-translational modifications, disease-associated mutations and pharmacological inhibitors.

    Science.gov (United States)

    Olson, Michael F

    2018-05-04

    The 20 members of the Rho GTPase family are key regulators of a wide-variety of biological activities. In response to activation, they signal via downstream effector proteins to induce dynamic alterations in the organization of the actomyosin cytoskeleton. In this review, post-translational modifications, mechanisms of dysregulation identified in human pathological conditions, and the ways that Rho GTPases might be targeted for chemotherapy will be discussed.

  7. Hunting for unexpected post-translational modifications by spectral library searching with tier-wise scoring.

    Science.gov (United States)

    Ma, Chun Wai Manson; Lam, Henry

    2014-05-02

    Discovering novel post-translational modifications (PTMs) to proteins and detecting specific modification sites on proteins is one of the last frontiers of proteomics. At present, hunting for post-translational modifications remains challenging in widely practiced shotgun proteomics workflows due to the typically low abundance of modified peptides and the greatly inflated search space as more potential mass shifts are considered by the search engines. Moreover, most popular search methods require that the user specifies the modification(s) for which to search; therefore, unexpected and novel PTMs will not be detected. Here a new algorithm is proposed to apply spectral library searching to the problem of open modification searches, namely, hunting for PTMs without prior knowledge of what PTMs are in the sample. The proposed tier-wise scoring method intelligently looks for unexpected PTMs by allowing mass-shifted peak matches but only when the number of matches found is deemed statistically significant. This allows the search engine to search for unexpected modifications while maintaining its ability to identify unmodified peptides effectively at the same time. The utility of the method is demonstrated using three different data sets, in which the numbers of spectrum identifications to both unmodified and modified peptides were substantially increased relative to a regular spectral library search as well as to another open modification spectral search method, pMatch.

  8. Tubulin post-translational modifications in the primitive protist Trichomonas vaginalis.

    Science.gov (United States)

    Delgado-Viscogliosi, P; Brugerolle, G; Viscogliosi, E

    1996-01-01

    Using several specific monoclonal antibodies, we investigated the occurrence and distribution of different post-translationally modified tubulin during interphase and division of the primitive flagellated protist Trichomonas vaginalis. Immunoblotting and immunofluorescence experiments revealed that interphasic microtubular structures of T. vaginalis contained acetylated and glutamylated but non-tyrosinated and non-glycylated [Brugerolle and Adoutte, 1988: Bio Systems 21: 255-268] tubulin. Immunofluorescence studies performed on dividing cells showed that the extranuclear mitotic spindle (or paradesmosis) was acetylated and glutamylated, which contrast with the ephemeral nature of this structure. Newly formed short axostyles also contained acetylated and glutamylated tubulin suggesting that both post-translational modifications might take place very early after assembly of microtubular structures. Our results indicate that acetylation and glutamylation of tubulin appeared early in the history of eukaryotes and could reflect the occurrence of post-translational modifications of tubulin in the primitive eukaryotic cells. These cells probably had a highly ordered cross-linked microtubular cytoskeleton in which microtubules showed a low level of subunit exchange dynamics.

  9. Profiling Changes in Histone Post-translational Modifications by Top-Down Mass Spectrometry

    Energy Technology Data Exchange (ETDEWEB)

    Zhou, Mowei; Wu, Si; Stenoien, David L.; Zhang, Zhaorui; Connolly, Lanelle; Freitag, Michael; Pasa-Tolic, Ljiljana

    2016-11-11

    Top-down mass spectrometry is a valuable tool for charactering post-translational modifications on histones for understanding of gene control and expression. In this protocol, we describe a top-down workflow using liquid chromatography coupled to mass spectrometry for fast global profiling of changes in histone proteoforms between a wild-type and a mutant of a fungal species. The proteoforms exhibiting different abundances can be subjected to further targeted studies by other mass spectrometric or biochemical assays. This method can be generally adapted for preliminary screening for changes in histone modifications between samples such as wild-type vs. mutant, and control vs. disease.

  10. Plant cytoplasmic GAPDH: redox post-translational modifications and moonlighting properties

    Directory of Open Access Journals (Sweden)

    Mirko eZaffagnini

    2013-11-01

    Full Text Available Glyceraldehyde-3-phosphate dehydrogenase (GAPDH is a ubiquitous enzyme involved in glycolysis and shown, particularly in animal cells, to play additional roles in several unrelated non-metabolic processes such as control of gene expression and apoptosis. This functional versatility is regulated, in part at least, by redox post-translational modifications that alter GAPDH catalytic activity and influence the subcellular localization of the enzyme. In spite of the well established moonlighting (multifunctional properties of animal GAPDH, little is known about non-metabolic roles of GAPDH in plants. Plant cells contain several GAPDH isoforms with different catalytic and regulatory properties, located both in the cytoplasm and in plastids, and participating in glycolysis and the Calvin-Benson cycle. A general feature of all GAPDH proteins is the presence of an acidic catalytic cysteine in the active site that is overly sensitive to oxidative modifications, including glutathionylation and S-nitrosylation. In Arabidopsis, oxidatively-modified cytoplasmic GAPDH has been successfully used as a tool to investigate the role of reduced glutathione, thioredoxins and glutaredoxins in the control of different types of redox post-translational modifications. Oxidative modifications inhibit GAPDH activity, but might enable additional functions in plant cells. Mounting evidence support the concept that plant cytoplasmic GAPDH may fulfill alternative, non-metabolic functions that are triggered by redox post-translational modifications of the protein under stress conditions. The aim of this review is to detail the molecular mechanisms underlying the redox regulation of plant cytoplasmic GAPDH in the light of its crystal structure, and to provide a brief inventory of the well known redox-dependent multi-facetted properties of animal GAPDH, together with the emerging roles of oxidatively-modified GAPDH in stress signaling pathways in plants.

  11. Comparative proteomic analysis of histone post-translational modifications upon ischemia/reperfusion-induced retinal injury

    DEFF Research Database (Denmark)

    Zhao, Xiaolu; Sidoli, Simone; Wang, Leilei

    2014-01-01

    We present a detailed quantitative map of single and coexisting histone post-translational modifications (PTMs) in rat retinas affected by ischemia and reperfusion (I/R) injury. Retinal I/R injury contributes to serious ocular diseases, which can lead to vision loss and blindness. We applied linear...... ion trap-orbitrap hybrid tandem mass spectrometry (MS/MS) to quantify 131 single histone marks and 143 combinations of multiple histone marks in noninjured and injured retinas. We observed 34 histone PTMs that exhibited significantly (p

  12. Prediction of human protein function from post-translational modifications and localization features

    DEFF Research Database (Denmark)

    Jensen, Lars Juhl; Gupta, Ramneek; Blom, Nikolaj

    2002-01-01

    a number of functional attributes that are more directly related to the linear sequence of amino acids, and hence easier to predict, than protein structure. These attributes include features associated with post-translational modifications and protein sorting, but also much simpler aspects......We have developed an entirely sequence-based method that identifies and integrates relevant features that can be used to assign proteins of unknown function to functional classes, and enzyme categories for enzymes. We show that strategies for the elucidation of protein function may benefit from...

  13. Proteomic profiling and post-translational modifications in human keratinocytes treated with Mucuna pruriens leaf extract.

    Science.gov (United States)

    Cortelazzo, Alessio; Lampariello, Raffaella L; Sticozzi, Claudia; Guerranti, Roberto; Mirasole, Cristiana; Zolla, Lello; Sacchetti, Gianni; Hajek, Joussef; Valacchi, Giuseppe

    2014-02-03

    Mucuna pruriens (Mp) is a plant belonging to the Fabaceae family, with several medicinal properties among which its potential to treat diseases where reactive oxygen species (ROS) play an important role in the pathogeneses. The aim was to investigate the effects of Mp leaf methanolic extract (MPME) on human keratinocytes protein expression and its role in preventing proteins oxidation after oxidative stress (OS) exposure. The effects of MPME on HaCaT cells protein expression were evaluated treating cells with different concentrations of MPME, with glucose oxidase (GO, source of OS) and with MPME subsequently treated with GO. The protein patterns of treated HaCaT cells are analyzed by two-dimensional gel electrophoresis (2-DE) and compared with that of untreated HaCaT. Immunoblotting was then used to evaluate the role of MPME in preventing the 4-hydroxynonenal protein adducts (4-HNE PAs) formation (marker of OS). Eighteen proteins, identified by mass spectrometry (LC-ESI-CID-MS/MS), were modulated distinctly by MPME in HaCaT. Overall, MPME counteract GO effect, reducing the GO-induced overexpression of several proteins involved in stress response (T-complex protein 1, Protein disulfide-isomerase A3, Protein DJ-1, and Stress-induced-phosphoprotein 1), in cell energy methabolism (Inorganic pyrophosphatase, Triosephosphate isomerase isoform 1, 2-phosphopyruvate-hydratase alpha-enolase, and Fructose-bisphosphate aldolase A isoform 1), in cytoskeletal organization (Cytokeratins 18, 9, 2, Cofilin-1, Annexin A2 and F-actin-capping protein subunit beta isoform 1) and in cell cycle progression (Eukaryotic translation initiation factor 5A-1 isoform B). In addition, MPME decreased the 4-HNE PAs levels, in particular on 2-phosphopyruvate-hydratase alpha-enolase and Cytokeratin 9. Our findings show that MPME might be helpful in the treatment of OS-related skin diseases by preventing protein post-translational modifications (4-HNE PAs). © 2013 Published by Elsevier Ireland Ltd.

  14. Post-Translational Modifications of Desulfovibrio vulgaris Hildenborough Sulfate Reduction Pathway Proteins

    Energy Technology Data Exchange (ETDEWEB)

    Gaucher, S.P.; Redding, A.M.; Mukhopadhyay, A.; Keasling, J.D.; Singh, A.K.

    2008-03-01

    Recent developments in shotgun proteomics have enabled high-throughput studies of a variety of microorganisms at a proteome level and provide experimental validation for predicted open reading frames in the corresponding genome. More importantly, advances in mass spectrometric data analysis now allow mining of large proteomics data sets for the presence of post-translational modifications(PTMs). Although PTMs are a critical aspectof cellular activity, such information eludes cell-wide studies conducted at the transcript level. Here, we analyze several mass spectrometric data sets acquired using two-dimensional liquid chromatography tandem mass spectrometry, 2D-LC/MS/MS, for the sulfate reducing bacterium, Desulfovibrio vulgaris Hildenborough. Our searches of the raw spectra led us to discover several post-translationally modified peptides in D. vulgaris. Of these, several peptides containing a lysine with a +42 Da modification were found reproducibly across all data sets. Both acetylation and trimethylation have the same nominal +42 Da mass, and are therefore candidates for this modification. Several spectra were identified having markers for trimethylation, while one is consistent with an acetylation. Surprisingly, these modified peptides predominantly mapped to proteins involved in sulfate respiration. Other highly expressed proteins in D. vulgaris, such as enzymes involved in electron transport and other central metabolic processes, did not contain this modification. Decoy database searches were used to control for random spectrum/sequence matches. Additional validation for these modifications was provided by alternate workflows, for example, two-dimensional gel electrophoresis followed by mass spectrometry analysis of the dissimilatory sulfite reductase gamma-subunit(DsrC) protein. MS data for DsrC in this alternate workflow also contained the +42 Da modification at the same loci. Furthermore, the DsrC homologue in another sulfate reducing bacterium

  15. A novel post-translational modification in nerve terminals: O-linked N-acetylglucosamine phosphorylation

    DEFF Research Database (Denmark)

    Graham, Mark E; Thaysen-Andersen, Morten; Bache, Nicolai

    2011-01-01

    Protein phosphorylation and glycosylation are the most common post-translational modifications observed in biology, frequently on the same protein. Assembly protein AP180 is a synapse-specific phosphoprotein and O-linked beta-N-acetylglucosamine (O-GlcNAc) modified glycoprotein. AP180 is involved......NAc-P to a Thr residue was confirmed by electron transfer dissociation MS. A second AP180 tryptic peptide was also glycosyl phosphorylated, but the site of modification was not assigned. Sequence similarities suggest there may be a common motif within AP180 involving glycosyl phosphorylation and dual flanking...... phosphorylation sites within 4 amino acid residues. This novel type of protein glycosyl phosphorylation adds a new signaling mechanism to the regulation of neurotransmission and more complexity to the study of O-GlcNAc modification....

  16. Status of large-scale analysis of post-translational modifications by mass spectrometry

    DEFF Research Database (Denmark)

    Olsen, Jesper V; Mann, Matthias

    2013-01-01

    Cellular function can be controlled through the gene expression program but often protein post translations modifications (PTMs) provide a more precisely and elegant mechanism. Key functional roles of specific modification events for instance during the cell cycle have been known for decades...... of protein modifications. For many PTMs, including phosphorylation, ubiquitination, glycosylation and acetylation, tens of thousands of sites can now be confidently identified and localized in the sequence of the protein. Quantitation of PTM levels between different cellular states is likewise established......, with label-free methods showing particular promise. It is also becoming possible to determine the absolute occupancy or stoichiometry of PTMS sites on a large scale. Powerful software for the bioinformatic analysis of thousands of PTM sites has been developed. However, a complete inventory of sites has...

  17. Identification of novel post-translational modifications in linker histones from chicken erythrocytes.

    Science.gov (United States)

    Sarg, Bettina; Lopez, Rita; Lindner, Herbert; Ponte, Inma; Suau, Pedro; Roque, Alicia

    2015-01-15

    Chicken erythrocyte nuclei were digested with micrococcal nuclease and fractionated by centrifugation in low-salt buffer into soluble and insoluble fractions. Post-translational modifications of the purified linker histones of both fractions were analyzed by LC-ESI-MS/MS. All six histone H1 subtypes (H1.01, H1.02, H1.03, H1.10, H1.1L and H1.1R) and histone H5 were identified. Mass spectrometry analysis enabled the identification of a wide range of PTMs, including N(α)-terminal acetylation, acetylation, formylation, phosphorylation and oxidation. A total of nine new modifications in chicken linker histones were mapped, most of them located in the N-terminal and globular domains. Relative quantification of the modified peptides showed that linker histone PTMs were differentially distributed among both chromatin fractions, suggesting their relevance in the regulation of chromatin structure. The analysis of our results combined with previously reported data for chicken and some mammalian species showed that most of the modified positions were conserved throughout evolution, highlighting their importance in specific linker histone functions and epigenetics. Post-translational modifications of linker histones could have a role in the regulation of gene expression through the modulation of chromatin higher-order structure and chromatin remodeling. Finding new PTMs in linker histones is the first step to elucidate their role in the histone code. In this manuscript we report nine new post-translational modifications of the linker histones from chicken erythrocytes, one in H5 and eight in the H1 subtypes. Chromatin fractionated by centrifugation in low-salt buffer resulted in two fractions with different contents and compositions of linker histones and enriched in specific core histone PTMs. Of particular interest is the fact that linker histone PTMs were differentially distributed in both chromatin fractions, suggesting specific functions. Future studies are needed to

  18. AMS 4.0: consensus prediction of post-translational modifications in protein sequences.

    Science.gov (United States)

    Plewczynski, Dariusz; Basu, Subhadip; Saha, Indrajit

    2012-08-01

    We present here the 2011 update of the AutoMotif Service (AMS 4.0) that predicts the wide selection of 88 different types of the single amino acid post-translational modifications (PTM) in protein sequences. The selection of experimentally confirmed modifications is acquired from the latest UniProt and Phospho.ELM databases for training. The sequence vicinity of each modified residue is represented using amino acids physico-chemical features encoded using high quality indices (HQI) obtaining by automatic clustering of known indices extracted from AAindex database. For each type of the numerical representation, the method builds the ensemble of Multi-Layer Perceptron (MLP) pattern classifiers, each optimising different objectives during the training (for example the recall, precision or area under the ROC curve (AUC)). The consensus is built using brainstorming technology, which combines multi-objective instances of machine learning algorithm, and the data fusion of different training objects representations, in order to boost the overall prediction accuracy of conserved short sequence motifs. The performance of AMS 4.0 is compared with the accuracy of previous versions, which were constructed using single machine learning methods (artificial neural networks, support vector machine). Our software improves the average AUC score of the earlier version by close to 7 % as calculated on the test datasets of all 88 PTM types. Moreover, for the selected most-difficult sequence motifs types it is able to improve the prediction performance by almost 32 %, when compared with previously used single machine learning methods. Summarising, the brainstorming consensus meta-learning methodology on the average boosts the AUC score up to around 89 %, averaged over all 88 PTM types. Detailed results for single machine learning methods and the consensus methodology are also provided, together with the comparison to previously published methods and state-of-the-art software tools. The

  19. Mass-spectrometry analysis of histone post-translational modifications in pathology tissue using the PAT-H-MS approach

    Directory of Open Access Journals (Sweden)

    Roberta Noberini

    2016-06-01

    Full Text Available Aberrant histone post-translational modifications (hPTMs have been implicated with various pathologies, including cancer, and may represent useful epigenetic biomarkers. The data described here provide a mass spectrometry-based quantitative analysis of hPTMs from formalin-fixed paraffin-embedded (FFPE tissues, from which histones were extracted through the recently developed PAT-H-MS method. First, we analyzed FFPE samples from mouse spleen and liver or human breast cancer up to six years old, together with their corresponding fresh frozen tissue. We then combined the PAT-H-MS approach with a histone-focused version of the super-SILAC strategy-using a mix of histones from four breast cancer cell lines as a spike-in standard- to accurately quantify hPTMs from breast cancer specimens belonging to different subtypes. The data, which are associated with a recent publication (Pathology tissue-quantitative mass spectrometry analysis to profile histone post-translational modification patterns in patient samples (Noberini, 2015 [1], are deposited at the ProteomeXchange Consortium via the PRIDE partner repository with the dataset identifier http://www.ebi.ac.uk/pride/archive/projects/PXD002669.

  20. Prediction of protein post-translational modifications: main trends and methods

    Science.gov (United States)

    Sobolev, B. N.; Veselovsky, A. V.; Poroikov, V. V.

    2014-02-01

    The review summarizes main trends in the development of methods for the prediction of protein post-translational modifications (PTMs) by considering the three most common types of PTMs — phosphorylation, acetylation and glycosylation. Considerable attention is given to general characteristics of regulatory interactions associated with PTMs. Different approaches to the prediction of PTMs are analyzed. Most of the methods are based only on the analysis of the neighbouring environment of modification sites. The related software is characterized by relatively low accuracy of PTM predictions, which may be due both to the incompleteness of training data and the features of PTM regulation. Advantages and limitations of the phylogenetic approach are considered. The prediction of PTMs using data on regulatory interactions, including the modular organization of interacting proteins, is a promising field, provided that a more carefully selected training data will be used. The bibliography includes 145 references.

  1. A homology-based pipeline for global prediction of post-translational modification sites

    Science.gov (United States)

    Chen, Xiang; Shi, Shao-Ping; Xu, Hao-Dong; Suo, Sheng-Bao; Qiu, Jian-Ding

    2016-05-01

    The pathways of protein post-translational modifications (PTMs) have been shown to play particularly important roles for almost any biological process. Identification of PTM substrates along with information on the exact sites is fundamental for fully understanding or controlling biological processes. Alternative computational strategies would help to annotate PTMs in a high-throughput manner. Traditional algorithms are suited for identifying the common organisms and tissues that have a complete PTM atlas or extensive experimental data. While annotation of rare PTMs in most organisms is a clear challenge. In this work, to this end we have developed a novel homology-based pipeline named PTMProber that allows identification of potential modification sites for most of the proteomes lacking PTMs data. Cross-promotion E-value (CPE) as stringent benchmark has been used in our pipeline to evaluate homology to known modification sites. Independent-validation tests show that PTMProber achieves over 58.8% recall with high precision by CPE benchmark. Comparisons with other machine-learning tools show that PTMProber pipeline performs better on general predictions. In addition, we developed a web-based tool to integrate this pipeline at http://bioinfo.ncu.edu.cn/PTMProber/index.aspx. In addition to pre-constructed prediction models of PTM, the website provides an extensional functionality to allow users to customize models.

  2. Protein redox chemistry: post-translational cysteine modifications that regulate signal transduction and drug pharmacology

    Directory of Open Access Journals (Sweden)

    Revati eWani

    2014-10-01

    Full Text Available The perception of reactive oxygen species (ROS has evolved over the past decade from agents of cellular damage to secondary messengers which modify signaling proteins in physiology and the disease state (e.g. cancer. New protein targets of specific oxidation are rapidly being identified. One emerging class of redox modification occurs to the thiol side chain of cysteine residues which can produce multiple chemically-distinct alterations to the protein (e.g. sulfenic/sulfinic/sulfonic acid, disulfides. These post-translational modifications (PTM are shown to affect the protein structure and function. Because redox-sensitive proteins can traffic between subcellular compartments that have different redox environments, cysteine oxidation enables a spatio-temporal control to signaling. Understanding ramifications of these oxidative modifications to the functions of signaling proteins is crucial for understanding cellular regulation as well as for informed-drug discovery process. The effects of EGFR oxidation of Cys797 on inhibitor pharmacology are presented to illustrate the principle. Taken together, cysteine redox PTM can impact both cell biology and drug pharmacology.

  3. Beyond gene expression: the impact of protein post-translational modifications in bacteria.

    Science.gov (United States)

    Cain, Joel A; Solis, Nestor; Cordwell, Stuart J

    2014-01-31

    The post-translational modification (PTM) of proteins plays a critical role in the regulation of a broad range of cellular processes in eukaryotes. Yet their role in governing similar systems in the conventionally presumed 'simpler' forms of life has been largely neglected and, until recently, was thought to occur only rarely, with some modifications assumed to be limited to higher organisms alone. Recent developments in mass spectrometry-based proteomics have provided an unparalleled power to enrich, identify and quantify peptides with PTMs. Additional modifications to biological molecules such as lipids and carbohydrates that are essential for bacterial pathophysiology have only recently been detected on proteins. Here we review bacterial protein PTMs, focusing on phosphorylation, acetylation, proteolytic degradation, methylation and lipidation and the roles they play in bacterial adaptation - thus highlighting the importance of proteomic techniques in a field that is only just in its infancy. This article is part of a Special Issue entitled: Trends in Microbial Proteomics. © 2013 Elsevier B.V. All rights reserved.

  4. Quantitative proteomic characterization of redox-dependent post-translational modifications on protein cysteines

    Energy Technology Data Exchange (ETDEWEB)

    Duan, Jicheng; Gaffrey, Matthew J.; Qian, Wei-Jun

    2017-01-01

    Protein cysteine thiols play a crucial role in redox signaling, regulation of enzymatic activity and protein function, and maintaining redox homeostasis in living systems. The unique chemical reactivity of thiol groups makes cysteine susceptible to oxidative modifications by reactive oxygen and nitrogen species to form a broad array of reversible and irreversible protein post-translational modifications (PTMs). The reversible modifications in particular are one of the major components of redox signaling and are involved in regulation of various cellular processes under physiological and pathological conditions. The biological significance of these redox PTMs in health and diseases has been increasingly recognized. Herein, we review the recent advances of quantitative proteomic approaches for investigating redox PTMs in complex biological systems, including the general considerations of sample processing, various chemical or affinity enrichment strategies, and quantitative approaches. We also highlight a number of redox proteomic approaches that enable effective profiling of redox PTMs for addressing specific biological questions. Although some technological limitations remain, redox proteomics is paving the way towards a better understanding of redox signaling and regulation in human health and diseases.

  5. Global turnover of histone post-translational modifications and variants in human cells

    Directory of Open Access Journals (Sweden)

    Zee Barry M

    2010-12-01

    Full Text Available Abstract Background Post-translational modifications (PTMs on the N-terminal tails of histones and histone variants regulate distinct transcriptional states and nuclear events. Whereas the functional effects of specific PTMs are the current subject of intense investigation, most studies characterize histone PTMs/variants in a non-temporal fashion and very few studies have reported kinetic information about these histone forms. Previous studies have used radiolabeling, fluorescence microscopy and chromatin immunoprecipitation to determine rates of histone turnover, and have found interesting correlations between increased turnover and increased gene expression. Therefore, histone turnover is an understudied yet potentially important parameter that may contribute to epigenetic regulation. Understanding turnover in the context of histone modifications and sequence variants could provide valuable additional insight into the function of histone replacement. Results In this study, we measured the metabolic rate of labeled isotope incorporation into the histone proteins of HeLa cells by combining stable isotope labeling of amino acids in cell culture (SILAC pulse experiments with quantitative mass spectrometry-based proteomics. In general, we found that most core histones have similar turnover rates, with the exception of the H2A variants, which exhibit a wider range of rates, potentially consistent with their epigenetic function. In addition, acetylated histones have a significantly faster turnover compared with general histone protein and methylated histones, although these rates vary considerably, depending on the site and overall degree of methylation. Histones containing transcriptionally active marks have been consistently found to have faster turnover rates than histones containing silent marks. Interestingly, the presence of both active and silent marks on the same peptide resulted in a slower turnover rate than either mark alone on that same

  6. Identification and characterization of HTLV-1 HBZ post-translational modifications.

    Directory of Open Access Journals (Sweden)

    Nathan Dissinger

    Full Text Available Human T-cell leukemia virus type-1 (HTLV-1 is estimated to infect 15-25 million people worldwide, with several areas including southern Japan and the Caribbean basin being endemic. The virus is the etiological agent of debilitating and fatal diseases, for which there is currently no long-term cure. In the majority of cases of leukemia caused by HTLV-1, only a single viral gene, hbz, and its cognate protein, HBZ, are expressed and their importance is increasingly being recognized in the development of HTLV-1-associated disease. We hypothesized that HBZ, like other HTLV-1 proteins, has properties and functions regulated by post-translational modifications (PTMs that affect specific signaling pathways important for disease development. To date, PTM of HBZ has not been described. We used an affinity-tagged protein and mass spectrometry method to identify seven modifications of HBZ for the first time. We examined how these PTMs affected the ability of HBZ to modulate several pathways, as measured using luciferase reporter assays. Herein, we report that none of the identified PTMs affected HBZ stability or its regulation of tested pathways.

  7. Post-Translational Modifications of H2A Histone Variants and Their Role in Cancer

    Directory of Open Access Journals (Sweden)

    David Corujo

    2018-02-01

    Full Text Available Histone variants are chromatin components that replace replication-coupled histones in a fraction of nucleosomes and confer particular characteristics to chromatin. H2A variants represent the most numerous and diverse group among histone protein families. In the nucleosomal structure, H2A-H2B dimers can be removed and exchanged more easily than the stable H3-H4 core. The unstructured N-terminal histone tails of all histones, but also the C-terminal tails of H2A histones protrude out of the compact structure of the nucleosome core. These accessible tails are the preferential target sites for a large number of post-translational modifications (PTMs. While some PTMs are shared between replication-coupled H2A and H2A variants, many modifications are limited to a specific histone variant. The present review focuses on the H2A variants H2A.Z, H2A.X, and macroH2A, and summarizes their functions in chromatin and how these are linked to cancer development and progression. H2A.Z primarily acts as an oncogene and macroH2A and H2A.X as tumour suppressors. We further focus on the regulation by PTMs, which helps to understand a degree of context dependency.

  8. Multiple post-translational modifications in hepatocyte nuclear factor 4α

    International Nuclear Information System (INIS)

    Yokoyama, Atsushi; Katsura, Shogo; Ito, Ryo; Hashiba, Waka; Sekine, Hiroki; Fujiki, Ryoji; Kato, Shigeaki

    2011-01-01

    Highlights: → We performed comprehensive PTM analysis for HNF4α protein. → We identified 8 PTMs in HNF4α protein including newly identified PTMs. → Among them, we found acetylation at lysine 458 was one of the prime PTMs for HNF4α function. → Acetylation at lysine 458 was inhibitory for HNF4α transcription function. → This modification fluctuated in response to extracellular condition. -- Abstract: To investigate the role of post-translational modifications (PTMs) in the hepatocyte nuclear factor 4α (HNF4α)-mediated transcription, we took a comprehensive survey of PTMs in HNF4α protein by massspectrometry and identified totally 8 PTM sites including newly identified ubiquitilation and acetylation sites. To assess the impact of identified PTMs in HNF4α-function, we introduced point mutations at the identified PTM sites and, tested transcriptional activity of the HNF4α. Among the point-mutations, an acetylation site at lysine 458 was found significant in the HNF4α-mediated transcriptional control. An acetylation negative mutant at lysine 458 showed an increased transcriptional activity by about 2-fold, while an acetylation mimic mutant had a lowered transcriptional activation. Furthermore, this acetylation appeared to be fluctuated in response to extracellular nutrient conditions. Thus, by applying an comprehensive analysis of PTMs, multiple PTMs were newly identified in HNF4α and unexpected role of an HNF4α acetylation could be uncovered.

  9. Overview of xeroderma pigmentosum proteins architecture, mutations and post-translational modifications.

    Science.gov (United States)

    Feltes, Bruno César; Bonatto, Diego

    2015-01-01

    The xeroderma pigmentosum complementation group proteins (XPs), which include XPA through XPG, play a critical role in coordinating and promoting global genome and transcription-coupled nucleotide excision repair (GG-NER and TC-NER, respectively) pathways in eukaryotic cells. GG-NER and TC-NER are both required for the repair of bulky DNA lesions, such as those induced by UV radiation. Mutations in genes that encode XPs lead to the clinical condition xeroderma pigmentosum (XP). Although the roles of XPs in the GG-NER/TC-NER subpathways have been extensively studied, complete knowledge of their three-dimensional structure is only beginning to emerge. Hence, this review aims to summarize the current knowledge of mapped mutations and other structural information on XP proteins that influence their function and protein-protein interactions. We also review the possible post-translational modifications for each protein and the impact of these modifications on XP protein functions. Copyright © 2014 Elsevier B.V. All rights reserved.

  10. Electrospray mass spectrometry characterization of post-translational modifications of barley alpha-amylase 1 produced in yeast

    DEFF Research Database (Denmark)

    Søgaard, M; Andersen, Jens S.; Roepstorff, P

    1993-01-01

    We have used electrospray mass spectrometry (ESMS) in combination with protein chemistry and genetics to delineate post-translational modifications in yeast of barley alpha-amylase 1 (AMY1), a 45 kD enzyme crucial for production of malt, an important starting material in the manufacture of beer...

  11. Role of post-translational modifications on structure, function and pharmacology of class C G protein-coupled receptors

    DEFF Research Database (Denmark)

    Nørskov-Lauritsen, Lenea; Bräuner-Osborne, Hans

    2015-01-01

    taste receptors (T1R1-3), one calcium-sensing (CaS) receptor, one GPCR, class C, group 6, subtype A (GPRC6) receptor, and seven orphan receptors. G protein-coupled receptors undergo a number of post-translational modifications, which regulate their structure, function and/or pharmacology. Here, we...

  12. An integrative analysis of post-translational histone modifications in the marine diatom Phaeodactylum tricornutum

    KAUST Repository

    Veluchamy, Alaguraj

    2015-05-20

    Background: Nucleosomes are the building blocks of chromatin where gene regulation takes place. Chromatin landscapes have been profiled for several species, providing insights into the fundamental mechanisms of chromatin-mediated transcriptional regulation of gene expression. However, knowledge is missing for several major and deep-branching eukaryotic groups, such as the Stramenopiles, which include the diatoms. Diatoms are highly diverse and ubiquitous species of phytoplankton that play a key role in global biogeochemical cycles. Dissecting chromatin-mediated regulation of genes in diatoms will help understand the ecological success of these organisms in contemporary oceans. Results: Here, we use high resolution mass spectrometry to identify a full repertoire of post-translational modifications on histones of the marine diatom Phaeodactylum tricornutum, including eight novel modifications. We map five histone marks coupled with expression data and show that P. tricornutum displays both unique and broadly conserved chromatin features, reflecting the chimeric nature of its genome. Combinatorial analysis of histone marks and DNA methylation demonstrates the presence of an epigenetic code defining activating or repressive chromatin states. We further profile three specific histone marks under conditions of nitrate depletion and show that the histone code is dynamic and targets specific sets of genes. Conclusions: This study is the first genome-wide characterization of the histone code from a stramenopile and a marine phytoplankton. The work represents an important initial step for understanding the evolutionary history of chromatin and how epigenetic modifications affect gene expression in response to environmental cues in marine environments. © 2015 Veluchamy et al.

  13. Large scale analysis of co-existing post-translational modifications in histone tails reveals global fine structure of cross-talk

    DEFF Research Database (Denmark)

    Schwämmle, Veit; Aspalter, Claudia-Maria; Sidoli, Simone

    2014-01-01

    Mass spectrometry (MS) is a powerful analytical method for the identification and quantification of co-existing post-translational modifications in histone proteins. One of the most important challenges in current chromatin biology is to characterize the relationships between co-existing histone...... sample-specific patterns for the co-frequency of histone post-translational modifications. We implemented a new method to identify positive and negative interplay between pairs of methylation and acetylation marks in proteins. Many of the detected features were conserved between different cell types...... sites but negative cross-talk for distant ones, and for discrete methylation states at Lys-9, Lys-27, and Lys-36 of histone H3, suggesting a more differentiated functional role of methylation beyond the general expectation of enhanced activity at higher methylation states....

  14. In Silico Analysis of Correlations between Protein Disorder and Post-Translational Modifications in Algae

    Directory of Open Access Journals (Sweden)

    Atsushi Kurotani

    2015-08-01

    Full Text Available Recent proteome analyses have reported that intrinsically disordered regions (IDRs of proteins play important roles in biological processes. In higher plants whose genomes have been sequenced, the correlation between IDRs and post-translational modifications (PTMs has been reported. The genomes of various eukaryotic algae as common ancestors of plants have also been sequenced. However, no analysis of the relationship to protein properties such as structure and PTMs in algae has been reported. Here, we describe correlations between IDR content and the number of PTM sites for phosphorylation, glycosylation, and ubiquitination, and between IDR content and regions rich in proline, glutamic acid, serine, and threonine (PEST and transmembrane helices in the sequences of 20 algae proteomes. Phosphorylation, O-glycosylation, ubiquitination, and PEST preferentially occurred in disordered regions. In contrast, transmembrane helices were favored in ordered regions. N-glycosylation tended to occur in ordered regions in most of the studied algae; however, it correlated positively with disordered protein content in diatoms. Additionally, we observed that disordered protein content and the number of PTM sites were significantly increased in the species-specific protein clusters compared to common protein clusters among the algae. Moreover, there were specific relationships between IDRs and PTMs among the algae from different groups.

  15. In Silico Analysis of Correlations between Protein Disorder and Post-Translational Modifications in Algae.

    Science.gov (United States)

    Kurotani, Atsushi; Sakurai, Tetsuya

    2015-08-20

    Recent proteome analyses have reported that intrinsically disordered regions (IDRs) of proteins play important roles in biological processes. In higher plants whose genomes have been sequenced, the correlation between IDRs and post-translational modifications (PTMs) has been reported. The genomes of various eukaryotic algae as common ancestors of plants have also been sequenced. However, no analysis of the relationship to protein properties such as structure and PTMs in algae has been reported. Here, we describe correlations between IDR content and the number of PTM sites for phosphorylation, glycosylation, and ubiquitination, and between IDR content and regions rich in proline, glutamic acid, serine, and threonine (PEST) and transmembrane helices in the sequences of 20 algae proteomes. Phosphorylation, O-glycosylation, ubiquitination, and PEST preferentially occurred in disordered regions. In contrast, transmembrane helices were favored in ordered regions. N-glycosylation tended to occur in ordered regions in most of the studied algae; however, it correlated positively with disordered protein content in diatoms. Additionally, we observed that disordered protein content and the number of PTM sites were significantly increased in the species-specific protein clusters compared to common protein clusters among the algae. Moreover, there were specific relationships between IDRs and PTMs among the algae from different groups.

  16. Molecular classification of fatty liver by high-throughput profiling of protein post-translational modifications.

    Science.gov (United States)

    Urasaki, Yasuyo; Fiscus, Ronald R; Le, Thuc T

    2016-04-01

    We describe an alternative approach to classifying fatty liver by profiling protein post-translational modifications (PTMs) with high-throughput capillary isoelectric focusing (cIEF) immunoassays. Four strains of mice were studied, with fatty livers induced by different causes, such as ageing, genetic mutation, acute drug usage, and high-fat diet. Nutrient-sensitive PTMs of a panel of 12 liver metabolic and signalling proteins were simultaneously evaluated with cIEF immunoassays, using nanograms of total cellular protein per assay. Changes to liver protein acetylation, phosphorylation, and O-N-acetylglucosamine glycosylation were quantified and compared between normal and diseased states. Fatty liver tissues could be distinguished from one another by distinctive protein PTM profiles. Fatty liver is currently classified by morphological assessment of lipid droplets, without identifying the underlying molecular causes. In contrast, high-throughput profiling of protein PTMs has the potential to provide molecular classification of fatty liver. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

  17. Extensive and systematic rewiring of histone post-translational modifications in cancer model systems.

    Science.gov (United States)

    Noberini, Roberta; Osti, Daniela; Miccolo, Claudia; Richichi, Cristina; Lupia, Michela; Corleone, Giacomo; Hong, Sung-Pil; Colombo, Piergiuseppe; Pollo, Bianca; Fornasari, Lorenzo; Pruneri, Giancarlo; Magnani, Luca; Cavallaro, Ugo; Chiocca, Susanna; Minucci, Saverio; Pelicci, Giuliana; Bonaldi, Tiziana

    2018-05-04

    Histone post-translational modifications (PTMs) generate a complex combinatorial code that regulates gene expression and nuclear functions, and whose deregulation has been documented in different types of cancers. Therefore, the availability of relevant culture models that can be manipulated and that retain the epigenetic features of the tissue of origin is absolutely crucial for studying the epigenetic mechanisms underlying cancer and testing epigenetic drugs. In this study, we took advantage of quantitative mass spectrometry to comprehensively profile histone PTMs in patient tumor tissues, primary cultures and cell lines from three representative tumor models, breast cancer, glioblastoma and ovarian cancer, revealing an extensive and systematic rewiring of histone marks in cell culture conditions, which includes a decrease of H3K27me2/me3, H3K79me1/me2 and H3K9ac/K14ac, and an increase of H3K36me1/me2. While some changes occur in short-term primary cultures, most of them are instead time-dependent and appear only in long-term cultures. Remarkably, such changes mostly revert in cell line- and primary cell-derived in vivo xenograft models. Taken together, these results support the use of xenografts as the most representative models of in vivo epigenetic processes, suggesting caution when using cultured cells, in particular cell lines and long-term primary cultures, for epigenetic investigations.

  18. Antioxidant systems are regulated by nitric oxide-mediated post-translational modifications (NO-PTMs

    Directory of Open Access Journals (Sweden)

    Juan Carlos Begara-Morales

    2016-02-01

    Full Text Available Nitric oxide (NO is a biological messenger that orchestrates a plethora of plant functions, mainly through post-translational modifications (PTMs such as S-nitrosylation or tyrosine nitration. In plants, hundreds of proteins have been identified as potential targets of these NO-PTMs under physiological and stress conditions indicating the relevance of NO in plant-signaling mechanisms. Among these NO protein targets, there are different antioxidant enzymes involved in the control of reactive oxygen species (ROS, such as H2O2, which is also a signal molecule. This highlights the close relationship between ROS/NO signaling pathways. The major plant antioxidant enzymes, including catalase, superoxide dismutases (SODs peroxiredoxins (Prx and all the enzymatic components of the ascorbate-glutathione (Asa-GSH cycle, have been shown to be modulated to different degrees by NO-PTMs. This mini-review will update the recent knowledge concerning the interaction of NO with these antioxidant enzymes, with a special focus on the components of the Asa-GSH cycle and their physiological relevance.

  19. Post-translational modifications are key players of the Legionella pneumophila infection strategy

    Science.gov (United States)

    Michard, Céline; Doublet, Patricia

    2015-01-01

    Post-translational modifications (PTMs) are widely used by eukaryotes to control the enzymatic activity, localization or stability of their proteins. Traditionally, it was believed that the broad biochemical diversity of the PTMs is restricted to eukaryotic cells, which exploit it in extensive networks to fine-tune various and complex cellular functions. During the last decade, the advanced detection methods of PTMs and functional studies of the host–pathogen relationships highlight that bacteria have also developed a large arsenal of PTMs, particularly to subvert host cell pathways to their benefit. Legionella pneumophila, the etiological agent of the severe pneumonia legionellosis, is the paradigm of highly adapted intravacuolar pathogens that have set up sophisticated biochemical strategies. Among them, L. pneumophila has evolved eukaryotic-like and rare/novel PTMs to hijack host cell processes. Here, we review recent progress about the diversity of PTMs catalyzed by Legionella: ubiquitination, prenylation, phosphorylation, glycosylation, methylation, AMPylation, and de-AMPylation, phosphocholination, and de-phosphocholination. We focus on the host cell pathways targeted by the bacteria catalyzed PTMs and we stress the importance of the PTMs in the Legionella infection strategy. Finally, we highlight that the discovery of these PTMs undoubtedly made significant breakthroughs on the molecular basis of Legionella pathogenesis but also lead the way in improving our knowledge of the eukaryotic PTMs and complex cellular processes that are associated to. PMID:25713573

  20. Developmentally Regulated Post-translational Modification of Nucleoplasmin Controls Histone Sequestration and Deposition

    Directory of Open Access Journals (Sweden)

    Takashi Onikubo

    2015-03-01

    Full Text Available Nucleoplasmin (Npm is an abundant histone chaperone in vertebrate oocytes and embryos. During embryogenesis, regulation of Npm histone binding is critical for its function in storing and releasing maternal histones to establish and maintain the zygotic epigenome. Here, we demonstrate that Xenopus laevis Npm post-translational modifications (PTMs specific to the oocyte and egg promote either histone deposition or sequestration, respectively. Mass spectrometry and Npm phosphomimetic mutations used in chromatin assembly assays identified hyperphosphorylation on the N-terminal tail as a critical regulator for sequestration. C-terminal tail phosphorylation and PRMT5-catalyzed arginine methylation enhance nucleosome assembly by promoting histone interaction with the second acidic tract of Npm. Electron microscopy reconstructions of Npm and TTLL4 activity toward the C-terminal tail demonstrate that oocyte- and egg-specific PTMs cause Npm conformational changes. Our results reveal that PTMs regulate Npm chaperoning activity by modulating Npm conformation and Npm-histone interaction, leading to histone sequestration in the egg.

  1. Impact of Post-Translational Modifications of Crop Proteins under Abiotic Stress

    Directory of Open Access Journals (Sweden)

    Akiko Hashiguchi

    2016-12-01

    Full Text Available The efficiency of stress-induced adaptive responses of plants depends on intricate coordination of multiple signal transduction pathways that act coordinately or, in some cases, antagonistically. Protein post-translational modifications (PTMs can regulate protein activity and localization as well as protein–protein interactions in numerous cellular processes, thus leading to elaborate regulation of plant responses to various external stimuli. Understanding responses of crop plants under field conditions is crucial to design novel stress-tolerant cultivars that maintain robust homeostasis even under extreme conditions. In this review, proteomic studies of PTMs in crops are summarized. Although the research on the roles of crop PTMs in regulating stress response mechanisms is still in its early stage, several novel insights have been retrieved so far. This review covers techniques for detection of PTMs in plants, representative PTMs in plants under abiotic stress, and how PTMs control functions of representative proteins. In addition, because PTMs under abiotic stresses are well described in soybeans under submergence, recent findings in PTMs of soybean proteins under flooding stress are introduced. This review provides information on advances in PTM study in relation to plant adaptations to abiotic stresses, underlining the importance of PTM study to ensure adequate agricultural production in the future.

  2. Bug22 influences cilium morphology and the post-translational modification of ciliary microtubules

    Directory of Open Access Journals (Sweden)

    Teresa Mendes Maia

    2014-01-01

    Cilia and flagella are organelles essential for motility and sensing of environmental stimuli. Depending on the cell type, cilia acquire a defined set of functions and, accordingly, are built with an appropriate length and molecular composition. Several ciliary proteins display a high degree of conservation throughout evolution and mutations in ciliary genes are associated with various diseases such as ciliopathies and infertility. Here, we describe the role of the highly conserved ciliary protein, Bug22, in Drosophila. Previous studies in unicellular organisms have shown that Bug22 is required for proper cilia function, but its exact role in ciliogenesis has not been investigated yet. Null Bug22 mutant flies display cilia-associated phenotypes and nervous system defects. Furthermore, sperm differentiation is blocked at the individualization stage, due to impaired migration of the individualization machinery. Tubulin post-translational modifications (PTMs such as polyglycylation, polyglutamylation or acetylation, are determinants of microtubule (MT functions and stability in centrioles, cilia and neurons. We found defects in the timely incorporation of polyglycylation in sperm axonemal MTs of Bug22 mutants. In addition, we found that depletion of human Bug22 in RPE1 cells resulted in the appearance of longer cilia and reduced axonemal polyglutamylation. Our work identifies Bug22 as a protein that plays a conserved role in the regulation of PTMs of the ciliary axoneme.

  3. Unrestrictive identification of post-translational modifications in the urine proteome without enrichment

    Science.gov (United States)

    2013-01-01

    Background Research on the human urine proteome may lay the foundation for the discovery of relevant disease biomarkers. Post-translational modifications (PTMs) have important effects on the functions of protein biomarkers. Identifying PTMs without enrichment adds no extra steps to conventional identification procedures for urine proteomics. The only difference is that this method requires software that can conduct unrestrictive identifications of PTMs. In this study, routine urine proteomics techniques were used to identify urine proteins. Unspecified PTMs were searched by MODa and PEAKS 6 automated software, followed by a manual search to screen out in vivo PTMs by removing all in vitro PTMs and amino acid substitutions. Results There were 75 peptides with 6 in vivo PTMs that were found by both MODa and PEAKS 6. Of these, 34 peptides in 18 proteins have novel in vivo PTMs compared with the annotation information of these proteins on the Universal Protein Resource website. These new in vivo PTMs had undergone methylation, dehydration, oxidation, hydroxylation, phosphorylation, or dihydroxylation. Conclusions In this study, we identified PTMs of urine proteins without the need for enrichment. Our investigation may provide a useful reference for biomarker discovery in the future. PMID:23317149

  4. Evolutionary constraint and disease associations of post-translational modification sites in human genomes.

    Directory of Open Access Journals (Sweden)

    Jüri Reimand

    2015-01-01

    Full Text Available Interpreting the impact of human genome variation on phenotype is challenging. The functional effect of protein-coding variants is often predicted using sequence conservation and population frequency data, however other factors are likely relevant. We hypothesized that variants in protein post-translational modification (PTM sites contribute to phenotype variation and disease. We analyzed fraction of rare variants and non-synonymous to synonymous variant ratio (Ka/Ks in 7,500 human genomes and found a significant negative selection signal in PTM regions independent of six factors, including conservation, codon usage, and GC-content, that is widely distributed across tissue-specific genes and function classes. PTM regions are also enriched in known disease mutations, suggesting that PTM variation is more likely deleterious. PTM constraint also affects flanking sequence around modified residues and increases around clustered sites, indicating presence of functionally important short linear motifs. Using target site motifs of 124 kinases, we predict that at least ∼180,000 motif-breaker amino acid residues that disrupt PTM sites when substituted, and highlight kinase motifs that show specific negative selection and enrichment of disease mutations. We provide this dataset with corresponding hypothesized mechanisms as a community resource. As an example of our integrative approach, we propose that PTPN11 variants in Noonan syndrome aberrantly activate the protein by disrupting an uncharacterized cluster of phosphorylation sites. Further, as PTMs are molecular switches that are modulated by drugs, we study mutated binding sites of PTM enzymes in disease genes and define a drug-disease network containing 413 novel predicted disease-gene links.

  5. Pathogenic Leptospires Modulate Protein Expression and Post-translational Modifications in Response to Mammalian Host Signals.

    Science.gov (United States)

    Nally, Jarlath E; Grassmann, Andre A; Planchon, Sébastien; Sergeant, Kjell; Renaut, Jenny; Seshu, Janakiram; McBride, Alan J; Caimano, Melissa J

    2017-01-01

    Pathogenic species of Leptospira cause leptospirosis, a bacterial zoonotic disease with a global distribution affecting over one million people annually. Reservoir hosts of leptospirosis, including rodents, dogs, and cattle, exhibit little to no signs of disease but shed large numbers of organisms in their urine. Transmission occurs when mucosal surfaces or abraded skin come into contact with infected urine or urine-contaminated water or soil. Whilst little is known about how Leptospira adapt to and persist within a reservoir host, in vitro studies suggest that leptospires alter their transcriptomic and proteomic profiles in response to environmental signals encountered during mammalian infection. We applied the dialysis membrane chamber (DMC) peritoneal implant model to compare the whole cell proteome of in vivo derived leptospires with that of leptospires cultivated in vitro at 30°C and 37°C by 2-dimensional difference in-gel electrophoresis (2-D DIGE). Of 1,735 protein spots aligned across 9 2-D DIGE gels, 202 protein spots were differentially expressed ( p 1.25 or expressed proteins were excised for identification by mass spectrometry. Data are available via ProteomeXchange with identifier PXD006995. The greatest differences were detected when DMC-cultivated leptospires were compared with IV30- or IV37-cultivated leptospires, including the increased expression of multiple isoforms of Loa22, a known virulence factor. Unexpectedly, 20 protein isoforms of LipL32 and 7 isoforms of LipL41 were uniformly identified by DIGE as differentially expressed, suggesting that unique post-translational modifications (PTMs) are operative in response to mammalian host conditions. To test this hypothesis, a rat model of persistent renal colonization was used to isolate leptospires directly from the urine of experimentally infected rats. Comparison of urinary derived leptospires to IV30 leptospires by 2-D immunoblotting confirmed that modification of proteins with

  6. Pathogenic Leptospires Modulate Protein Expression and Post-translational Modifications in Response to Mammalian Host Signals

    Directory of Open Access Journals (Sweden)

    Jarlath E. Nally

    2017-08-01

    Full Text Available Pathogenic species of Leptospira cause leptospirosis, a bacterial zoonotic disease with a global distribution affecting over one million people annually. Reservoir hosts of leptospirosis, including rodents, dogs, and cattle, exhibit little to no signs of disease but shed large numbers of organisms in their urine. Transmission occurs when mucosal surfaces or abraded skin come into contact with infected urine or urine-contaminated water or soil. Whilst little is known about how Leptospira adapt to and persist within a reservoir host, in vitro studies suggest that leptospires alter their transcriptomic and proteomic profiles in response to environmental signals encountered during mammalian infection. We applied the dialysis membrane chamber (DMC peritoneal implant model to compare the whole cell proteome of in vivo derived leptospires with that of leptospires cultivated in vitro at 30°C and 37°C by 2-dimensional difference in-gel electrophoresis (2-D DIGE. Of 1,735 protein spots aligned across 9 2-D DIGE gels, 202 protein spots were differentially expressed (p < 0.05, fold change >1.25 or < −1.25 across all three conditions. Differentially expressed proteins were excised for identification by mass spectrometry. Data are available via ProteomeXchange with identifier PXD006995. The greatest differences were detected when DMC-cultivated leptospires were compared with IV30- or IV37-cultivated leptospires, including the increased expression of multiple isoforms of Loa22, a known virulence factor. Unexpectedly, 20 protein isoforms of LipL32 and 7 isoforms of LipL41 were uniformly identified by DIGE as differentially expressed, suggesting that unique post-translational modifications (PTMs are operative in response to mammalian host conditions. To test this hypothesis, a rat model of persistent renal colonization was used to isolate leptospires directly from the urine of experimentally infected rats. Comparison of urinary derived leptospires to IV30

  7. Analysis of Histones H3 and H4 Reveals Novel and Conserved Post-Translational Modifications in Sugarcane.

    Science.gov (United States)

    Moraes, Izabel; Yuan, Zuo-Fei; Liu, Shichong; Souza, Glaucia Mendes; Garcia, Benjamin A; Casas-Mollano, J Armando

    2015-01-01

    Histones are the main structural components of the nucleosome, hence targets of many regulatory proteins that mediate processes involving changes in chromatin. The functional outcome of many pathways is "written" in the histones in the form of post-translational modifications that determine the final gene expression readout. As a result, modifications, alone or in combination, are important determinants of chromatin states. Histone modifications are accomplished by the addition of different chemical groups such as methyl, acetyl and phosphate. Thus, identifying and characterizing these modifications and the proteins related to them is the initial step to understanding the mechanisms of gene regulation and in the future may even provide tools for breeding programs. Several studies over the past years have contributed to increase our knowledge of epigenetic gene regulation in model organisms like Arabidopsis, yet this field remains relatively unexplored in crops. In this study we identified and initially characterized histones H3 and H4 in the monocot crop sugarcane. We discovered a number of histone genes by searching the sugarcane ESTs database. The proteins encoded correspond to canonical histones, and their variants. We also purified bulk histones and used them to map post-translational modifications in the histones H3 and H4 using mass spectrometry. Several modifications conserved in other plants, and also novel modified residues, were identified. In particular, we report O-acetylation of serine, threonine and tyrosine, a recently identified modification conserved in several eukaryotes. Additionally, the sub-nuclear localization of some well-studied modifications (i.e., H3K4me3, H3K9me2, H3K27me3, H3K9ac, H3T3ph) is described and compared to other plant species. To our knowledge, this is the first report of histones H3 and H4 as well as their post-translational modifications in sugarcane, and will provide a starting point for the study of chromatin regulation in

  8. Prediction of post translational modifications in avicennia marina Cu-Zn superoxide dismutase: implication of glycation on the enzyme structure

    International Nuclear Information System (INIS)

    Jabeen, U.; Salim, A.; Abbasi, A.

    2012-01-01

    3D homology model of Cu-Zn superoxide dismutase (SOD) from Avicennia marina (AMSOD) was constructed using the structural coordinates of Spinach SOD (SSOD). Prediction of post translational modification was done by PROSITE. The predicted sites were examined in the 3D model. AMSOD model was glycated using modeling software and changes in the structure was analyzed after glycation. The analysis revealed some potential sites and structural changes after glycation. (author)

  9. Proteomic analysis reveals APC-dependent post-translational modifications and identifies a novel regulator of β-catenin.

    Science.gov (United States)

    Blundon, Malachi A; Schlesinger, Danielle R; Parthasarathy, Amritha; Smith, Samantha L; Kolev, Hannah M; Vinson, David A; Kunttas-Tatli, Ezgi; McCartney, Brooke M; Minden, Jonathan S

    2016-07-15

    Wnt signaling generates patterns in all embryos, from flies to humans, and controls cell fate, proliferation and metabolic homeostasis. Inappropriate Wnt pathway activation results in diseases, including colorectal cancer. The adenomatous polyposis coli (APC) tumor suppressor gene encodes a multifunctional protein that is an essential regulator of Wnt signaling and cytoskeletal organization. Although progress has been made in defining the role of APC in a normal cellular context, there are still significant gaps in our understanding of APC-dependent cellular function and dysfunction. We expanded the APC-associated protein network using a combination of genetics and a proteomic technique called two-dimensional difference gel electrophoresis (2D-DIGE). We show that loss of Drosophila Apc2 causes protein isoform changes reflecting misregulation of post-translational modifications (PTMs), which are not dependent on β-catenin transcriptional activity. Mass spectrometry revealed that proteins involved in metabolic and biosynthetic pathways, protein synthesis and degradation, and cell signaling are affected by Apc2 loss. We demonstrate that changes in phosphorylation partially account for the altered PTMs in APC mutants, suggesting that APC mutants affect other types of PTM. Finally, through this approach Aminopeptidase P was identified as a new regulator of β-catenin abundance in Drosophila embryos. This study provides new perspectives on the cellular effects of APC that might lead to a deeper understanding of its role in development. © 2016. Published by The Company of Biologists Ltd.

  10. Post-translational modifications of voltage-gated sodium channels in chronic pain syndromes.

    Directory of Open Access Journals (Sweden)

    Cédric James Laedermann

    2015-11-01

    Full Text Available In the peripheral sensory nervous system the neuronal expression of voltage-gated sodium channels (Navs is a very important for the transmission of nociceptive information since they give rise to the upstroke of the action potential. Navs are composed of 9 different isoforms with distinct biophysical properties. Studying the mutations associated with the increase or absence of pain sensitivity in humans, as well as other expression studies, have highlighted Nav1.7, Nav1.8 and Nav1.9 as being the most important contributors to the control of nociceptive neuronal electrogenesis. Modulating their expression and/or function can impact the shape of the action potential and consequently modify pain transmission, a process that is observed in persistent pain conditions.Post-translational modification (PTM of Navs is a well-known process that modifies their expression and function. In chronic pain syndromes, the release of inflammatory molecules into the direct environment of dorsal root ganglia (DRG sensory neurons leads to an abnormal activation of enzymes that induce Navs PTM. The addition of small molecules, i.e. peptides, phosphoryl groups, ubiquitin moieties and/or carbohydrates, can modify the function of Navs in two different ways: via direct physical interference with the subunit of Nav gating, or via the control of Nav trafficking. Both mechanisms have a profound impact on neuronal excitability. In this review we will discuss the role of Protein Kinase A, B and C, Mitogen Activated Protein Kinases and Ca++/Calmodulin-dependent Kinase II in peripheral chronic pain syndromes. We will also discuss more recent findings that the ubiquitination of Nav1.7 by Nedd4-2 and the effect of methylglyoxal on Nav1.8 are also implicated in the development of experimental neuropathic pain. We will address the potential roles of other PTMs in chronic pain and highlight the need for further investigation of PTMs of Navs in order to develop new pharmacological

  11. The phosphopantetheinyl transferases: catalysis of a post-translational modification crucial for life

    DEFF Research Database (Denmark)

    Beld, Joris; Sonnenschein, Eva; Vickery, Christopher R.

    2013-01-01

    Covering: up to 2013 Although holo-acyl carrier protein synthase, AcpS, a phosphopantetheinyl transferase (PPTase), was characterized in the 1960s, it was not until the publication of the landmark paper by Lambalot et al. in 1996 that PPTases garnered wide-spread attention being classified...... knowledge on this class of enzymes that post-translationally install a 4′-phosphopantetheine arm on various carrier proteins....

  12. Comparison at the peptide level with post-translational modification consideration reveals more differences between two unenriched samples.

    Science.gov (United States)

    Yin, Jianrui; Shao, Chen; Jia, Lulu; Gao, Youhe

    2014-06-30

    In shotgun strategies, peptide sequences are first identified from tandem mass (MS/MS) spectra, and the existence and abundance of the proteins are then inferred from the peptide information. However, the protein inference step can produce errors and a loss of information. To identify the information that is lost using the traditional approaches, this study compared the proteomic data of two leukemia cell lines (Jurkat and K562) at the peptide level with consideration of post-translational modifications (PTMs). The raw files from the two cell lines were searched against the decoy IPI-human database version 3.68, which contains forward and reverse sequences. Then the observed modification name in the results was matched with the modification classification on the Unimod website by a manual search. Only the peptides with 'post-translational' modifications were compared between the two cell lines. After searching the database with consideration of PTMs, a total of 44046 non-redundant peptides were identified in both the Jurkat and K562 cell lines. Of these peptides, even without specific PTM enrichment, 11.43% of them (with at least two spectra in one cell line) existed in different PTM forms between the two cell lines, and 1.73% of the peptides were modified in both cell lines, but with different modifications or possibly on different sites. Comparing proteomic data at the peptide level with consideration of PTMs can reveal more differences between two unenriched samples. Copyright © 2014 John Wiley & Sons, Ltd.

  13. Quantitative proteomic analysis of post-translational modifications of human histones

    DEFF Research Database (Denmark)

    Beck, Hans Christian; Nielsen, Eva C; Matthiesen, Rune

    2006-01-01

    , and H4 in a site-specific and dose-dependent manner. This unbiased analysis revealed that a relative increase in acetylated peptide from the histone variants H2A, H2B, and H4 was accompanied by a relative decrease of dimethylated Lys(57) from histone H2B. The dose-response results obtained...... by quantitative proteomics of histones from HDACi-treated cells were consistent with Western blot analysis of histone acetylation, cytotoxicity, and dose-dependent expression profiles of p21 and cyclin A2. This demonstrates that mass spectrometry-based quantitative proteomic analysis of post-translational...

  14. Multiple γ-glutamylation: A novel type of post-translational modification in a diapausing Artemia cyst protein

    International Nuclear Information System (INIS)

    Hasegawa, Mai; Ikeda, Yuka; Kanzawa, Hideaki; Sakamoto, Mika; Goto, Mina; Tsunasawa, Susumu; Uchiumi, Toshio; Odani, Shoji

    2010-01-01

    A highly hydrophilic, glutamate-rich protein was identified in the aqueous phenol extract from the cytosolic fraction of brine shrimp (Artemia franciscana) diapausing cysts and termed Artemia phenol soluble protein (PSP). Mass spectrometric analysis revealed the presence of many protein peaks around m/z 11,000, separated by 129 atomic mass units; this value corresponds to that of glutamate, which is strongly suggestive of heterogeneous polyglutamylation. Polyglutamylation has long been known as the functionally important post-translational modification of tubulins, which carry poly(L-glutamic acid) chains of heterogeneous length branching off from the main chain at the γ-carboxy groups of a few specific glutamate residues. In Artemia PSP, however, Edman degradation of enzymatic peptides revealed that at least 13, and presumably 16, glutamate residues were modified by the attachment of a single L-glutamate, representing a hitherto undescribed type of post-translational modification: namely, multiple γ-glutamylation or the addition of a large number of glutamate residues along the polypeptide chain. Although biological significance of PSP and its modification is yet to be established, suppression of in vitro thermal aggregation of lactate dehydrogenase by glutamylated PSP was observed.

  15. ReportSites - A Computational Method to Extract Positional and Physico- Chemical Information from Large-Scale Proteomic Post-Translational Modification Datasets

    DEFF Research Database (Denmark)

    Edwards, Alistair; Edwards, Gregory; Larsen, Martin Røssel

    2012-01-01

    -translational modification data sets, wherein patterns of sequence surrounding processed sites may reveal more about the functional and structural requirements of the modification and the biochemical processes that regulate them. Results: We developed Report Sites using a test set of phosphoproteomic data from rat......-chemical environment (local pI and hydrophobicity). These were then also compared to corresponding values extracted from the full database to allow comparison of phosphorylation trends. Conclusions: Report Sites enabled physico-chemical aspects of protein phosphorylation to be deciphered in a test set of eleven...... thousand phospho sites. Basic properties of modified proteins, such as site location in the context of the complete protein, were also documented. This program can be easily adapted to any post-translational modification (or, indeed, to any defined amino acid sequence), or expanded to include more...

  16. Importance of post-translational modifications for functionality of a chloroplast-localized carbonic anhydrase (CAH1 in Arabidopsis thaliana.

    Directory of Open Access Journals (Sweden)

    Stefan Burén

    Full Text Available BACKGROUND: The Arabidopsis CAH1 alpha-type carbonic anhydrase is one of the few plant proteins known to be targeted to the chloroplast through the secretory pathway. CAH1 is post-translationally modified at several residues by the attachment of N-glycans, resulting in a mature protein harbouring complex-type glycans. The reason of why trafficking through this non-canonical pathway is beneficial for certain chloroplast resident proteins is not yet known. Therefore, to elucidate the significance of glycosylation in trafficking and the effect of glycosylation on the stability and function of the protein, epitope-labelled wild type and mutated versions of CAH1 were expressed in plant cells. METHODOLOGY/PRINCIPAL FINDINGS: Transient expression of mutant CAH1 with disrupted glycosylation sites showed that the protein harbours four, or in certain cases five, N-glycans. While the wild type protein trafficked through the secretory pathway to the chloroplast, the non-glycosylated protein formed aggregates and associated with the ER chaperone BiP, indicating that glycosylation of CAH1 facilitates folding and ER-export. Using cysteine mutants we also assessed the role of disulphide bridge formation in the folding and stability of CAH1. We found that a disulphide bridge between cysteines at positions 27 and 191 in the mature protein was required for correct folding of the protein. Using a mass spectrometric approach we were able to measure the enzymatic activity of CAH1 protein. Under circumstances where protein N-glycosylation is blocked in vivo, the activity of CAH1 is completely inhibited. CONCLUSIONS/SIGNIFICANCE: We show for the first time the importance of post-translational modifications such as N-glycosylation and intramolecular disulphide bridge formation in folding and trafficking of a protein from the secretory pathway to the chloroplast in higher plants. Requirements for these post-translational modifications for a fully functional native

  17. Amyloid β production is regulated by β2-adrenergic signaling-mediated post-translational modifications of the ryanodine receptor.

    Science.gov (United States)

    Bussiere, Renaud; Lacampagne, Alain; Reiken, Steven; Liu, Xiaoping; Scheuerman, Valerie; Zalk, Ran; Martin, Cécile; Checler, Frederic; Marks, Andrew R; Chami, Mounia

    2017-06-16

    Alteration of ryanodine receptor (RyR)-mediated calcium (Ca 2+ ) signaling has been reported in Alzheimer disease (AD) models. However, the molecular mechanisms underlying altered RyR-mediated intracellular Ca 2+ release in AD remain to be fully elucidated. We report here that RyR2 undergoes post-translational modifications (phosphorylation, oxidation, and nitrosylation) in SH-SY5Y neuroblastoma cells expressing the β-amyloid precursor protein (βAPP) harboring the familial double Swedish mutations (APPswe). RyR2 macromolecular complex remodeling, characterized by depletion of the regulatory protein calstabin2, resulted in increased cytosolic Ca 2+ levels and mitochondrial oxidative stress. We also report a functional interplay between amyloid β (Aβ), β-adrenergic signaling, and altered Ca 2+ signaling via leaky RyR2 channels. Thus, post-translational modifications of RyR occur downstream of Aβ through a β2-adrenergic signaling cascade that activates PKA. RyR2 remodeling in turn enhances βAPP processing. Importantly, pharmacological stabilization of the binding of calstabin2 to RyR2 channels, which prevents Ca 2+ leakage, or blocking the β2-adrenergic signaling cascade reduced βAPP processing and the production of Aβ in APPswe-expressing SH-SY5Y cells. We conclude that targeting RyR-mediated Ca 2+ leakage may be a therapeutic approach to treat AD. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  18. Tax-1 and Tax-2 similarities and differences: focus on post-translational modifications and NF-κB activation

    Science.gov (United States)

    Shirinian, Margret; Kfoury, Youmna; Dassouki, Zeina; El-Hajj, Hiba; Bazarbachi, Ali

    2013-01-01

    Although human T cell leukemia virus type 1 and 2 (HTLV-1 and HTLV-2) share similar genetic organization, they have major differences in their pathogenesis and disease manifestation. HTLV-1 is capable of transforming T lymphocytes in infected patients resulting in adult T cell leukemia/lymphoma whereas HTLV-2 is not clearly associated with lymphoproliferative diseases. Numerous studies have provided accumulating evidence on the involvement of the viral transactivators Tax-1 versus Tax-2 in T cell transformation. Tax-1 is a potent transcriptional activator of both viral and cellular genes. Tax-1 post-translational modifications and specifically ubiquitylation and SUMOylation have been implicated in nuclear factor-kappaB (NF-κB) activation and may contribute to its transformation capacity. Although Tax-2 has similar protein structure compared to Tax-1, the two proteins display differences both in their protein–protein interaction and activation of signal transduction pathways. Recent studies on Tax-2 have suggested ubiquitylation and SUMOylation independent mechanisms of NF-κB activation. In this present review, structural and functional differences between Tax-1 and Tax-2 will be summarized. Specifically, we will address their subcellular localization, nuclear trafficking and their effect on cellular regulatory proteins. A special attention will be given to Tax-1/Tax-2 post-translational modification such as ubiquitylation, SUMOylation, phosphorylation, acetylation, NF-κB activation, and protein–protein interactions involved in oncogenecity both in vivo and in vitro. PMID:23966989

  19. Post-translational modification of ribosomally synthesized peptides by a radical SAM epimerase in Bacillus subtilis

    Science.gov (United States)

    Benjdia, Alhosna; Guillot, Alain; Ruffié, Pauline; Leprince, Jérôme; Berteau, Olivier

    2017-07-01

    Ribosomally synthesized peptides are built out of L-amino acids, whereas D-amino acids are generally the hallmark of non-ribosomal synthetic processes. Here we show that the model bacterium Bacillus subtilis is able to produce a novel type of ribosomally synthesized and post-translationally modified peptide that contains D-amino acids, and which we propose to call epipeptides. We demonstrate that a two [4Fe-4S]-cluster radical S-adenosyl-L-methionine (SAM) enzyme converts L-amino acids into their D-counterparts by catalysing Cα-hydrogen-atom abstraction and using a critical cysteine residue as the hydrogen-atom donor. Unexpectedly, these D-amino acid residues proved to be essential for the activity of a peptide that induces the expression of LiaRS, a major component of the bacterial cell envelope stress-response system. Present in B. subtilis and in several members of the human microbiome, these epipeptides and radical SAM epimerases broaden the landscape of peptidyl structures accessible to living organisms.

  20. Radiation-induced alterations of histone post-translational modification levels in lymphoblastoid cell lines

    International Nuclear Information System (INIS)

    Maroschik, Belinda; Gürtler, Anne; Krämer, Anne; Rößler, Ute; Gomolka, Maria; Hornhardt, Sabine; Mörtl, Simone; Friedl, Anna A

    2014-01-01

    Radiation-induced alterations in posttranslational histone modifications (PTMs) may affect the cellular response to radiation damage in the DNA. If not reverted appropriately, altered PTM patterns may cause long-term alterations in gene expression regulation and thus lead to cancer. It is therefore important to characterize radiation-induced alterations in PTM patterns and the factors affecting them. A lymphoblastoid cell line established from a normal donor was used to screen for alterations in methylation levels at H3K4, H3K9, H3K27, and H4K20, as well as acetylation at H3K9, H3K56, H4K5, and H4K16, by quantitative Western Blot analysis at 15 min, 1 h and 24 h after irradiation with 2 Gy and 10 Gy. The variability of alterations in acetylation marks was in addition investigated in a panel of lymphoblastoid cell lines with differing radiosensitivity established from lung cancer patients. The screening procedure demonstrated consistent hypomethylation at H3K4me3 and hypoacetylation at all acetylation marks tested. In the panel of lymphoblastoid cell lines, however, a high degree of inter-individual variability became apparent. Radiosensitive cell lines showed more pronounced and longer lasting H4K16 hypoacetylation than radioresistant lines, which correlates with higher levels of residual γ-H2AX foci after 24 h. So far, the factors affecting extent and duration of radiation-induced histone alterations are poorly defined. The present work hints at a high degree of inter-individual variability and a potential correlation of DNA damage repair capacity and alterations in PTM levels

  1. Salivary Cystatins: Exploring New Post-Translational Modifications and Polymorphisms by Top-Down High-Resolution Mass Spectrometry.

    Science.gov (United States)

    Manconi, Barbara; Liori, Barbara; Cabras, Tiziana; Vincenzoni, Federica; Iavarone, Federica; Castagnola, Massimo; Messana, Irene; Olianas, Alessandra

    2017-11-03

    Cystatins are a complex family of cysteine peptidase inhibitors. In the present study, various proteoforms of cystatin A, cystatin B, cystatin S, cystatin SN, and cystatin SA were detected in the acid-soluble fraction of human saliva and characterized by a top-down HPLC-ESI-MS approach. Proteoforms of cystatin D were also detected and characterized by an integrated top-down and bottom-up strategy. The proteoforms derive from coding sequence polymorphisms and post-translational modifications, in particular, phosphorylation, N-terminal processing, and oxidation. This study increases the current knowledge of salivary cystatin proteoforms and provides the basis to evaluate possible qualitative/quantitative variations of these proteoforms in different pathological states and reveal new potential salivary biomarkers of disease. Data are available via ProteomeXchange with identifier PXD007170.

  2. Post-translational modification of osteopontin: Effects on in vitro hydroxyapatite formation and growth

    DEFF Research Database (Denmark)

    Boskey, Adele L.; Christensen, Brian Søndergaard; Taleb, Hayat

    2012-01-01

    The manuscript tests the hypothesis that posttranslational modification of the SIBLING family of proteins in general and osteopontin in particular modify the abilities of these proteins to regulate in vitro hydroxyapatite (HA) formation. Osteopontin has diverse effects on hydroxyapatite (HA...

  3. Role of post-translational modifications at the β-subunit ectodomain in complex association with a promiscuous plant P4-ATPase

    DEFF Research Database (Denmark)

    Costa, Sara; Marek, Magdalena; Axelsen, Kristian Buhl

    2016-01-01

    and can interact with several isoforms. In the present study, we used a site-directed mutagenesis approach to assess the role of post-translational modifications at the plant ALIS5 β-subunit ectodomain in the functionality of the promiscuous plant P4-ATPase ALA2. We identified two N-glycosylated residues......) compromises complex association, but the mutant β-subunits still promote complex trafficking and activity to some extent. In contrast, disruption of a conserved disulfide bond between Cys(158) and Cys(172) has no effect on the P4-ATPase complex. Our results demonstrate that post-translational modifications...

  4. Tyrosine sulfation, a post-translational modification of microvillar enzymes in the small intestinal enterocyte

    DEFF Research Database (Denmark)

    Danielsen, E M

    1987-01-01

    Protein sulfation in small intestinal epithelial cells was studied by labelling of organ cultured mucosal explants with [35S]-sulfate. Six bands in SDS-PAGE became selectively labelled; four, of 250, 200, 166 and 130 kd, were membrane-bound and two, of 75 and 60 kd, were soluble. The sulfated mem...... sulfated. Most if not all the sulfate was bound to tyrosine residues rather than to the carbohydrate of the microvillar enzymes, showing that this type of modification can occur on plasma membrane proteins as well as on secretory proteins....

  5. The measurement of reversible redox dependent post-translational modifications and their regulation of mitochondrial and skeletal muscle function

    Directory of Open Access Journals (Sweden)

    Philip A Kramer

    2015-11-01

    Full Text Available Mitochondrial oxidative stress is a common feature of skeletal myopathies across multiple conditions; however, the mechanism by which it contributes to skeletal muscle dysfunction remains controversial. Oxidative damage to proteins, lipids, and DNA has received the most attention, yet an important role for reversible redox post-translational modifications (PTMs in pathophysiology is emerging. The possibility that these PTMs can exert dynamic control of muscle function implicates them as a mechanism contributing to skeletal muscle dysfunction in chronic disease. Herein, we discuss the significance of thiol-based redox dependent modifications to mitochondrial, myofibrillar and excitation-contraction (EC coupling proteins with an emphasis on how these changes could alter skeletal muscle performance under chronically stressed conditions. A major barrier to a better mechanistic understanding of the role of reversible redox PTMs in muscle function is the technical challenges associated with accurately measuring the changes of site-specific redox PTMs. Here we will critically review current approaches with an emphasis on sample preparation artifacts, quantitation, and specificity. Despite these challenges, the ability to accurately quantify reversible redox PTMs is critical to understanding the mechanisms by which mitochondrial oxidative stress contributes to skeletal muscle dysfunction in chronic diseases.

  6. RPA-coated single-stranded DNA as a platform for post-translational modifications in the DNA damage response.

    Science.gov (United States)

    Maréchal, Alexandre; Zou, Lee

    2015-01-01

    The Replication Protein A (RPA) complex is an essential regulator of eukaryotic DNA metabolism. RPA avidly binds to single-stranded DNA (ssDNA) through multiple oligonucleotide/oligosaccharide-binding folds and coordinates the recruitment and exchange of genome maintenance factors to regulate DNA replication, recombination and repair. The RPA-ssDNA platform also constitutes a key physiological signal which activates the master ATR kinase to protect and repair stalled or collapsed replication forks during replication stress. In recent years, the RPA complex has emerged as a key target and an important regulator of post-translational modifications in response to DNA damage, which is critical for its genome guardian functions. Phosphorylation and SUMOylation of the RPA complex, and more recently RPA-regulated ubiquitination, have all been shown to control specific aspects of DNA damage signaling and repair by modulating the interactions between RPA and its partners. Here, we review our current understanding of the critical functions of the RPA-ssDNA platform in the maintenance of genome stability and its regulation through an elaborate network of covalent modifications.

  7. The Measurement of Reversible Redox Dependent Post-translational Modifications and Their Regulation of Mitochondrial and Skeletal Muscle Function

    Energy Technology Data Exchange (ETDEWEB)

    Kramer, Philip A.; Duan, Jicheng; Qian, Wei-Jun; Marcinek, David J.

    2015-11-25

    Mitochondrial oxidative stress is a common feature of skeletal myopathies across multiple conditions; however, the mechanism by which it contributes to skeletal muscle dysfunction remains controversial. Oxidative damage to proteins, lipids, and DNA has received the most attention, yet an important role for reversible redox post-translational modifications (PTMs) in pathophysiology is emerging. The possibility that these PTMs can exert dynamic control of muscle function implicates them as a mechanism contributing to skeletal muscle dysfunction in chronic disease. Herein, we discuss the significance of thiol-based redox dependent modifications to mitochondrial, myofibrillar and excitation-contraction (EC) coupling proteins with an emphasis on how these changes could alter skeletal muscle performance under chronically stressed conditions. A major barrier to a better mechanistic understanding of the role of reversible redox PTMs in muscle function is the technical challenges associated with accurately measuring the changes of site-specific redox PTMs. Here we will critically review current approaches with an emphasis on sample preparation artifacts, quantitation, and specificity. Despite these challenges, the ability to accurately quantify reversible redox PTMs is critical to understanding the mechanisms by which mitochondrial oxidative stress contributes to skeletal muscle dysfunction in chronic diseases.

  8. Tax-1 and Tax-2 similarities and differences: Focus on post-translational modifications and NF-кB activation

    Directory of Open Access Journals (Sweden)

    Margret eShirinian

    2013-08-01

    Full Text Available ABSTRACTAlthough human T-cell leukemia virus type 1 and 2 (HTLV-1 and HTLV-2 share similar genetic organization, they have major differences in their pathogenesis and disease manifestation. HTLV-1 is capable of transforming T lymphocytes in infected patients and subsequently leads to adult T cell leukemia/lymphoma (ATL whereas HTLV-2 is not clearly associated with lymphoproliferative diseases. Numerous studies have provided accumulating evidence on the involvement of the viral transactivators Tax-1 versus Tax-2 in T cell transformation. Tax-1 is a potent transcriptional activator of both viral and cellular genes. Tax-1 posttranslational modifications and specifically ubiquitylation and SUMOylation have been implicated in NF-кB activation and may contribute to its transformation capacity. Although Tax-2 has similar protein structure compared to Tax-1, the two proteins display differences both in their protein-protein interaction and activation of signal transduction pathways. Recent studies on Tax-2 have suggested ubiquitylation and SUMOylation independent mechanisms of NF-кB activation. In this present review, structural and functional differences between Tax-1 and Tax- 2 will be summarized. Specifically, we will address their subcellular localization, nuclear trafficking and their effect on cellular regulatory proteins. A special attention will be given to Tax-1/Tax-2 post-translational modification such as ubiquitylation, SUMOylation, phosphorylation, acetylation, NF-кB activation and protein-protein interactions involved in oncogenecity both in vivo and in vitro.

  9. Peptidomics of Peptic Digest of Selected Potato Tuber Proteins: Post-Translational Modifications and Limited Cleavage Specificity.

    Science.gov (United States)

    C K Rajendran, Subin R; Mason, Beth; Udenigwe, Chibuike C

    2016-03-23

    Bioinformatic tools are useful in predicting bioactive peptides from food proteins. This study was focused on using bioinformatics and peptidomics to evaluate the specificity of peptide release and post-translational modifications (PTMs) in a peptic digest of potato protein isolate. Peptides in the protein hydrolysate were identified by LC-MS/MS and subsequently aligned to their parent potato tuber proteins. Five major proteins were selected for further analysis, namely, lipoxygenase, α-1,4-glucan phosphorylase, annexin, patatin, and polyubiquitin, based on protein coverage, abundance, confidence levels, and function. Comparison of the in silico peptide profile generated with ExPASy PeptideCutter and experimental peptidomics data revealed several differences. The experimental peptic cleavage sites were found to vary in number and specificity from PeptideCutter predictions. Average peptide chain length was also found to be higher than predicted with hexapeptides as the smallest detected peptides. Moreover, PTMs, particularly Met oxidation and Glu/Asp deamidation, were observed in some peptides, and these were unaccounted for during in silico analysis. PTMs can be formed during aging of potato tubers, or as a result of processing conditions during protein isolation and hydrolysis. The findings provide insights on the limitations of current bioinformatics tools for predicting bioactive peptide release from proteins, and on the existence of structural modifications that can alter the peptide bioactivity and functionality.

  10. Basolateral cholesterol depletion alters Aquaporin-2 post-translational modifications and disrupts apical plasma membrane targeting.

    Science.gov (United States)

    Moeller, Hanne B; Fuglsang, Cecilia Hvitfeldt; Pedersen, Cecilie Nøhr; Fenton, Robert A

    2018-01-01

    Apical plasma membrane accumulation of the water channel Aquaporin-2 (AQP2) in kidney collecting duct principal cells is critical for body water homeostasis. Posttranslational modification (PTM) of AQP2 is important for regulating AQP2 trafficking. The aim of this study was to determine the role of cholesterol in regulation of AQP2 PTM and in apical plasma membrane targeting of AQP2. Cholesterol depletion from the basolateral plasma membrane of a collecting duct cell line (mpkCCD14) using methyl-beta-cyclodextrin (MBCD) increased AQP2 ubiquitylation. Forskolin, cAMP or dDAVP-mediated AQP2 phosphorylation at Ser269 (pS269-AQP2) was prevented by cholesterol depletion from the basolateral membrane. None of these effects on pS269-AQP2 were observed when cholesterol was depleted from the apical side of cells, or when MBCD was applied subsequent to dDAVP stimulation. Basolateral, but not apical, MBCD application prevented cAMP-induced apical plasma membrane accumulation of AQP2. These studies indicate that manipulation of the cholesterol content of the basolateral plasma membrane interferes with AQP2 PTM and subsequently regulated apical plasma membrane targeting of AQP2. Copyright © 2017 Elsevier Inc. All rights reserved.

  11. Post-translational modification of osteopontin: Effects on in vitro hydroxyapatite formation and growth

    International Nuclear Information System (INIS)

    Boskey, Adele L.; Christensen, Brian; Taleb, Hayat; Sørensen, Esben S.

    2012-01-01

    Highlights: ► Thrombin-cleaved fragments of milk-osteopontin effect hydroxyapatite formation differently. ► N- and C-terminal fragments promoted hydroxyapatite formation and growth. ► A central fragment inhibited hydroxyapatite formation and growth. ► Binding to collagen or hydroxyapatite seed crystals modified these effects. -- Abstract: The manuscript tests the hypothesis that posttranslational modification of the SIBLING family of proteins in general and osteopontin in particular modify the abilities of these proteins to regulate in vitro hydroxyapatite (HA) formation. Osteopontin has diverse effects on hydroxyapatite (HA) mineral crystallite formation and growth depending on the extent of phosphorylation. We hypothesized that different regions of full-length OPN would also have distinct effects on the mineralization process. Thrombin fragmentation of milk OPN (mOPN) was used to test this hypothesis. Three fragments were tested in a de novo HA formation assay; an N-terminal fragment (aa 1–147), a central fragment (aa 148–204) denoted SKK-fragment and a C-terminal fragment (aa 205–262). Compared to intact mOPN the C- and N-terminal fragments behaved comparably, promoting HA formation and growth, but the central SKK-fragment acted as a mineralization inhibitor. In a seeded growth experiment all fragments inhibited mineral proliferation, but the SKK-fragment was the most effective inhibitor. These effects, seen in HA-formation and seeded growth assays in a gelatin gel system and in a pH-stat experiment were lost when the protein or fragments were dephosphorylated. Effects of the fully phosphorylated protein and fragments were also altered in the presence of fibrillar collagen. The diverse effects can be explained in terms of the intrinsically disordered nature of OPN and its fragments which enable them to interact with their multiple partners.

  12. Post-translational modifications near the quinone binding site of mammalian complex I.

    Science.gov (United States)

    Carroll, Joe; Ding, Shujing; Fearnley, Ian M; Walker, John E

    2013-08-23

    Complex I (NADH:ubiquinone oxidoreductase) in mammalian mitochondria is an L-shaped assembly of 44 protein subunits with one arm buried in the inner membrane of the mitochondrion and the orthogonal arm protruding about 100 Å into the matrix. The protruding arm contains the binding sites for NADH, the primary acceptor of electrons flavin mononucleotide (FMN), and a chain of seven iron-sulfur clusters that carries the electrons one at a time from FMN to a coenzyme Q molecule bound in the vicinity of the junction between the two arms. In the structure of the closely related bacterial enzyme from Thermus thermophilus, the quinone is thought to bind in a tunnel that spans the interface between the two arms, with the quinone head group close to the terminal iron-sulfur cluster, N2. The tail of the bound quinone is thought to extend from the tunnel into the lipid bilayer. In the mammalian enzyme, it is likely that this tunnel involves three of the subunits of the complex, ND1, PSST, and the 49-kDa subunit. An arginine residue in the 49-kDa subunit is symmetrically dimethylated on the ω-N(G) and ω-N(G') nitrogen atoms of the guanidino group and is likely to be close to cluster N2 and to influence its properties. Another arginine residue in the PSST subunit is hydroxylated and probably lies near to the quinone. Both modifications are conserved in mammalian enzymes, and the former is additionally conserved in Pichia pastoris and Paracoccus denitrificans, suggesting that they are functionally significant.

  13. The membrane-topogenic vectorial behaviour of Nrf1 controls its post-translational modification and transactivation activity.

    Science.gov (United States)

    Zhang, Yiguo; Hayes, John D

    2013-01-01

    The integral membrane-bound Nrf1 transcription factor fulfils important functions in maintaining cellular homeostasis and organ integrity, but how it is controlled vectorially is unknown. Herein, creative use of Gal4-based reporter assays with protease protection assays (GRAPPA), and double fluorescence protease protection (dFPP), reveals that the membrane-topogenic vectorial behaviour of Nrf1 dictates its post-translational modification and transactivation activity. Nrf1 is integrated within endoplasmic reticulum (ER) membranes through its NHB1-associated TM1 in cooperation with other semihydrophobic amphipathic regions. The transactivation domains (TADs) of Nrf1, including its Asn/Ser/Thr-rich (NST) glycodomain, are transiently translocated into the ER lumen, where it is glycosylated in the presence of glucose to become a 120-kDa isoform. Thereafter, the NST-adjoining TADs are partially repartitioned out of membranes into the cyto/nucleoplasmic side, where Nrf1 is subject to deglycosylation and/or proteolysis to generate 95-kDa and 85-kDa isoforms. Therefore, the vectorial process of Nrf1 controls its target gene expression.

  14. Biosynthetic Tailoring of Microcin E492m: Post-Translational Modification Affords an Antibacterial Siderophore-Peptide Conjugate

    Science.gov (United States)

    Nolan, Elizabeth M.; Fischbach, Michael A.; Koglin, Alexander; Walsh, Christopher T.

    2008-01-01

    The present work reveals that four proteins, MceCDIJ, encoded by the MccE492 gene cluster are responsible for the remarkable post-translational tailoring of Microcin E492 (MccE492), an 84-residue protein toxin secreted by Klebsiella pneumonaie RYC492 that targets neighboring gram-negative species. This modification results in attachment of a linearized and monoglycosylated derivative of enterobactin, a nonribosomal peptide and iron scavenger (siderophore), to the MccE492m C-terminus. MceC and MceD derivatize enterobactin by C-glycosylation at the C5 position of a N-(2,3-dihydroxybenzoyl) serine (DHB-Ser) moiety and regiospecific hydrolysis of an ester linkage in the trilactone scaffold, respectively. MceI and MceJ form a protein complex that attaches C-glycosylated enterobactins to the C-terminal serine residue of both aC10 model peptide and full-length MccE492. In the enzymatic product, the terminal serine residue is covalently attached to the C4′ oxygen of the glucose moiety. Non-enzymatic and base-catalyzed migration of the peptide to the C6′ position affords the C6′ glycosyl ester linkage observed in the mature toxin, MccE492m, isolated from bacterial cultures. PMID:17973380

  15. Predicting Post-Translational Modifications from Local Sequence Fragments Using Machine Learning Algorithms: Overview and Best Practices.

    Science.gov (United States)

    Tatjewski, Marcin; Kierczak, Marcin; Plewczynski, Dariusz

    2017-01-01

    Here, we present two perspectives on the task of predicting post translational modifications (PTMs) from local sequence fragments using machine learning algorithms. The first is the description of the fundamental steps required to construct a PTM predictor from the very beginning. These steps include data gathering, feature extraction, or machine-learning classifier selection. The second part of our work contains the detailed discussion of more advanced problems which are encountered in PTM prediction task. Probably the most challenging issues which we have covered here are: (1) how to address the training data class imbalance problem (we also present statistics describing the problem); (2) how to properly set up cross-validation folds with an approach which takes into account the homology of protein data records, to address this problem we present our folds-over-clusters algorithm; and (3) how to efficiently reach for new sources of learning features. Presented techniques and notes resulted from intense studies in the field, performed by our and other groups, and can be useful both for researchers beginning in the field of PTM prediction and for those who want to extend the repertoire of their research techniques.

  16. Identification of a Post-translational Modification with Ribitol-Phosphate and Its Defect in Muscular Dystrophy

    Directory of Open Access Journals (Sweden)

    Motoi Kanagawa

    2016-03-01

    Full Text Available Glycosylation is an essential post-translational modification that underlies many biological processes and diseases. α-dystroglycan (α-DG is a receptor for matrix and synaptic proteins that causes muscular dystrophy and lissencephaly upon its abnormal glycosylation (α-dystroglycanopathies. Here we identify the glycan unit ribitol 5-phosphate (Rbo5P, a phosphoric ester of pentose alcohol, in α-DG. Rbo5P forms a tandem repeat and functions as a scaffold for the formation of the ligand-binding moiety. We show that enzyme activities of three major α-dystroglycanopathy-causing proteins are involved in the synthesis of tandem Rbo5P. Isoprenoid synthase domain-containing (ISPD is cytidine diphosphate ribitol (CDP-Rbo synthase. Fukutin and fukutin-related protein are sequentially acting Rbo5P transferases that use CDP-Rbo. Consequently, Rbo5P glycosylation is defective in α-dystroglycanopathy models. Supplementation of CDP-Rbo to ISPD-deficient cells restored α-DG glycosylation. These findings establish the molecular basis of mammalian Rbo5P glycosylation and provide insight into pathogenesis and therapeutic strategies in α-DG-associated diseases.

  17. Notch-mediated post-translational control of Ngn3 protein stability regulates pancreatic patterning and cell fate commitment

    DEFF Research Database (Denmark)

    Qu, Xiaoling; Afelik, Solomon; Jensen, Jan Nygaard

    2013-01-01

    of ducts. On one hand, Ngn3 cell-intrinsically activates endocrine target genes; on the other, Ngn3 cell-extrinsically promotes lateral signaling via the Dll1>Notch>Hes1 pathway which substantially limits its ability to sustain endocrine formation. Prior to endocrine commitment, the Ngn3-mediated...... involves transcriptional repression as previously shown, but also incorporates a novel post-translational mechanism. In addition to its ability to promote endocrine fate, we provide evidence of a competing ability of Ngn3 in the patterning of multipotent progenitor cells in turn controlling the formation...

  18. PhosphOrtholog: a web-based tool for cross-species mapping of orthologous protein post-translational modifications.

    Science.gov (United States)

    Chaudhuri, Rima; Sadrieh, Arash; Hoffman, Nolan J; Parker, Benjamin L; Humphrey, Sean J; Stöckli, Jacqueline; Hill, Adam P; James, David E; Yang, Jean Yee Hwa

    2015-08-19

    Most biological processes are influenced by protein post-translational modifications (PTMs). Identifying novel PTM sites in different organisms, including humans and model organisms, has expedited our understanding of key signal transduction mechanisms. However, with increasing availability of deep, quantitative datasets in diverse species, there is a growing need for tools to facilitate cross-species comparison of PTM data. This is particularly important because functionally important modification sites are more likely to be evolutionarily conserved; yet cross-species comparison of PTMs is difficult since they often lie in structurally disordered protein domains. Current tools that address this can only map known PTMs between species based on known orthologous phosphosites, and do not enable the cross-species mapping of newly identified modification sites. Here, we addressed this by developing a web-based software tool, PhosphOrtholog ( www.phosphortholog.com ) that accurately maps protein modification sites between different species. This facilitates the comparison of datasets derived from multiple species, and should be a valuable tool for the proteomics community. Here we describe PhosphOrtholog, a web-based application for mapping known and novel orthologous PTM sites from experimental data obtained from different species. PhosphOrtholog is the only generic and automated tool that enables cross-species comparison of large-scale PTM datasets without relying on existing PTM databases. This is achieved through pairwise sequence alignment of orthologous protein residues. To demonstrate its utility we apply it to two sets of human and rat muscle phosphoproteomes generated following insulin and exercise stimulation, respectively, and one publicly available mouse phosphoproteome following cellular stress revealing high mapping and coverage efficiency. Although coverage statistics are dataset dependent, PhosphOrtholog increased the number of cross-species mapped sites

  19. Tandem affinity purification of histones, coupled to mass spectrometry, identifies associated proteins and new sites of post-translational modification in Saccharomyces cerevisiae.

    Science.gov (United States)

    Valero, M Luz; Sendra, Ramon; Pamblanco, Mercè

    2016-03-16

    Histones and their post-translational modifications contribute to regulating fundamental biological processes in all eukaryotic cells. We have applied a conventional tandem affinity purification strategy to histones H3 and H4 of the yeast Saccharomyces cerevisiae. Mass spectrometry analysis of the co-purified proteins revealed multiple associated proteins, including core histones, which indicates that tagged histones may be incorporated to the nucleosome particle. Among the many other co-isolated proteins there are histone chaperones, elements of chromatin remodeling, of nucleosome assembly/disassembly, and of histone modification complexes. The histone chaperone Rtt106p, two members of chromatin assembly FACT complex and Psh1p, an ubiquitin ligase, were the most abundant proteins obtained with both H3-TAP and H4-TAP, regardless of the cell extraction medium stringency. Our mass spectrometry analyses have also revealed numerous novel post-translational modifications, including 30 new chemical modifications in histones, mainly by ubiquitination. We have discovered not only new sites of ubiquitination but that, besides lysine, also serine and threonine residues are targets of ubiquitination on yeast histones. Our results show the standard tandem affinity purification procedure is suitable for application to yeast histones, in order to isolate and characterize histone-binding proteins and post-translational modifications, avoiding the bias caused by histone purification from a chromatin-enriched fraction. Copyright © 2016 Elsevier B.V. All rights reserved.

  20. Functional anthology of intrinsic disorder. 3. Ligands, post-translational modifications, and diseases associated with intrinsically disordered proteins.

    Science.gov (United States)

    Xie, Hongbo; Vucetic, Slobodan; Iakoucheva, Lilia M; Oldfield, Christopher J; Dunker, A Keith; Obradovic, Zoran; Uversky, Vladimir N

    2007-05-01

    devoted to the presentation of 87 Swiss-Prot keywords attributed to the cellular components, domains, technical terms, developmental processes, and coding sequence diversities possessing strong positive and negative correlation with long disordered regions (Vucetic, S.; Xie, H.; Iakoucheva, L. M.; Oldfield, C. J.; Dunker, A. K.; Obradovic, Z.; Uversky, V. N. Functional anthology of intrinsic disorder. 2. Cellular components, domains, technical terms, developmental processes, and coding sequence diversities correlated with long disordered regions. J. Proteome Res. 2007, 5, 1899-1916). Protein structure and functionality can be modulated by various post-translational modifications or/and as a result of binding of specific ligands. Numerous human diseases are associated with protein misfolding/misassembly/misfunctioning. This work concludes the series of papers dedicated to the functional anthology of intrinsic disorder and describes approximately 80 Swiss-Prot functional keywords that are related to ligands, post-translational modifications, and diseases possessing strong positive or negative correlation with the predicted long disordered regions in proteins.

  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. Investigation and identification of functional post-translational modification sites associated with drug binding and protein-protein interactions.

    Science.gov (United States)

    Su, Min-Gang; Weng, Julia Tzu-Ya; Hsu, Justin Bo-Kai; Huang, Kai-Yao; Chi, Yu-Hsiang; Lee, Tzong-Yi

    2017-12-21

    Protein post-translational modification (PTM) plays an essential role in various cellular processes that modulates the physical and chemical properties, folding, conformation, stability and activity of proteins, thereby modifying the functions of proteins. The improved throughput of mass spectrometry (MS) or MS/MS technology has not only brought about a surge in proteome-scale studies, but also contributed to a fruitful list of identified PTMs. However, with the increase in the number of identified PTMs, perhaps the more crucial question is what kind of biological mechanisms these PTMs are involved in. This is particularly important in light of the fact that most protein-based pharmaceuticals deliver their therapeutic effects through some form of PTM. Yet, our understanding is still limited with respect to the local effects and frequency of PTM sites near pharmaceutical binding sites and the interfaces of protein-protein interaction (PPI). Understanding PTM's function is critical to our ability to manipulate the biological mechanisms of protein. In this study, to understand the regulation of protein functions by PTMs, we mapped 25,835 PTM sites to proteins with available three-dimensional (3D) structural information in the Protein Data Bank (PDB), including 1785 modified PTM sites on the 3D structure. Based on the acquired structural PTM sites, we proposed to use five properties for the structural characterization of PTM substrate sites: the spatial composition of amino acids, residues and side-chain orientations surrounding the PTM substrate sites, as well as the secondary structure, division of acidity and alkaline residues, and solvent-accessible surface area. We further mapped the structural PTM sites to the structures of drug binding and PPI sites, identifying a total of 1917 PTM sites that may affect PPI and 3951 PTM sites associated with drug-target binding. An integrated analytical platform (CruxPTM), with a variety of methods and online molecular docking

  3. Mass spectrometry analysis of the variants of histone H3 and H4 of soybean and their post-translational modifications

    Directory of Open Access Journals (Sweden)

    Lam Hon-Ming

    2009-07-01

    Full Text Available Abstract Background Histone modifications and histone variants are of importance in many biological processes. To understand the biological functions of the global dynamics of histone modifications and histone variants in higher plants, we elucidated the variants and post-translational modifications of histones in soybean, a legume plant with a much bigger genome than that of Arabidopsis thaliana. Results In soybean leaves, mono-, di- and tri-methylation at Lysine 4, Lysine 27 and Lysine 36, and acetylation at Lysine 14, 18 and 23 were detected in HISTONE H3. Lysine 27 was prone to being mono-methylated, while tri-methylation was predominant at Lysine 36. We also observed that Lysine 27 methylation and Lysine 36 methylation usually excluded each other in HISTONE H3. Although methylation at HISTONE H3 Lysine 79 was not reported in A. thaliana, mono- and di-methylated HISTONE H3 Lysine 79 were detected in soybean. Besides, acetylation at Lysine 8 and 12 of HISTONE H4 in soybean were identified. Using a combination of mass spectrometry and nano-liquid chromatography, two variants of HISTONE H3 were detected and their modifications were determined. They were different at positions of A31F41S87S90 (HISTONE variant H3.1 and T31Y41H87L90 (HISTONE variant H3.2, respectively. The methylation patterns in these two HISTONE H3 variants also exhibited differences. Lysine 4 and Lysine 36 methylation were only detected in HISTONE H3.2, suggesting that HISTONE variant H3.2 might be associated with actively transcribing genes. In addition, two variants of histone H4 (H4.1 and H4.2 were also detected, which were missing in other organisms. In the histone variant H4.1 and H4.2, the amino acid 60 was isoleucine and valine, respectively. Conclusion This work revealed several distinct variants of soybean histone and their modifications that were different from A. thaliana, thus providing important biological information toward further understanding of the histone

  4. Tissue-specific expression and post-translational modifications of plant- and bacterial-type phosphoenolpyruvate carboxylase isozymes of the castor oil plant, Ricinus communis L.

    Science.gov (United States)

    O’Leary, Brendan; Fedosejevs, Eric T.; Hill, Allyson T.; Bettridge, James; Park, Joonho; Rao, Srinath K.; Leach, Craig A.; Plaxton, William C.

    2011-01-01

    This study employs transcript profiling together with immunoblotting and co-immunopurification to assess the tissue-specific expression, protein:protein interactions, and post-translational modifications (PTMs) of plant- and bacterial-type phosphoenolpyruvate carboxylase (PEPC) isozymes (PTPC and BTPC, respectively) in the castor plant, Ricinus communis. Previous studies established that the Class-1 PEPC (PTPC homotetramer) of castor oil seeds (COS) is activated by phosphorylation at Ser-11 and inhibited by monoubiquitination at Lys-628 during endosperm development and germination, respectively. Elimination of photosynthate supply to developing COS by depodding caused the PTPC of the endosperm and cotyledon to be dephosphorylated, and then subsequently monoubiquitinated in vivo. PTPC monoubiquitination rather than phosphorylation is widespread throughout the castor plant and appears to be the predominant PTM of Class-1 PEPC that occurs in planta. The distinctive developmental patterns of PTPC phosphorylation versus monoubiquitination indicates that these two PTMs are mutually exclusive. By contrast, the BTPC: (i) is abundant in the inner integument, cotyledon, and endosperm of developing COS, but occurs at low levels in roots and cotyledons of germinated COS, (ii) shows a unique developmental pattern in leaves such that it is present in leaf buds and young expanding leaves, but undetectable in fully expanded leaves, and (iii) tightly interacts with co-expressed PTPC to form the novel and allosterically-desensitized Class-2 PEPC heteromeric complex. BTPC and thus Class-2 PEPC up-regulation appears to be a distinctive feature of rapidly growing and/or biosynthetically active tissues that require a large anaplerotic flux from phosphoenolpyruvate to replenish tricarboxylic acid cycle C-skeletons being withdrawn for anabolism. PMID:21841182

  5. 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.

  6. Tandem Affinity Purification Approach Coupled to Mass Spectrometry to Identify Post-translational Modifications of Histones Associated with Chromatin-Binding Proteins.

    Science.gov (United States)

    Beyer, Sophie; Robin, Philippe; Ait-Si-Ali, Slimane

    2017-01-01

    Protein purification by tandem affinity purification (TAP)-tag coupled to mass spectrometry analysis is usually used to reveal protein complex composition. Here we describe a TAP-tag purification of chromatin-bound proteins along with associated nucleosomes, which allow exhaustive identification of protein partners. Moreover, this method allows exhaustive identification of the post-translational modifications (PTMs) of the associated histones. Thus, in addition to partner characterization, this approach reveals the associated epigenetic landscape that can shed light on the function and properties of the studied chromatin-bound protein.

  7. Mitochondrial Reactive Oxygen Species in Lipotoxic Hearts Induce Post-Translational Modifications of AKAP121, DRP1, and OPA1 That Promote Mitochondrial Fission.

    Science.gov (United States)

    Tsushima, Kensuke; Bugger, Heiko; Wende, Adam R; Soto, Jamie; Jenson, Gregory A; Tor, Austin R; McGlauflin, Rose; Kenny, Helena C; Zhang, Yuan; Souvenir, Rhonda; Hu, Xiao X; Sloan, Crystal L; Pereira, Renata O; Lira, Vitor A; Spitzer, Kenneth W; Sharp, Terry L; Shoghi, Kooresh I; Sparagna, Genevieve C; Rog-Zielinska, Eva A; Kohl, Peter; Khalimonchuk, Oleh; Schaffer, Jean E; Abel, E Dale

    2018-01-05

    Cardiac lipotoxicity, characterized by increased uptake, oxidation, and accumulation of lipid intermediates, contributes to cardiac dysfunction in obesity and diabetes mellitus. However, mechanisms linking lipid overload and mitochondrial dysfunction are incompletely understood. To elucidate the mechanisms for mitochondrial adaptations to lipid overload in postnatal hearts in vivo. Using a transgenic mouse model of cardiac lipotoxicity overexpressing ACSL1 (long-chain acyl-CoA synthetase 1) in cardiomyocytes, we show that modestly increased myocardial fatty acid uptake leads to mitochondrial structural remodeling with significant reduction in minimum diameter. This is associated with increased palmitoyl-carnitine oxidation and increased reactive oxygen species (ROS) generation in isolated mitochondria. Mitochondrial morphological changes and elevated ROS generation are also observed in palmitate-treated neonatal rat ventricular cardiomyocytes. Palmitate exposure to neonatal rat ventricular cardiomyocytes initially activates mitochondrial respiration, coupled with increased mitochondrial polarization and ATP synthesis. However, long-term exposure to palmitate (>8 hours) enhances ROS generation, which is accompanied by loss of the mitochondrial reticulum and a pattern suggesting increased mitochondrial fission. Mechanistically, lipid-induced changes in mitochondrial redox status increased mitochondrial fission by increased ubiquitination of AKAP121 (A-kinase anchor protein 121) leading to reduced phosphorylation of DRP1 (dynamin-related protein 1) at Ser637 and altered proteolytic processing of OPA1 (optic atrophy 1). Scavenging mitochondrial ROS restored mitochondrial morphology in vivo and in vitro. Our results reveal a molecular mechanism by which lipid overload-induced mitochondrial ROS generation causes mitochondrial dysfunction by inducing post-translational modifications of mitochondrial proteins that regulate mitochondrial dynamics. These findings provide a

  8. Characterizing the Range of Extracellular Protein Post-Translational Modifications in a Cellulose-Degrading Bacteria Using a Multiple Proteolyic Digestion/Peptide Fragmentation Approach

    Energy Technology Data Exchange (ETDEWEB)

    Dykstra, Andrew B [ORNL; Rodriguez, Jr., Miguel [ORNL; Raman, Babu [Dow Chemical Company, The; Cook, Kelsey [ORNL; Hettich, Robert {Bob} L [ORNL

    2013-01-01

    Post-translational modifications (PTMs) are known to play a significant role in many biological functions. The focus of this study is to characterize the post-translational modifications of the cellulosome protein complex used by the bacterium Clostridium thermocellum to better understand how this protein machine is tuned for enzymatic cellulose solubilization. To enhance comprehensive characterization, the extracellular cellulosome proteins were analyzed using multiple proteolytic digests (trypsin, Lys-C, Glu-C) and multiple fragmentation techniques (collisionally-activated dissociation, electron transfer dissociation, decision tree). As expected, peptide and protein identifications were increased by utilizing alternate proteases and fragmentation methods, in addition to the increase in protein sequence coverage. The complementarity of these experiments also allowed for a global exploration of PTMs associated with the cellulosome based upon a set of defined PTMs that included methylation, oxidation, acetylation, phosphorylation, and signal peptide cleavage. In these experiments, 85 modified peptides corresponding to 28 cellulosome proteins were identified. Many of these modifications were located in active cellulolytic or structural domains of the cellulosome proteins, suggesting a level of possible regulatory control of protein function in various cellulotyic conditions. The use of multiple enzymes and fragmentation technologies allowed for independent verification of PTMs in different experiments, thus leading to increased confidence in PTM identifications.

  9. Role of post-translational modifications at the β-subunit ectodomain in complex association with a promiscuous plant P4-ATPase.

    Science.gov (United States)

    Costa, Sara R; Marek, Magdalena; Axelsen, Kristian B; Theorin, Lisa; Pomorski, Thomas G; López-Marqués, Rosa L

    2016-06-01

    P-type ATPases of subfamily IV (P4-ATPases) constitute a major group of phospholipid flippases that form heteromeric complexes with members of the Cdc50 (cell division control 50) protein family. Some P4-ATPases interact specifically with only one β-subunit isoform, whereas others are promiscuous and can interact with several isoforms. In the present study, we used a site-directed mutagenesis approach to assess the role of post-translational modifications at the plant ALIS5 β-subunit ectodomain in the functionality of the promiscuous plant P4-ATPase ALA2. We identified two N-glycosylated residues, Asn(181) and Asn(231) Whereas mutation of Asn(231) seems to have a small effect on P4-ATPase complex formation, mutation of evolutionarily conserved Asn(181) disrupts interaction between the two subunits. Of the four cysteine residues located in the ALIS5 ectodomain, mutation of Cys(86) and Cys(107) compromises complex association, but the mutant β-subunits still promote complex trafficking and activity to some extent. In contrast, disruption of a conserved disulfide bond between Cys(158) and Cys(172) has no effect on the P4-ATPase complex. Our results demonstrate that post-translational modifications in the β-subunit have different functional roles in different organisms, which may be related to the promiscuity of the P4-ATPase. © 2016 The Author(s). published by Portland Press Limited on behalf of the Biochemical Society.

  10. Extraction and Characterization of Extracellular Proteins and Their Post-Translational Modifications from Arabidopsis thaliana Suspension Cell Cultures and Seedlings: A Critical Review

    Directory of Open Access Journals (Sweden)

    Mina Ghahremani

    2016-09-01

    Full Text Available Proteins secreted by plant cells into the extracellular space, consisting of the cell wall, apoplastic fluid, and rhizosphere, play crucial roles during development, nutrient acquisition, and stress acclimation. However, isolating the full range of secreted proteins has proven difficult, and new strategies are constantly evolving to increase the number of proteins that can be detected and identified. In addition, the dynamic nature of the extracellular proteome presents the further challenge of identifying and characterizing the post-translational modifications (PTMs of secreted proteins, particularly glycosylation and phosphorylation. Such PTMs are common and important regulatory modifications of proteins, playing a key role in many biological processes. This review explores the most recent methods in isolating and characterizing the plant extracellular proteome with a focus on the model plant Arabidopsis thaliana, highlighting the current challenges yet to be overcome. Moreover, the crucial role of protein PTMs in cell wall signalling, development, and plant responses to biotic and abiotic stress is discussed.

  11. Modulations of DNA Contacts by Linker Histones and Post-translational Modifications Determine the Mobility and Modifiability of Nucleosomal H3 Tails.

    Science.gov (United States)

    Stützer, Alexandra; Liokatis, Stamatios; Kiesel, Anja; Schwarzer, Dirk; Sprangers, Remco; Söding, Johannes; Selenko, Philipp; Fischle, Wolfgang

    2016-01-21

    Post-translational histone modifications and linker histone incorporation regulate chromatin structure and genome activity. How these systems interface on a molecular level is unclear. Using biochemistry and NMR spectroscopy, we deduced mechanistic insights into the modification behavior of N-terminal histone H3 tails in different nucleosomal contexts. We find that linker histones generally inhibit modifications of different H3 sites and reduce H3 tail dynamics in nucleosomes. These effects are caused by modulations of electrostatic interactions of H3 tails with linker DNA and largely depend on the C-terminal domains of linker histones. In agreement, linker histone occupancy and H3 tail modifications segregate on a genome-wide level. Charge-modulating modifications such as phosphorylation and acetylation weaken transient H3 tail-linker DNA interactions, increase H3 tail dynamics, and, concomitantly, enhance general modifiability. We propose that alterations of H3 tail-linker DNA interactions by linker histones and charge-modulating modifications execute basal control mechanisms of chromatin function. Copyright © 2016 Elsevier Inc. All rights reserved.

  12. DbPTM 3.0: an informative resource for investigating substrate site specificity and functional association of protein post-translational modifications.

    Science.gov (United States)

    Lu, Cheng-Tsung; Huang, Kai-Yao; Su, Min-Gang; Lee, Tzong-Yi; Bretaña, Neil Arvin; Chang, Wen-Chi; Chen, Yi-Ju; Chen, Yu-Ju; Huang, Hsien-Da

    2013-01-01

    Protein modification is an extremely important post-translational regulation that adjusts the physical and chemical properties, conformation, stability and activity of a protein; thus altering protein function. Due to the high throughput of mass spectrometry (MS)-based methods in identifying site-specific post-translational modifications (PTMs), dbPTM (http://dbPTM.mbc.nctu.edu.tw/) is updated to integrate experimental PTMs obtained from public resources as well as manually curated MS/MS peptides associated with PTMs from research articles. Version 3.0 of dbPTM aims to be an informative resource for investigating the substrate specificity of PTM sites and functional association of PTMs between substrates and their interacting proteins. In order to investigate the substrate specificity for modification sites, a newly developed statistical method has been applied to identify the significant substrate motifs for each type of PTMs containing sufficient experimental data. According to the data statistics in dbPTM, >60% of PTM sites are located in the functional domains of proteins. It is known that most PTMs can create binding sites for specific protein-interaction domains that work together for cellular function. Thus, this update integrates protein-protein interaction and domain-domain interaction to determine the functional association of PTM sites located in protein-interacting domains. Additionally, the information of structural topologies on transmembrane (TM) proteins is integrated in dbPTM in order to delineate the structural correlation between the reported PTM sites and TM topologies. To facilitate the investigation of PTMs on TM proteins, the PTM substrate sites and the structural topology are graphically represented. Also, literature information related to PTMs, orthologous conservations and substrate motifs of PTMs are also provided in the resource. Finally, this version features an improved web interface to facilitate convenient access to the resource.

  13. Analysis of the post-translational modifications of the individual amino acids in lens proteins which were induced by aging and irradiation

    International Nuclear Information System (INIS)

    Fujii, Noriko; Kim, Ingu; Saito, Takeshi; Takata, Takumi

    2017-01-01

    The eye lens is a transparent organ that functions to focus light and images on the retina. The transparency and high refraction of the lens are maintained by the function of α-, β- and γ-crystallins. These long-lived proteins are subject to various post-translational modifications, such as oxidation, deamidation, truncation and isomerization, which occur gradually during the aging process. Such modifications, which are generated by UV light and oxidative stress, decrease crystallin solubility and lens transparency, and ultimately lead to the development of age-related cataracts. Here, we irradiated young rat lenses with γ-rays (5-500 Gy) and extracted the water-soluble (WS) and insoluble (WI) protein fractions. The WS and WI lens proteins were digested with trypsin, and the resulting peptides were analyzed by one-shot LC-MS/MS to determine the specific sites of oxidation of methionine and tryptophan, deamidation of asparagine and glutamine, and isomerization of aspartyl in rat α- and β-crystallins in the WS and WI fractions. Oxidation and deamidation occurred in several crystallins after irradiation at more than, respectively, 50 Gy and 5 Gy; however, isomerization did not occur in any crystallin even after exposure to 500 Gy of irradiation. The number of oxidation and deamidation sites was much higher in the WI than in the WS fraction. Furthermore, the oxidation and deamidation sites in rat crystallins resemble those reported in crystallins from human age-related cataracts. Thus, this study on post-translational modifications of crystallins induced by ionizing irradiation may provide useful information relevant to the formation of human age-related cataracts. (author)

  14. Abnormal Type I Collagen Post-translational Modification and Crosslinking in a Cyclophilin B KO Mouse Model of Recessive Osteogenesis Imperfecta

    Science.gov (United States)

    Cabral, Wayne A.; Perdivara, Irina; Weis, MaryAnn; Terajima, Masahiko; Blissett, Angela R.; Chang, Weizhong; Perosky, Joseph E.; Makareeva, Elena N.; Mertz, Edward L.; Leikin, Sergey; Tomer, Kenneth B.; Kozloff, Kenneth M.; Eyre, David R.; Yamauchi, Mitsuo; Marini, Joan C.

    2014-01-01

    Cyclophilin B (CyPB), encoded by PPIB, is an ER-resident peptidyl-prolyl cis-trans isomerase (PPIase) that functions independently and as a component of the collagen prolyl 3-hydroxylation complex. CyPB is proposed to be the major PPIase catalyzing the rate-limiting step in collagen folding. Mutations in PPIB cause recessively inherited osteogenesis imperfecta type IX, a moderately severe to lethal bone dysplasia. To investigate the role of CyPB in collagen folding and post-translational modifications, we generated Ppib−/− mice that recapitulate the OI phenotype. Knock-out (KO) mice are small, with reduced femoral areal bone mineral density (aBMD), bone volume per total volume (BV/TV) and mechanical properties, as well as increased femoral brittleness. Ppib transcripts are absent in skin, fibroblasts, femora and calvarial osteoblasts, and CyPB is absent from KO osteoblasts and fibroblasts on western blots. Only residual (2–11%) collagen prolyl 3-hydroxylation is detectable in KO cells and tissues. Collagen folds more slowly in the absence of CyPB, supporting its rate-limiting role in folding. However, treatment of KO cells with cyclosporine A causes further delay in folding, indicating the potential existence of another collagen PPIase. We confirmed and extended the reported role of CyPB in supporting collagen lysyl hydroxylase (LH1) activity. Ppib−/− fibroblast and osteoblast collagen has normal total lysyl hydroxylation, while increased collagen diglycosylation is observed. Liquid chromatography/mass spectrometry (LC/MS) analysis of bone and osteoblast type I collagen revealed site-specific alterations of helical lysine hydroxylation, in particular, significantly reduced hydroxylation of helical crosslinking residue K87. Consequently, underhydroxylated forms of di- and trivalent crosslinks are strikingly increased in KO bone, leading to increased total crosslinks and decreased helical hydroxylysine- to lysine-derived crosslink ratios. The altered

  15. Abnormal type I collagen post-translational modification and crosslinking in a cyclophilin B KO mouse model of recessive osteogenesis imperfecta.

    Directory of Open Access Journals (Sweden)

    Wayne A Cabral

    2014-06-01

    Full Text Available Cyclophilin B (CyPB, encoded by PPIB, is an ER-resident peptidyl-prolyl cis-trans isomerase (PPIase that functions independently and as a component of the collagen prolyl 3-hydroxylation complex. CyPB is proposed to be the major PPIase catalyzing the rate-limiting step in collagen folding. Mutations in PPIB cause recessively inherited osteogenesis imperfecta type IX, a moderately severe to lethal bone dysplasia. To investigate the role of CyPB in collagen folding and post-translational modifications, we generated Ppib-/- mice that recapitulate the OI phenotype. Knock-out (KO mice are small, with reduced femoral areal bone mineral density (aBMD, bone volume per total volume (BV/TV and mechanical properties, as well as increased femoral brittleness. Ppib transcripts are absent in skin, fibroblasts, femora and calvarial osteoblasts, and CyPB is absent from KO osteoblasts and fibroblasts on western blots. Only residual (2-11% collagen prolyl 3-hydroxylation is detectable in KO cells and tissues. Collagen folds more slowly in the absence of CyPB, supporting its rate-limiting role in folding. However, treatment of KO cells with cyclosporine A causes further delay in folding, indicating the potential existence of another collagen PPIase. We confirmed and extended the reported role of CyPB in supporting collagen lysyl hydroxylase (LH1 activity. Ppib-/- fibroblast and osteoblast collagen has normal total lysyl hydroxylation, while increased collagen diglycosylation is observed. Liquid chromatography/mass spectrometry (LC/MS analysis of bone and osteoblast type I collagen revealed site-specific alterations of helical lysine hydroxylation, in particular, significantly reduced hydroxylation of helical crosslinking residue K87. Consequently, underhydroxylated forms of di- and trivalent crosslinks are strikingly increased in KO bone, leading to increased total crosslinks and decreased helical hydroxylysine- to lysine-derived crosslink ratios. The altered

  16. Abnormal type I collagen post-translational modification and crosslinking in a cyclophilin B KO mouse model of recessive osteogenesis imperfecta.

    Science.gov (United States)

    Cabral, Wayne A; Perdivara, Irina; Weis, MaryAnn; Terajima, Masahiko; Blissett, Angela R; Chang, Weizhong; Perosky, Joseph E; Makareeva, Elena N; Mertz, Edward L; Leikin, Sergey; Tomer, Kenneth B; Kozloff, Kenneth M; Eyre, David R; Yamauchi, Mitsuo; Marini, Joan C

    2014-06-01

    Cyclophilin B (CyPB), encoded by PPIB, is an ER-resident peptidyl-prolyl cis-trans isomerase (PPIase) that functions independently and as a component of the collagen prolyl 3-hydroxylation complex. CyPB is proposed to be the major PPIase catalyzing the rate-limiting step in collagen folding. Mutations in PPIB cause recessively inherited osteogenesis imperfecta type IX, a moderately severe to lethal bone dysplasia. To investigate the role of CyPB in collagen folding and post-translational modifications, we generated Ppib-/- mice that recapitulate the OI phenotype. Knock-out (KO) mice are small, with reduced femoral areal bone mineral density (aBMD), bone volume per total volume (BV/TV) and mechanical properties, as well as increased femoral brittleness. Ppib transcripts are absent in skin, fibroblasts, femora and calvarial osteoblasts, and CyPB is absent from KO osteoblasts and fibroblasts on western blots. Only residual (2-11%) collagen prolyl 3-hydroxylation is detectable in KO cells and tissues. Collagen folds more slowly in the absence of CyPB, supporting its rate-limiting role in folding. However, treatment of KO cells with cyclosporine A causes further delay in folding, indicating the potential existence of another collagen PPIase. We confirmed and extended the reported role of CyPB in supporting collagen lysyl hydroxylase (LH1) activity. Ppib-/- fibroblast and osteoblast collagen has normal total lysyl hydroxylation, while increased collagen diglycosylation is observed. Liquid chromatography/mass spectrometry (LC/MS) analysis of bone and osteoblast type I collagen revealed site-specific alterations of helical lysine hydroxylation, in particular, significantly reduced hydroxylation of helical crosslinking residue K87. Consequently, underhydroxylated forms of di- and trivalent crosslinks are strikingly increased in KO bone, leading to increased total crosslinks and decreased helical hydroxylysine- to lysine-derived crosslink ratios. The altered crosslink

  17. Barley lipid transfer protein, LTP1, contains a new type of lipid-like post-translational modification

    DEFF Research Database (Denmark)

    Lindorff-Larsen, Kresten; Lerche, Mathilde H.; Poulsen, Flemming Martin

    2001-01-01

    in which an aspartic acid in LTP1 is bound to the modification through what most likely is an ester bond. The chemical structure of the modification has been characterized by means of two-dimensional homo- and heteronuclear nuclear magnetic resonance spectroscopy as well as mass spectrometry and is found...

  18. Biosynthesis of the D2-cell adhesion molecule: post-translational modifications, intracellular transport, and developmental changes

    DEFF Research Database (Denmark)

    Lyles, J M; Linnemann, D; Bock, E

    1984-01-01

    Posttranslational modifications and intracellular transport of the D2-cell adhesion molecule (D2-CAM) were examined in cultured fetal rat neuronal cells. Developmental changes in biosynthesis were studied in rat forebrain explant cultures. Two D2-CAM polypeptides with Mr of 187,000-210,000 (A...

  19. Profiling of Histone Post-Translational Modifications in Mouse Brain with High-Resolution Top-Down Mass Spectrometry

    Energy Technology Data Exchange (ETDEWEB)

    Zhou, Mowei; Paša-Tolić, Ljiljana; Stenoien, David L.

    2016-12-21

    Histones play central roles in most chromosomal functions and both their basic biology and roles in disease have been the subject of intense study. Since multiple PTMs along the entire protein sequence are potential regulators of histones, a top-down approach, where intact proteins are analyzed, is ultimately required for complete characterization of proteoforms. However, significant challenges remain for top-down histone analysis primarily because of deficiencies in separation/resolving power and effective identification algorithms. Here, we used state of the art mass spectrometry and a bioinformatics workflow for targeted data analysis and visualization. The workflow uses ProMex for intact mass deconvolution, MSPathFinder as search engine, and LcMsSpectator as a data visualization tool. ProMex sums across retention time to maximize sensitivity and accuracy for low abundance species in MS1deconvolution. MSPathFinder searches the MS2 data against protein sequence databases with user-defined modifications. LcMsSpectator presents the results from ProMex and MSPathFinder in a format that allows quick manual evaluation of critical attributes for high-confidence identifications. When complemented with the open-modification tool TopPIC, this workflow enabled identification of novel histone PTMs including tyrosine bromination on histone H4 and H2A, H3 glutathionylation, and mapping of conventional PTMs along the entire protein for many histone subunits.

  20. Native pyroglutamation of huwentoxin-IV: a post-translational modification that increases the trapping ability to the sodium channel.

    Science.gov (United States)

    Rong, Mingqiang; Duan, Zhigui; Chen, Juliang; Li, Jianglin; Xiao, Yuchen; Liang, Songping

    2013-01-01

    Huwentoxin-IV (HWTX-IV), a tetrodotoxin-sensitive (TTX-s) sodium channel antagonist, is found in the venom of the Chinese spider Ornithoctonus huwena. A naturally modified HWTX-IV (mHWTX-IV), having a molecular mass 18 Da lower than HWTX-IV, has also been isolated from the venom of the same spider. By a combination of enzymatic fragmentation and MS/MS de novo sequencing, mHWTX-IV has been shown to have the same amino acid sequence as that of HWTX-IV, except that the N-terminal glutamic acid replaced by pyroglutamic acid. mHWTX-IV inhibited tetrodotoxin-sensitive voltage-gated sodium channels of dorsal root ganglion neurons with an IC50 nearly equal to native HWTX-IV. mHWTX-IV showed the same activation and inactivation kinetics seen for native HWTX-IV. In contrast with HWTX-IV, which dissociates at moderate voltage depolarization voltages (+50 mV, 180000 ms), mHWTX-IV inhibition of TTX-sensitive sodium channels is not reversed by strong depolarization voltages (+200 mV, 500 ms). Recovery of Nav1.7current was voltage-dependent and was induced by extreme depolarization in the presence of HWTX-IV, but no obvious current was elicited after application of mHWTX-IV. Our data indicate that the N-terminal modification of HWTX-IV gives the peptide toxin a greater ability to trap the voltage sensor in the sodium channel. Loss of a negative charge, caused by cyclization at the N-terminus, is a possible reason why the modified toxin binds much stronger. To our knowledge, this is the first report of a pyroglutamic acid residue in a spider toxin; this modification seems to increase the trapping ability of the voltage sensor in the sodium channel.

  1. Native pyroglutamation of huwentoxin-IV: a post-translational modification that increases the trapping ability to the sodium channel.

    Directory of Open Access Journals (Sweden)

    Mingqiang Rong

    Full Text Available Huwentoxin-IV (HWTX-IV, a tetrodotoxin-sensitive (TTX-s sodium channel antagonist, is found in the venom of the Chinese spider Ornithoctonus huwena. A naturally modified HWTX-IV (mHWTX-IV, having a molecular mass 18 Da lower than HWTX-IV, has also been isolated from the venom of the same spider. By a combination of enzymatic fragmentation and MS/MS de novo sequencing, mHWTX-IV has been shown to have the same amino acid sequence as that of HWTX-IV, except that the N-terminal glutamic acid replaced by pyroglutamic acid. mHWTX-IV inhibited tetrodotoxin-sensitive voltage-gated sodium channels of dorsal root ganglion neurons with an IC50 nearly equal to native HWTX-IV. mHWTX-IV showed the same activation and inactivation kinetics seen for native HWTX-IV. In contrast with HWTX-IV, which dissociates at moderate voltage depolarization voltages (+50 mV, 180000 ms, mHWTX-IV inhibition of TTX-sensitive sodium channels is not reversed by strong depolarization voltages (+200 mV, 500 ms. Recovery of Nav1.7current was voltage-dependent and was induced by extreme depolarization in the presence of HWTX-IV, but no obvious current was elicited after application of mHWTX-IV. Our data indicate that the N-terminal modification of HWTX-IV gives the peptide toxin a greater ability to trap the voltage sensor in the sodium channel. Loss of a negative charge, caused by cyclization at the N-terminus, is a possible reason why the modified toxin binds much stronger. To our knowledge, this is the first report of a pyroglutamic acid residue in a spider toxin; this modification seems to increase the trapping ability of the voltage sensor in the sodium channel.

  2. Assigning Quantitative Function to Post-Translational Modifications Reveals Multiple Sites of Phosphorylation That Tune Yeast Pheromone Signaling Output

    Energy Technology Data Exchange (ETDEWEB)

    Pincus, David; Ryan, Christopher J.; Smith, Richard D.; Brent, Roger; Resnekov, Orna; Hakimi, Mohamed Ali

    2013-03-12

    Cell signaling systems transmit information by post-­translationally modifying signaling proteins, often via phosphorylation. While thousands of sites of phosphorylation have been identified in proteomic studies, the vast majority of sites have no known function. Assigning functional roles to the catalog of uncharacterized phosphorylation sites is a key research challenge. Here we present a general approach to address this challenge and apply it to a prototypical signaling pathway, the pheromone response pathway in Saccharomyces cerevisiae. The pheromone pathway includes a mitogen activated protein kinase (MAPK) cascade activated by a G-­protein coupled receptor (GPCR). We used mass spectrometry-based proteomics to identify sites whose phosphorylation changed when the system was active, and evolutionary conservation to assign priority to a list of candidate MAPK regulatory sites. We made targeted alterations in those sites, and measured the effects of the mutations on pheromone pathway output in single cells. Our work identified six new sites that quantitatively tuned system output. We developed simple computational models to find system architectures that recapitulated the quantitative phenotypes of the mutants. Our results identify a number of regulated phosphorylation events that contribute to adjust the input-­output relationship of this model eukaryotic signaling system. We believe this combined approach constitutes a general means not only to reveal modification sites required to turn a pathway on and off, but also those required for more subtle quantitative effects that tune pathway output. Our results further suggest that relatively small quantitative influences from individual regulatory phosphorylation events endow signaling systems with plasticity that evolution may exploit to quantitatively tailor signaling outcomes.

  3. The crystal structure of Giardia duodenalis 14-3-3 in the apo form: when protein post-translational modifications make the difference.

    KAUST Repository

    Fiorillo, Annarita

    2014-03-21

    The 14-3-3s are a family of dimeric evolutionary conserved pSer/pThr binding proteins that play a key role in multiple biological processes by interacting with a plethora of client proteins. Giardia duodenalis is a flagellated protozoan that affects millions of people worldwide causing an acute and chronic diarrheal disease. The single giardial 14-3-3 isoform (g14-3-3), unique in the 14-3-3 family, needs the constitutive phosphorylation of Thr214 and the polyglycylation of its C-terminus to be fully functional in vivo. Alteration of the phosphorylation and polyglycylation status affects the parasite differentiation into the cyst stage. To further investigate the role of these post-translational modifications, the crystal structure of the g14-3-3 was solved in the unmodified apo form. Oligomers of g14-3-3 were observed due to domain swapping events at the protein C-terminus. The formation of filaments was supported by TEM. Mutational analysis, in combination with native PAGE and chemical cross-linking, proved that polyglycylation prevents oligomerization. In silico phosphorylation and molecular dynamics simulations supported a structural role for the phosphorylation of Thr214 in promoting target binding. Our findings highlight unique structural features of g14-3-3 opening novel perspectives on the evolutionary history of this protein family and envisaging the possibility to develop anti-giardial drugs targeting g14-3-3.

  4. The crystal structure of Giardia duodenalis 14-3-3 in the apo form: when protein post-translational modifications make the difference.

    Directory of Open Access Journals (Sweden)

    Annarita Fiorillo

    Full Text Available The 14-3-3s are a family of dimeric evolutionary conserved pSer/pThr binding proteins that play a key role in multiple biological processes by interacting with a plethora of client proteins. Giardia duodenalis is a flagellated protozoan that affects millions of people worldwide causing an acute and chronic diarrheal disease. The single giardial 14-3-3 isoform (g14-3-3, unique in the 14-3-3 family, needs the constitutive phosphorylation of Thr214 and the polyglycylation of its C-terminus to be fully functional in vivo. Alteration of the phosphorylation and polyglycylation status affects the parasite differentiation into the cyst stage. To further investigate the role of these post-translational modifications, the crystal structure of the g14-3-3 was solved in the unmodified apo form. Oligomers of g14-3-3 were observed due to domain swapping events at the protein C-terminus. The formation of filaments was supported by TEM. Mutational analysis, in combination with native PAGE and chemical cross-linking, proved that polyglycylation prevents oligomerization. In silico phosphorylation and molecular dynamics simulations supported a structural role for the phosphorylation of Thr214 in promoting target binding. Our findings highlight unique structural features of g14-3-3 opening novel perspectives on the evolutionary history of this protein family and envisaging the possibility to develop anti-giardial drugs targeting g14-3-3.

  5. The crystal structure of Giardia duodenalis 14-3-3 in the apo form: when protein post-translational modifications make the difference.

    KAUST Repository

    Fiorillo, Annarita; di Marino, Daniele; Bertuccini, Lucia; Via, Allegra; Pozio, Edoardo; Camerini, Serena; Ilari, Andrea; Lalle, Marco

    2014-01-01

    The 14-3-3s are a family of dimeric evolutionary conserved pSer/pThr binding proteins that play a key role in multiple biological processes by interacting with a plethora of client proteins. Giardia duodenalis is a flagellated protozoan that affects millions of people worldwide causing an acute and chronic diarrheal disease. The single giardial 14-3-3 isoform (g14-3-3), unique in the 14-3-3 family, needs the constitutive phosphorylation of Thr214 and the polyglycylation of its C-terminus to be fully functional in vivo. Alteration of the phosphorylation and polyglycylation status affects the parasite differentiation into the cyst stage. To further investigate the role of these post-translational modifications, the crystal structure of the g14-3-3 was solved in the unmodified apo form. Oligomers of g14-3-3 were observed due to domain swapping events at the protein C-terminus. The formation of filaments was supported by TEM. Mutational analysis, in combination with native PAGE and chemical cross-linking, proved that polyglycylation prevents oligomerization. In silico phosphorylation and molecular dynamics simulations supported a structural role for the phosphorylation of Thr214 in promoting target binding. Our findings highlight unique structural features of g14-3-3 opening novel perspectives on the evolutionary history of this protein family and envisaging the possibility to develop anti-giardial drugs targeting g14-3-3.

  6. An update on post-translational modifications of hydroxyproline-rich glycoproteins: Towards a model highlighting their contribution to plant cell wall architecture

    Directory of Open Access Journals (Sweden)

    May eHijazi

    2014-08-01

    Full Text Available Plant cell walls are composite structures mainly composed of polysaccharides, also containing a large set of proteins involved in diverse functions such as growth, environmental sensing, signaling, and defense. Research on cell wall proteins (CWPs is a challenging field since present knowledge of their role into the structure and function of cell walls is very incomplete. Among CWPs, hydroxyproline (Hyp-rich O-glycoproteins (HRGPs were classified into three categories: (i moderately glycosylated extensins (EXTs able to form covalent scaffolds; (ii hyperglycosylated arabinogalactan proteins (AGPs; and (iii Hyp/proline (Pro-Rich proteins (H/PRPs that may be non-, weakly- or highly-glycosylated. In this review, we provide a description of the main features of their post-translational modifications (PTMs, biosynthesis, structure and function. We propose a new model integrating HRGPs and their partners in cell walls. Altogether, they could form a continuous glyco-network with non-cellulosic polysaccharides via covalent bonds or non-covalent interactions, thus strongly contributing to cell wall architecture.

  7. Characterization of Chlamydomonas reinhardtii Core Histones by Top-Down Mass Spectrometry Reveals Unique Algae-Specific Variants and Post-Translational Modifications.

    Science.gov (United States)

    Khan, Aliyya; Eikani, Carlo K; Khan, Hana; Iavarone, Anthony T; Pesavento, James J

    2018-01-05

    The unicellular microalga Chlamydomonas reinhardtii has played an instrumental role in the development of many new fields (bioproducts, biofuels, etc.) as well as the advancement of basic science (photosynthetic apparati, flagellar function, etc.). Chlamydomonas' versatility ultimately derives from the genes encoded in its genome and the way that the expression of these genes is regulated, which is largely influenced by a family of DNA binding proteins called histones. We characterize C. reinhardtii core histones, both variants and their post-translational modifications, by chromatographic separation, followed by top-down mass spectrometry (TDMS). Because TDMS has not been previously used to study Chlamydomonas proteins, we show rampant artifactual protein oxidation using established nuclei purification and histone extraction methods. After addressing oxidation, both histones H3 and H4 are found to each have a single polypeptide sequence that is minimally acetylated and methylated. Surprisingly, we uncover a novel monomethylation at lysine 79 on histone H4 present on all observed molecules. Histone H2B and H2A are found to have two and three variants, respectively, and both are minimally modified. This study provides an updated assessment of the core histone proteins in the green alga C. reinhardtii by top-down mass spectrometry and lays the foundation for further investigation of these essential proteins.

  8. Direct Profiling the Post-Translational Modification Codes of a Single Protein Immobilized on a Surface Using Cu-free Click Chemistry.

    Science.gov (United States)

    Kim, Kyung Lock; Park, Kyeng Min; Murray, James; Kim, Kimoon; Ryu, Sung Ho

    2018-05-23

    Combinatorial post-translational modifications (PTMs), which can serve as dynamic "molecular barcodes", have been proposed to regulate distinct protein functions. However, studies of combinatorial PTMs on single protein molecules have been hindered by a lack of suitable analytical methods. Here, we describe erasable single-molecule blotting (eSiMBlot) for combinatorial PTM profiling. This assay is performed in a highly multiplexed manner and leverages the benefits of covalent protein immobilization, cyclic probing with different antibodies, and single molecule fluorescence imaging. Especially, facile and efficient covalent immobilization on a surface using Cu-free click chemistry permits multiple rounds (>10) of antibody erasing/reprobing without loss of antigenicity. Moreover, cumulative detection of coregistered multiple data sets for immobilized single-epitope molecules, such as HA peptide, can be used to increase the antibody detection rate. Finally, eSiMBlot enables direct visualization and quantitative profiling of combinatorial PTM codes at the single-molecule level, as we demonstrate by revealing the novel phospho-codes of ligand-induced epidermal growth factor receptor. Thus, eSiMBlot provides an unprecedentedly simple, rapid, and versatile platform for analyzing the vast number of combinatorial PTMs in biological pathways.

  9. ELISA-PLA: A novel hybrid platform for the rapid, highly sensitive and specific quantification of proteins and post-translational modifications.

    Science.gov (United States)

    Tong, Qing-He; Tao, Tao; Xie, Li-Qi; Lu, Hao-Jie

    2016-06-15

    Detection of low-abundance proteins and their post-translational modifications (PTMs) remains a great challenge. A conventional enzyme-linked immunosorbent assay (ELISA) is not sensitive enough to detect low-abundance PTMs and suffers from nonspecific detection. Herein, a rapid, highly sensitive and specific platform integrating ELISA with a proximity ligation assay (PLA), termed ELISA-PLA, was developed. Using ELISA-PLA, the specificity was improved by the simultaneous and proximate recognition of targets through multiple probes, and the sensitivity was significantly improved by rolling circle amplification (RCA). For GFP, the limit of detection (LOD) was decreased by two orders of magnitude compared to that of ELISA. Using site-specific phospho-antibody and pan-specific phospho-antibody, ELISA-PLA was successfully applied to quantify the phosphorylation dynamics of ERK1/2 and the overall tyrosine phosphorylation level of ERK1/2, respectively. ELISA-PLA was also used to quantify the O-GlcNAcylation of AKT, c-Fos, CREB and STAT3, which is faster and more sensitive than the conventional immunoprecipitation and western blotting (IP-WB) method. As a result, the sample consumption of ELISA-PLA was reduced 40-fold compared to IP-WB. Therefore, ELISA-PLA could be a promising platform for the rapid, sensitive and specific detection of proteins and PTMs. Copyright © 2016 Elsevier B.V. All rights reserved.

  10. An Engineered Version of Human PON2 Opens the Way to Understand the Role of Its Post-Translational Modifications in Modulating Catalytic Activity.

    Directory of Open Access Journals (Sweden)

    Luigi Mandrich

    Full Text Available The human paraoxonase 2 (PON2 has been described as a highly specific lactonase hydrolysing the quorum sensing molecule N-(3-oxododecanoyl-L-homoserine lactone (3OC12-HSL and having secondary esterase but not phosphotriesterase activity, in contrast with the related enzymes PON1 and PON3. It has been suggested that PON2 enzyme activity is dependent on glycosylation and its N-terminal region has been recently demonstrated to be a transmembrane domain mediating association to membranes. In the present study we describe a mutated form of PON2, lacking the above N-terminal region, which has been further stabilized by the insertion of six amino acidic substitutions. The engineered version, hence forth called rPON2, has been over-expressed in E.coli, refolded from inclusion bodies and purified, yielding an enzyme with the same characteristics as the full length enzyme. Therefore the first conclusion of this work was that the catalytic activity is independent from the N-terminus and protein glycosylation. The kinetic characterization confirmed the primary activity on 3OC12-HSL; accordingly, in vitro experiments of inhibition of the biofilm formed by Pseudomonas aeruginosa (PAO1 have demonstrated that rPON2 is more effective than PON1. In addition, we observed small but significant activity against organophosphorothiotes pesticides, m-parathion, coumaphos and malathion.The availability of fair amount of active protein allowed to pinpoint, by mass-spectrometry, ubiquitination of Lys 168 induced in rPON2 by HeLa extract and to correlate such post-translational modification to the modulation of catalytic activity. A mutational analysis of the modified residue confirmed the result.

  11. Osteopontin binding to the alpha 4 integrin requires highest affinity integrin conformation, but is independent of post-translational modifications of osteopontin

    DEFF Research Database (Denmark)

    Hui, Tommy; Sørensen, Esben Skipper; Rittling, Susan R.

    2015-01-01

    Osteopontin (OPN) is a ligand for the α4 integrin, but the physiological importance of this binding is not well understood. Here, we have assessed the effect of posttranslational modifications on OPN binding to the α4 integrin on cultured human leukocyte cell lines, and compared OPN interaction...

  12. Barley peroxidase isozymes. Expression and post-translational modification in mature seeds as identified by two-dimensional gel electrophoresis and mass spectrometry

    DEFF Research Database (Denmark)

    Laugesen, Sabrina; Bak-Jensen, Kristian Sass; Hägglund, Per

    2007-01-01

    spectrometric analysis. Distinct peroxidase spot patterns divided the 16 cultivars tested into two groups. The distribution of the three isozymes in different seed tissues (endosperm, embryo, and aleurone layer) suggested the peroxidases to play individual albeit partially overlapping roles during germination...

  13. Patterns of low temperature induced accumulation of dehydrins in Rosaceae crops-Evidence for post-translational modification in apple.

    Science.gov (United States)

    Haimi, Perttu; Vinskienė, Jurgita; Stepulaitienė, Inga; Baniulis, Danas; Stanienė, Gražina; Šikšnianienė, Jūratė Bronė; Rugienius, Rytis

    2017-11-01

    Important crop plants of Rosaceae family are often damaged during winter due to the lack of acclimation and cold hardiness. One of the cellular responses of plants to cold stress is the accumulation of dehydrin proteins. We studied the expression of dehydrins in several Rosaceae species during low temperature treatment in vitro. Microshoots of Pyrus communis, Malus×domestica, Fragaria vesca, Fragaria×ananassa, Prunus cerasus and Prunus avium cultivars were grown in low temperature conditions. Genotype -specific accumulation of dehydrins was detected by immunoblot analysis of the extracted proteins. Untargeted difference gel electrophoresis of Malus x domestica microshoots revealed an extensive accumulation of three dehydrins. In a protein phosphatase assay, MdDHN2 and MdDHN4, but not MdDHN6 proteins were found to be extensively phosphorylated. In terms of the amount of protein synthesized, dehydrins are a major protein-level adaptation mechanism to low temperature in M. x domestica. In addition to dehydrins, the induction of proteins involved in the response for oxidative stress were observed. Additionally, a Xero2 -like dehydrin of F. vesca was detected by difference gel electrophoresis and identified by nano LC-MS/MS. Copyright © 2017 Elsevier GmbH. All rights reserved.

  14. Novel Antimicrobial Peptides EeCentrocins 1, 2 and EeStrongylocin 2 from the Edible Sea Urchin Echinus esculentus Have 6-Br-Trp Post-Translational Modifications.

    Directory of Open Access Journals (Sweden)

    Runar Gjerp Solstad

    Full Text Available The global problem of microbial resistance to antibiotics has resulted in an urgent need to develop new antimicrobial agents. Natural antimicrobial peptides are considered promising candidates for drug development. Echinoderms, which rely on innate immunity factors in the defence against harmful microorganisms, are sources of novel antimicrobial peptides. This study aimed to isolate and characterise antimicrobial peptides from the Edible sea urchin Echinus esculentus. Using bioassay-guided purification and cDNA cloning, three antimicrobial peptides were characterised from the haemocytes of the sea urchin; two heterodimeric peptides and a cysteine-rich peptide. The peptides were named EeCentrocin 1 and 2 and EeStrongylocin 2, respectively, due to their apparent homology to the published centrocins and strongylocins isolated from the green sea urchin Strongylocentrotus droebachiensis. The two centrocin-like peptides EeCentrocin 1 and 2 are intramolecularly connected via a disulphide bond to form a heterodimeric structure, containing a cationic heavy chain of 30 and 32 amino acids and a light chain of 13 amino acids. Additionally, the light chain of EeCentrocin 2 seems to be N-terminally blocked by a pyroglutamic acid residue. The heavy chains of EeCentrocins 1 and 2 were synthesised and shown to be responsible for the antimicrobial activity of the natural peptides. EeStrongylocin 2 contains 6 cysteines engaged in 3 disulphide bonds. A fourth peptide (Ee4635 was also discovered but not fully characterised. Using mass spectrometric and NMR analyses, EeCentrocins 1 and 2, EeStrongylocin 2 and Ee4635 were all shown to contain post-translationally brominated Trp residues in the 6 position of the indole ring.

  15. Quantitative mass spectrometry of histones H3.2 and H3.3 in Suz12-deficient mouse embryonic stem cells reveals distinct, dynamic post-translational modifications at Lys-27 and Lys-36

    DEFF Research Database (Denmark)

    Jung, Hye Ryung; Pasini, Diego; Helin, Kristian

    2010-01-01

    distinct coexisting modifications. In certain cases, high mass accuracy LTQ-Orbitrap MS/MS allowed precise localization of near isobaric coexisting PTMs such as trimethylation and acetylation within individual peptides. ETD MS/MS facilitated sequencing and annotation of phosphorylated histone peptides....... The combined use of ETD and CID MS/MS increased the total number of identified modified peptides. Comparative quantitative analysis of histones from wild type and Suz12-deficient ESCs using stable isotope labeling with amino acids in cell culture and LC-MS/MS revealed a dramatic reduction of H3K27me2 and H3K27......me3 and an increase of H3K27ac, thereby uncovering an antagonistic methyl/acetyl switch at H3K27. The reduction in H3K27 methylation and increase in H3K27 acetylation was accompanied by H3K36 acetylation and methylation. Estimation of the global isoform percentage of unmodified and modified histone...

  16. Opioid gene expression changes and post-translational histone modifications at promoter regions in the rat nucleus accumbens after acute and repeated 3,4-methylenedioxy-methamphetamine (MDMA) exposure.

    Science.gov (United States)

    Caputi, Francesca Felicia; Palmisano, Martina; Carboni, Lucia; Candeletti, Sanzio; Romualdi, Patrizia

    2016-12-01

    The recreational drug of abuse 3,4-methylenedioxymethamphetamine (MDMA) has been shown to produce neurotoxic damage and long-lasting changes in several brain areas. In addition to the involvement of serotoninergic and dopaminergic systems, little information exists about the contribution of nociceptin/orphaninFQ (N/OFQ)-NOP and dynorphin (DYN)-KOP systems in neuronal adaptations evoked by MDMA. Here we investigated the behavioral and molecular effects induced by acute (8mg/kg) or repeated (8mg/kg twice daily for seven days) MDMA exposure. MDMA exposure affected body weight gain and induced hyperlocomotion; this latter effect progressively decreased after repeated administration. Gene expression analysis indicated a down-regulation of the N/OFQ system and an up-regulation of the DYN system in the nucleus accumbens (NAc), highlighting an opposite systems regulation in response to MDMA exposure. Since histone modifications have been strongly associated to the addiction-related maladaptive changes, we examined two permissive (acH3K9 and me3H3K4) and two repressive transcription marks (me3H3K27 and me2H3K9) at the pertinent opioid gene promoter regions. Chromatin immunoprecipitation assays revealed that acute MDMA increased me3H3K4 at the pN/OFQ, pDYN and NOP promoters. Following acute and repeated treatment a significant decrease of acH3K9 at the pN/OFQ promoter was observed, which correlated with gene expression results. Acute treatment caused an acH3K9 increase and a me2H3K9 decrease at the pDYN promoter which matched its mRNA up-regulation. Our data indicate that the activation of the DYNergic stress system together with the inactivation of the N/OFQergic anti-stress system contribute to the neuroadaptive actions of MDMA and offer novel epigenetic information associated with MDMA abuse. Copyright © 2016 Elsevier Ltd. All rights reserved.

  17. Post-Translational Modifications of Histones in Human Sperm

    Czech Academy of Sciences Publication Activity Database

    Krejčí, Jana; Stixová, Lenka; Legartová, Soňa; Kozubek, Stanislav; Lochmanová, G.; Zdráhal, Z.; Sehnalová, Petra; Dabravolski, S.; Hejatko, J.; Bártová, Eva

    2015-01-01

    Roč. 116, č. 10 (2015), s. 2195-2209 ISSN 0730-2312 R&D Projects: GA MŠk(CZ) ED1.1.00/02.0068 Institutional support: RVO:68081707 Keywords : HUMAN SPERM * HISTONES * PROTAMINE P2 Subject RIV: BO - Biophysics Impact factor: 3.446, year: 2015

  18. Post-translational Control of Intracellular Pathogen Sensing Pathways.

    Science.gov (United States)

    Chiang, Cindy; Gack, Michaela U

    2017-01-01

    Mammalian cells recognize virus-derived nucleic acids using a defined set of intracellular sensors including the DNA sensors cyclic GMP-AMP (cGAMP) synthase (cGAS) and interferon gamma (IFNγ)-inducible protein 16 (IFI16) as well as viral RNA receptors of the retinoic acid-inducible gene I (RIG-I)-like receptor (RLR) family. Following innate immune recognition, these sensors launch an immune response that is characterized by the transcriptional upregulation of many antiviral molecules, including proinflammatory cytokines, chemokines, and IFN-stimulated genes. Recent studies have demonstrated that the signal transduction initiated by these sensors is sophisticatedly regulated by post-translational modifications (PTMs) resulting in a robust yet 'tunable' cytokine response to maintain immune homeostasis. Here we summarize recent advances in our understanding of how PTMs and regulatory enzymes control the signaling activity of RLRs, cGAS, and IFI16 as well as their proximal adaptor proteins. Copyright © 2016 Elsevier Ltd. All rights reserved.

  19. Variations of Histone Modification Patterns: Contributions of Inter-plant Variability and Technical Factors

    Directory of Open Access Journals (Sweden)

    Sylva Brabencová

    2017-12-01

    Full Text Available Inter-individual variability of conspecific plants is governed by differences in their genetically determined growth and development traits, environmental conditions, and adaptive responses under epigenetic control involving histone post-translational modifications. The apparent variability in histone modifications among plants might be increased by technical variation introduced in sample processing during epigenetic analyses. Thus, to detect true variations in epigenetic histone patterns associated with given factors, the basal variability among samples that is not associated with them must be estimated. To improve knowledge of relative contribution of biological and technical variation, mass spectrometry was used to examine histone modification patterns (acetylation and methylation among Arabidopsis thaliana plants of ecotypes Columbia 0 (Col-0 and Wassilewskija (Ws homogenized by two techniques (grinding in a cryomill or with a mortar and pestle. We found little difference in histone modification profiles between the ecotypes. However, in comparison of the biological and technical components of variability, we found consistently higher inter-individual variability in histone mark levels among Ws plants than among Col-0 plants (grown from seeds collected either from single plants or sets of plants. Thus, more replicates of Ws would be needed for rigorous analysis of epigenetic marks. Regarding technical variability, the cryomill introduced detectably more heterogeneity in the data than the mortar and pestle treatment, but mass spectrometric analyses had minor apparent effects. Our study shows that it is essential to consider inter-sample variance and estimate suitable numbers of biological replicates for statistical analysis for each studied organism when investigating changes in epigenetic histone profiles.

  20. Variations of Histone Modification Patterns: Contributions of Inter-plant Variability and Technical Factors.

    Science.gov (United States)

    Brabencová, Sylva; Ihnatová, Ivana; Potěšil, David; Fojtová, Miloslava; Fajkus, Jiří; Zdráhal, Zbyněk; Lochmanová, Gabriela

    2017-01-01

    Inter-individual variability of conspecific plants is governed by differences in their genetically determined growth and development traits, environmental conditions, and adaptive responses under epigenetic control involving histone post-translational modifications. The apparent variability in histone modifications among plants might be increased by technical variation introduced in sample processing during epigenetic analyses. Thus, to detect true variations in epigenetic histone patterns associated with given factors, the basal variability among samples that is not associated with them must be estimated. To improve knowledge of relative contribution of biological and technical variation, mass spectrometry was used to examine histone modification patterns (acetylation and methylation) among Arabidopsis thaliana plants of ecotypes Columbia 0 (Col-0) and Wassilewskija (Ws) homogenized by two techniques (grinding in a cryomill or with a mortar and pestle). We found little difference in histone modification profiles between the ecotypes. However, in comparison of the biological and technical components of variability, we found consistently higher inter-individual variability in histone mark levels among Ws plants than among Col-0 plants (grown from seeds collected either from single plants or sets of plants). Thus, more replicates of Ws would be needed for rigorous analysis of epigenetic marks. Regarding technical variability, the cryomill introduced detectably more heterogeneity in the data than the mortar and pestle treatment, but mass spectrometric analyses had minor apparent effects. Our study shows that it is essential to consider inter-sample variance and estimate suitable numbers of biological replicates for statistical analysis for each studied organism when investigating changes in epigenetic histone profiles.

  1. Statistical density modification using local pattern matching

    International Nuclear Information System (INIS)

    Terwilliger, Thomas C.

    2003-01-01

    Statistical density modification can make use of local patterns of density found in protein structures to improve crystallographic phases. A method for improving crystallographic phases is presented that is based on the preferential occurrence of certain local patterns of electron density in macromolecular electron-density maps. The method focuses on the relationship between the value of electron density at a point in the map and the pattern of density surrounding this point. Patterns of density that can be superimposed by rotation about the central point are considered equivalent. Standard templates are created from experimental or model electron-density maps by clustering and averaging local patterns of electron density. The clustering is based on correlation coefficients after rotation to maximize the correlation. Experimental or model maps are also used to create histograms relating the value of electron density at the central point to the correlation coefficient of the density surrounding this point with each member of the set of standard patterns. These histograms are then used to estimate the electron density at each point in a new experimental electron-density map using the pattern of electron density at points surrounding that point and the correlation coefficient of this density to each of the set of standard templates, again after rotation to maximize the correlation. The method is strengthened by excluding any information from the point in question from both the templates and the local pattern of density in the calculation. A function based on the origin of the Patterson function is used to remove information about the electron density at the point in question from nearby electron density. This allows an estimation of the electron density at each point in a map, using only information from other points in the process. The resulting estimates of electron density are shown to have errors that are nearly independent of the errors in the original map using

  2. Post-translational regulation enables robust p53 regulation.

    Science.gov (United States)

    Shin, Yong-Jun; Chen, Kai-Yuan; Sayed, Ali H; Hencey, Brandon; Shen, Xiling

    2013-08-30

    The tumor suppressor protein p53 plays important roles in DNA damage repair, cell cycle arrest and apoptosis. Due to its critical functions, the level of p53 is tightly regulated by a negative feedback mechanism to increase its tolerance towards fluctuations and disturbances. Interestingly, the p53 level is controlled by post-translational regulation rather than transcriptional regulation in this feedback mechanism. We analyzed the dynamics of this feedback to understand whether post-translational regulation provides any advantages over transcriptional regulation in regard to disturbance rejection. When a disturbance happens, even though negative feedback reduces the steady-state error, it can cause a system to become less stable and transiently overshoots, which may erroneously trigger downstream reactions. Therefore, the system needs to balance the trade-off between steady-state and transient errors. Feedback control and adaptive estimation theories revealed that post-translational regulation achieves a better trade-off than transcriptional regulation, contributing to a more steady level of p53 under the influence of noise and disturbances. Furthermore, post-translational regulation enables cells to respond more promptly to stress conditions with consistent amplitude. However, for better disturbance rejection, the p53- Mdm2 negative feedback has to pay a price of higher stochastic noise. Our analyses suggest that the p53-Mdm2 feedback favors regulatory mechanisms that provide the optimal trade-offs for dynamic control.

  3. Radical SAM Enzymes in the Biosynthesis of Ribosomally Synthesized and Post-translationally Modified Peptides (RiPPs

    Directory of Open Access Journals (Sweden)

    Alhosna Benjdia

    2017-11-01

    Full Text Available Ribosomally-synthesized and post-translationally modified peptides (RiPPs are a large and diverse family of natural products. They possess interesting biological properties such as antibiotic or anticancer activities, making them attractive for therapeutic applications. In contrast to polyketides and non-ribosomal peptides, RiPPs derive from ribosomal peptides and are post-translationally modified by diverse enzyme families. Among them, the emerging superfamily of radical SAM enzymes has been shown to play a major role. These enzymes catalyze the formation of a wide range of post-translational modifications some of them having no counterparts in living systems or synthetic chemistry. The investigation of radical SAM enzymes has not only illuminated unprecedented strategies used by living systems to tailor peptides into complex natural products but has also allowed to uncover novel RiPP families. In this review, we summarize the current knowledge on radical SAM enzymes catalyzing RiPP post-translational modifications and discuss their mechanisms and growing importance notably in the context of the human microbiota.

  4. Experimental annotation of post-translational features and translated coding regions in the pathogen Salmonella Typhimurium

    Energy Technology Data Exchange (ETDEWEB)

    Ansong, Charles; Tolic, Nikola; Purvine, Samuel O.; Porwollik, Steffen; Jones, Marcus B.; Yoon, Hyunjin; Payne, Samuel H.; Martin, Jessica L.; Burnet, Meagan C.; Monroe, Matthew E.; Venepally, Pratap; Smith, Richard D.; Peterson, Scott; Heffron, Fred; Mcclelland, Michael; Adkins, Joshua N.

    2011-08-25

    Complete and accurate genome annotation is crucial for comprehensive and systematic studies of biological systems. For example systems biology-oriented genome scale modeling efforts greatly benefit from accurate annotation of protein-coding genes to develop proper functioning models. However, determining protein-coding genes for most new genomes is almost completely performed by inference, using computational predictions with significant documented error rates (> 15%). Furthermore, gene prediction programs provide no information on biologically important post-translational processing events critical for protein function. With the ability to directly measure peptides arising from expressed proteins, mass spectrometry-based proteomics approaches can be used to augment and verify coding regions of a genomic sequence and importantly detect post-translational processing events. In this study we utilized “shotgun” proteomics to guide accurate primary genome annotation of the bacterial pathogen Salmonella Typhimurium 14028 to facilitate a systems-level understanding of Salmonella biology. The data provides protein-level experimental confirmation for 44% of predicted protein-coding genes, suggests revisions to 48 genes assigned incorrect translational start sites, and uncovers 13 non-annotated genes missed by gene prediction programs. We also present a comprehensive analysis of post-translational processing events in Salmonella, revealing a wide range of complex chemical modifications (70 distinct modifications) and confirming more than 130 signal peptide and N-terminal methionine cleavage events in Salmonella. This study highlights several ways in which proteomics data applied during the primary stages of annotation can improve the quality of genome annotations, especially with regards to the annotation of mature protein products.

  5. Post-translationally modified muscle-specific ubiquitin ligases as circulating biomarkers in experimental cancer cachexia

    Science.gov (United States)

    Mota, Roberto; Rodríguez, Jessica E; Bonetto, Andrea; O’Connell, Thomas M; Asher, Scott A; Parry, Traci L; Lockyer, Pamela; McCudden, Christopher R; Couch, Marion E; Willis, Monte S

    2017-01-01

    Cancer cachexia is a severe wasting syndrome characterized by the progressive loss of lean body mass and systemic inflammation. Up to 80% of cancer patients experience cachexia, with 20-30% of cancer-related deaths directly linked to cachexia. Despite efforts to identify early cachexia and cancer relapse, clinically useful markers are lacking. Recently, we identified the role of muscle-specific ubiquitin ligases Atrogin-1 (MAFbx, FBXO32) and Muscle Ring Finger-1 in the pathogenesis of cardiac atrophy and hypertrophy. We hypothesized that during cachexia, the Atrogin-1 and MuRF1 ubiquitin ligases are released from muscle and migrate to the circulation where they could be detected and serve as a cachexia biomarker. To test this, we induced cachexia in mice using the C26 adenocarcinoma cells or vehicle (control). Body weight, tumor volume, and food consumption were measured from inoculation until ~day 14 to document cachexia. Western blot analysis of serum identified the presence of Atrogin-1 and MuRF1 with unique post-translational modifications consistent with mono- and poly- ubiquitination of Atrogin-1 and MuRF1 found only in cachectic serum. These findings suggest that both increased Atrogin-1 and the presence of unique post-translational modifications may serve as a surrogate marker specific for cachexia. PMID:28979816

  6. Human Immunodeficiency Virus type-1 reverse transcriptase exists as post-translationally modified forms in virions and cells

    Directory of Open Access Journals (Sweden)

    Warrilow David

    2008-12-01

    Full Text Available Abstract Background HIV-1 reverse transcriptase (RT is a heterodimer composed of p66 and p51 subunits and is responsible for reverse transcription of the viral RNA genome into DNA. RT can be post-translationally modified in vitro which may be an important mechanism for regulating RT activity. Here we report detection of different p66 and p51 RT isoforms by 2D gel electrophoresis in virions and infected cells. Results Major isoforms of the p66 and p51 RT subunits were observed, with pI's of 8.44 and 8.31 respectively (p668.44 and p518.31. The same major isoforms were present in virions, virus-infected cell lysates and intracellular reverse transcription complexes (RTCs, and their presence in RTCs suggested that these are likely to be the forms that function in reverse transcription. Several minor RT isoforms were also observed. The observed pIs of the RT isoforms differed from the pI of theoretical unmodified RT (p668.53 and p518.60, suggesting that most of the RT protein in virions and cells is post-translationally modified. The modifications of p668.44 and p518.31 differed from each other indicating selective modification of the different RT subunits. The susceptibility of RT isoforms to phosphatase treatment suggested that some of these modifications were due to phosphorylation. Dephosphorylation, however, had no effect on in vitro RT activity associated with virions, infected cells or RTCs suggesting that the phospho-isoforms do not make a major contribution to RT activity in an in vitro assay. Conclusion The same major isoform of p66 and p51 RT is found in virions, infected cells and RTC's and both of these subunits are post-translationally modified. This post-translational modification of RT may be important for the function of RT inside the cell.

  7. Prediction of post-translational glycosylation and phosphorylation of proteins from the amino acid sequence

    DEFF Research Database (Denmark)

    Blom, Nikolaj; Sicheritz-Pontén, Thomas; Gupta, Ramneek

    2004-01-01

    Post-translational modifications (PTMs) occur on almost all proteins analyzed to date. The function of a modified protein is often strongly affected by these modifications and therefore increased knowledge about the potential PTMs of a target protein may increase our understanding of the molecular...... steps by integrating computational approaches into the validation procedures. Many advanced methods for the prediction of PTMs exist and many are made publicly available. We describe our experiences with the development of prediction methods for phosphorylation and glycosylation sites...... and the development of PTM-specific databases. In addition, we discuss novel ideas for PTM visualization (exemplified by kinase landscapes) and improvements for prediction specificity (by using ESS-evolutionary stable sites). As an example, we present a new method for kinase-specific prediction of phosphorylation...

  8. A post-translational balancing act: the good and the bad of SUMOylation in pancreatic islets.

    Science.gov (United States)

    MacDonald, Patrick E

    2018-04-01

    Post-translational modification of proteins contributes to the control of cell function and survival. The balance of these in insulin-producing pancreatic beta cells is important for the maintenance of glucose homeostasis. Protection from the damaging effects of reactive oxygen species is required for beta cell survival, but if this happens at the expense of insulin secretory function then the ability of islets to respond to changing metabolic conditions may be compromised. In this issue of Diabetologia, He et al ( https://doi.org/10.1007/s00125-017-4523-9 ) show that post-translational attachment of small ubiquitin-like modifier (SUMO) to target lysine residues (SUMOylation) strikes an important balance between the protection of beta cells from oxidative stress and the maintenance of insulin secretory function. They show that SUMOylation is required to stabilise nuclear factor erythroid 2-related factor 2 (NRF2) and increase antioxidant gene expression. Decreasing SUMOylation in beta cells impairs their antioxidant capacity, causes cell death, hyperglycaemia, and increased sensitivity to streptozotocin-induced diabetes, while increasing SUMOylation is protective. However, this protection from overt diabetes occurs in concert with glucose intolerance due to impaired beta cell function. A possible role for SUMOylation as a key factor balancing beta cell protection vs beta cell responsiveness to metabolic cues is discussed in this Commentary.

  9. Incorporation of post-translational modified amino acids as an approach to increase both chemical and biological diversity of conotoxins and conopeptides.

    Science.gov (United States)

    Espiritu, Michael J; Cabalteja, Chino C; Sugai, Christopher K; Bingham, Jon-Paul

    2014-01-01

    Bioactive peptides from Conus venom contain a natural abundance of post-translational modifications that affect their chemical diversity, structural stability, and neuroactive properties. These modifications have continually presented hurdles in their identification and characterization. Early endeavors in their analysis relied on classical biochemical techniques that have led to the progressive development and use of novel proteomic-based approaches. The critical importance of these post-translationally modified amino acids and their specific assignment cannot be understated, having impact on their folding, pharmacological selectivity, and potency. Such modifications at an amino acid level may also provide additional insight into the advancement of conopeptide drugs in the quest for precise pharmacological targeting. To achieve this end, a concerted effort between the classical and novel approaches is needed to completely elucidate the role of post-translational modifications in conopeptide structure and dynamics. This paper provides a reflection in the advancements observed in dealing with numerous and multiple post-translationally modified amino acids within conotoxins and conopeptides and provides a summary of the current techniques used in their identification.

  10. Post-translational glutamylation and tyrosination in tubulin of tritrichomonads and the diplomonad Giardia intestinalis.

    Science.gov (United States)

    Boggild, A K; Sundermann, C A; Estridge, B H

    2002-01-01

    Glutamylated and tyrosinated tubulin were localized in Giardia intestinalis and selected trichomonads of the Tritrichomonadinae subfamily, using specific monoclonal antibodies directed at each of the post-translational modifications. Analysis was carried out using indirect immunofluorescence microscopy. Although trichomonad tubulins remained unlabeled by anti-tyrosine tubulin (TUB-1A2), the presence of the glutamylation motif (GT 335) was confirmed and found to differ in distribution among tritrichomonads. Tritrichomonas muris was most heavily labeled with GT 335, while T. foetus was the least so. Like trichomonads, Giardia was unreactive to anti-tyrosine tubulin; however, the GT 335 antibody produced marked fluorescence in Giardia trophozoites. This study is the first to report immunofluorescent localization of tubulin glutamylation in Giardia and confirms previously reported mass spectrometry data.

  11. Printing-assisted surface modifications of patterned ultrafiltration membranes

    International Nuclear Information System (INIS)

    Wardrip, Nathaniel C.; Dsouza, Melissa; Urgun-Demirtas, Meltem; Snyder, Seth W.

    2016-01-01

    Understanding and restricting microbial surface attachment will enhance wastewater treatment with membranes. We report a maskless lithographic patterning technique for the generation of patterned polymer coatings on ultrafiltration membranes. Polyethylene glycol, zwitterionic, or negatively charged hydrophilic polymer compositions in parallel- or perpendicular-striped patterns with respect to feed flow were evaluated using wastewater. Membrane fouling was dependent on the orientation and chemical composition of the coatings. Modifications reduced alpha diversity in the attached microbial community (Shannon indices decreased from 2.63 to 1.89) which nevertheless increased with filtration time. Sphingomonas species, which condition membrane surfaces and facilitate cellular adhesion, were depleted in all modified membranes. Microbial community structure was significantly different between control, different patterns, and different chemistries. Lastly, this study broadens the tools for surface modification of membranes with polymer coatings and for understanding and optimization of antifouling surfaces.

  12. Post-translational control of nitrate reductase activity responding to light and photosynthesis evolved already in the early vascular plants.

    Science.gov (United States)

    Nemie-Feyissa, Dugassa; Królicka, Adriana; Førland, Nina; Hansen, Margarita; Heidari, Behzad; Lillo, Cathrine

    2013-05-01

    Regulation of nitrate reductase (NR) by reversible phosphorylation at a conserved motif is well established in higher plants, and enables regulation of NR in response to rapid fluctuations in light intensity. This regulation is not conserved in algae NR, and we wished to test the evolutionary origin of the regulatory mechanism by physiological examination of ancient land plants. Especially a member of the lycophytes is of interest since their NR is candidate for regulation by reversible phosphorylation based on sequence analysis. We compared Selaginella kraussiana, a member of the lycophytes and earliest vascular plants, with the angiosperm Arabidopsis thaliana, and also tested the moss Physcomitrella patens. Interestingly, optimization of assay conditions revealed that S. kraussiana NR used NADH as an electron donor like A. thaliana, whereas P. patens NR activity depended on NADPH. Examination of light/darkness effects showed that S. kraussiana NR was rapidly regulated similar to A. thaliana NR when a differential (Mg(2+) contra EDTA) assay was used to reveal activity state of NR. This implies that already existing NR enzyme was post-translationally activated by light in both species. Light had a positive effect also on de novo synthesis of NR in S. kraussiana, which could be shown after the plants had been exposed to a prolonged dark period (7 days). Daily variations in NR activity were mainly caused by post-translational modifications. As for angiosperms, the post-translational light activation of NR in S. kraussiana was inhibited by 3-(3,4-dichlorophenyl)-1*1-dimethylurea (DCMU), an inhibitor of photosynthesis and stomata opening. Evolutionary, a post-translational control mechanism for NR have occurred before or in parallel with development of vascular tissue in land plants, and appears to be part of a complex mechanisms for coordination of CO2 and nitrogen metabolism in these plants. Copyright © 2013 Elsevier GmbH. All rights reserved.

  13. Post-translational processing and secretion of atrial natriuretic factor

    International Nuclear Information System (INIS)

    Shields, P.P.

    1988-01-01

    The post-translational processing and regulated secretion of atrial natriuretic factor (ANF) were studied in primary cultures of rat cardiac myocytes. Cultures were established from neonatal rat atria or ventricles, and were maintained for 7-14 days in complete serum free medium. The cultures contained high and constant levels of ANF-(1-126), the known storage form of the hormone in vivo. The cultures also secreted ANF-(1-126), instead of the known circulating form of the hormone, ANF-(99-126). However, the inclusion of the glucocorticoids dexamethasone or hydrocortisone in the culture medium increased the levels of ir-ANF contained and secreted by the cultures, and caused both atrial and ventricular cultures to secrete principally ANF-(99-126) instead of ANF-(1-126). The secreted peptide was shown to be authentic ANF-(99-126) by chromatographic, amino acid composition and radiosequence analysis, thus confirming that the cultures were accurately processing ANF to the in vivo circulating form in the presence of glucocorticoids. Glucocorticoids also caused an increase in size and clustering of atrial myocytes as determined by immunocytochemical analysis, but the morphological effects could be dissociated from the stimulation of ANF-(99-126) secretion by manipulating the timing of glucocorticoid exposure. The location of ANF-(99-126) formation was investigated using biosynthetically labeled 35 S-Cys-ANF-(1-126) in conjunction with actively processing cultures

  14. Pdx1 is post-translationally modified in vivo and serine 61 is the principal site of phosphorylation

    DEFF Research Database (Denmark)

    Frogne, Thomas; Sylvestersen, Kathrine Beck; Kubicek, Stefan

    2012-01-01

    signaling. Several studies have shown that post-translational modifications are regulating Pdx1 activity through intracellular localization and binding to co-factors. Understanding the signaling cues converging on Pdx1 and modulating its activity is therefore an attractive approach in diabetes treatment. We...... alanine scanning and mass spectrometry we map this phosphorylation to serine 61 in both Min6 cells and in exogenous Pdx1 over-expressed in HEK293 cells. A single phosphorylation is also present in cultured islets but it remains unaffected by changes in glucose levels. It is present during embryogenesis...

  15. An improved ChIP-seq peak detection system for simultaneously identifying post-translational modified transcription factors by combinatorial fusion, using SUMOylation as an example.

    Science.gov (United States)

    Cheng, Chia-Yang; Chu, Chia-Han; Hsu, Hung-Wei; Hsu, Fang-Rong; Tang, Chung Yi; Wang, Wen-Ching; Kung, Hsing-Jien; Chang, Pei-Ching

    2014-01-01

    Post-translational modification (PTM) of transcriptional factors and chromatin remodelling proteins is recognized as a major mechanism by which transcriptional regulation occurs. Chromatin immunoprecipitation (ChIP) in combination with high-throughput sequencing (ChIP-seq) is being applied as a gold standard when studying the genome-wide binding sites of transcription factor (TFs). This has greatly improved our understanding of protein-DNA interactions on a genomic-wide scale. However, current ChIP-seq peak calling tools are not sufficiently sensitive and are unable to simultaneously identify post-translational modified TFs based on ChIP-seq analysis; this is largely due to the wide-spread presence of multiple modified TFs. Using SUMO-1 modification as an example; we describe here an improved approach that allows the simultaneous identification of the particular genomic binding regions of all TFs with SUMO-1 modification. Traditional peak calling methods are inadequate when identifying multiple TF binding sites that involve long genomic regions and therefore we designed a ChIP-seq processing pipeline for the detection of peaks via a combinatorial fusion method. Then, we annotate the peaks with known transcription factor binding sites (TFBS) using the Transfac Matrix Database (v7.0), which predicts potential SUMOylated TFs. Next, the peak calling result was further analyzed based on the promoter proximity, TFBS annotation, a literature review, and was validated by ChIP-real-time quantitative PCR (qPCR) and ChIP-reChIP real-time qPCR. The results show clearly that SUMOylated TFs are able to be pinpointed using our pipeline. A methodology is presented that analyzes SUMO-1 ChIP-seq patterns and predicts related TFs. Our analysis uses three peak calling tools. The fusion of these different tools increases the precision of the peak calling results. TFBS annotation method is able to predict potential SUMOylated TFs. Here, we offer a new approach that enhances Ch

  16. A single cysteine post-translational oxidation suffices to compromise globular proteins kinetic stability and promote amyloid formation

    Directory of Open Access Journals (Sweden)

    Patrizia Marinelli

    2018-04-01

    Full Text Available Oxidatively modified forms of proteins accumulate during aging. Oxidized protein conformers might act as intermediates in the formation of amyloids in age-related disorders. However, it is not known whether this amyloidogenic conversion requires an extensive protein oxidative damage or it can be promoted just by a discrete, localized post-translational modification of certain residues. Here, we demonstrate that the irreversible oxidation of a single free Cys suffices to severely perturb the folding energy landscape of a stable globular protein, compromise its kinetic stability, and lead to the formation of amyloids under physiological conditions. Experiments and simulations converge to indicate that this specific oxidation-promoted protein aggregation requires only local unfolding. Indeed, a large scale analysis indicates that many cellular proteins are at risk of undergoing this kind of deleterious transition; explaining how oxidative stress can impact cell proteostasis and subsequently lead to the onset of pathological states. Keywords: Protein oxidation, Protein misfolding, Protein aggregation, Oxidative stress, Post-translational modification

  17. Regulation of dynamin family proteins by post-translational

    Indian Academy of Sciences (India)

    2017-04-22

    Apr 22, 2017 ... School of Biological Sciences, National Institute of Science Education and Research-. Bhubaneswar .... translational modifications provides the cell a dynamic and ..... Drp1 which confers cellular protection from mitochondrial.

  18. Post-translational processing targets functionally diverse proteins in Mycoplasma hyopneumoniae.

    Science.gov (United States)

    Tacchi, Jessica L; Raymond, Benjamin B A; Haynes, Paul A; Berry, Iain J; Widjaja, Michael; Bogema, Daniel R; Woolley, Lauren K; Jenkins, Cheryl; Minion, F Chris; Padula, Matthew P; Djordjevic, Steven P

    2016-02-01

    Mycoplasma hyopneumoniae is a genome-reduced, cell wall-less, bacterial pathogen with a predicted coding capacity of less than 700 proteins and is one of the smallest self-replicating pathogens. The cell surface of M. hyopneumoniae is extensively modified by processing events that target the P97 and P102 adhesin families. Here, we present analyses of the proteome of M. hyopneumoniae-type strain J using protein-centric approaches (one- and two-dimensional GeLC-MS/MS) that enabled us to focus on global processing events in this species. While these approaches only identified 52% of the predicted proteome (347 proteins), our analyses identified 35 surface-associated proteins with widely divergent functions that were targets of unusual endoproteolytic processing events, including cell adhesins, lipoproteins and proteins with canonical functions in the cytosol that moonlight on the cell surface. Affinity chromatography assays that separately used heparin, fibronectin, actin and host epithelial cell surface proteins as bait recovered cleavage products derived from these processed proteins, suggesting these fragments interact directly with the bait proteins and display previously unrecognized adhesive functions. We hypothesize that protein processing is underestimated as a post-translational modification in genome-reduced bacteria and prokaryotes more broadly, and represents an important mechanism for creating cell surface protein diversity. © 2016 The Authors.

  19. The Oncoprotein BRD4-NUT Generates Aberrant Histone Modification Patterns.

    Directory of Open Access Journals (Sweden)

    Barry M Zee

    Full Text Available Defects in chromatin proteins frequently manifest in diseases. A striking case of a chromatin-centric disease is NUT-midline carcinoma (NMC, which is characterized by expression of NUT as a fusion partner most frequently with BRD4. ChIP-sequencing studies from NMC patients revealed that BRD4-NUT (B4N covers large genomic regions and elevates transcription within these domains. To investigate how B4N modulates chromatin, we performed affinity purification of B4N when ectopically expressed in 293-TREx cells and quantified the associated histone posttranslational modifications (PTM using proteomics. We observed significant enrichment of acetylation particularly on H3 K18 and of combinatorial patterns such as H3 K27 acetylation paired with K36 methylation. We postulate that B4N complexes override the preexisting histone code with new PTM patterns that reflect aberrant transcription and that epigenetically modulate the nucleosome environment toward the NMC state.

  20. Contribution of Post-translational Phosphorylation to Sarcomere-linked Cardiomyopathy Phenotypes

    Directory of Open Access Journals (Sweden)

    Margaret V Westfall

    2016-09-01

    Full Text Available Secondary shifts develop in post-translational phosphorylation of sarcomeric proteins in multi¬ple animal models of inherited cardiomyopathy. These signaling alterations together with the primary mutation are predicted to contribute to the overall cardiac phenotype. As a result, identification and integration of post-translational myofilament signaling responses are identified as priorities for gaining insights into sarcomeric cardiomyopathies. However, significant questions remain about the nature and contribution of post-translational phosphorylation to structural remodeling and cardiac dysfunction in animal models and human patients. This perspective essay discusses specific goals for filling critical gaps about post-translational signaling in response to these inherited mutations, especially within sarcomeric proteins. The discussion focuses primarily on pre-clinical analysis of animal models and defines challenges and future directions in this field.

  1. Fluorescent polymer-based post-translational differentiation and subtyping of breast cancer cells.

    Science.gov (United States)

    Scott, Michael D; Dutta, Rinku; Haldar, Manas K; Wagh, Anil; Gustad, Thomas R; Law, Benedict; Friesner, Daniel L; Mallik, Sanku

    2012-12-07

    Herein, we report the application of synthesized fluorescent, water soluble polymers for post-translational subtyping and differentiation of breast cancer cells in vitro. The fluorescence emission spectra from these polymers were modulated differently in the presence of conditioned cell culture media from various breast cancer cells. These polymers differentiate at a post-translation level possibly due to their ability to interact with extracellular enzymes that are over-expressed in cancerous conditions.

  2. Extracellular and Intracellular Cyclophilin A, Native and Post-Translationally Modified, Show Diverse and Specific Pathological Roles in Diseases.

    Science.gov (United States)

    Xue, Chao; Sowden, Mark P; Berk, Bradford C

    2018-05-01

    CypA (cyclophilin A) is a ubiquitous and highly conserved protein with peptidyl prolyl isomerase activity. Because of its highly abundant level in the cytoplasm, most studies have focused on the roles of CypA as an intracellular protein. However, emerging evidence suggests an important role for extracellular CypA in the pathogenesis of several diseases through receptor (CD147 or other)-mediated autocrine and paracrine signaling pathways. In this review, we will discuss the shared and unique pathological roles of extracellular and intracellular CypA in human cardiovascular diseases. In addition, the evolving role of post-translational modifications of CypA in the pathogenesis of disease is discussed. Finally, recent studies with drugs specific for extracellular CypA show its importance in disease pathogenesis in several animal models and make extracellular CypA a new therapeutic target. © 2018 American Heart Association, Inc.

  3. Mass spectrometric identification of proteins and characterization of their post-translational modifications in proteome analysis

    DEFF Research Database (Denmark)

    Roepstorff, P; Larsen, Martin Røssel

    2001-01-01

    High-throughput DNA sequencing has resulted in increasing input in protein sequence databases. Today more than 20 genomes have been sequenced and many more will be completed in the near future, including the largest of them all, the human genome. Presently, sequence databases contain entries for ...

  4. Post-translational modification of LipL32 during Leptospira interrogans infection

    Science.gov (United States)

    Leptospirosis, a re-emerging disease of global importance caused by pathogenic Leptospira spp., is considered the world’s most widespread zoonotic disease. Rats serve as asymptomatic carriers of pathogenic Leptospira and are critical for disease spread. In such reservoir hosts, leptospires colonize ...

  5. Glycoproteomic analysis of seven major allergenic proteins reveals novel post-translational modifications

    DEFF Research Database (Denmark)

    Halim, Adnan; Carlsson, Michael C; Mathiesen, Caroline Benedicte K

    2015-01-01

    Allergenic proteins such as grass pollen and house dust mite (HDM) proteins are known to trigger hypersensitivity reactions of the immune system, leading to what is commonly known as allergy. Key allergenic proteins including sequence variants have been identified but characterization of their post...... allergens. Moreover, we identified more complex glycan structures than previously reported on the major grass pollen group 1 and 5 allergens, implicating important roles for carbohydrates in allergen recognition and response by the immune system. The new findings are important for understanding basic...

  6. Detection of hydrolysis of lipid post-translational modifications during gel-electrophoresis-based proteomic protocol

    Czech Academy of Sciences Publication Activity Database

    Žídková, Jitka; Řehulka, Pavel; Chmelík, Josef

    2007-01-01

    Roč. 7, č. 15 (2007), s. 2507-2510 ISSN 1615-9853 R&D Projects: GA MŠk 1M0570 Institutional research plan: CEZ:AV0Z40310501 Keywords : ester hydrolysis * gel electrophoresis * MALDI-TOF MS Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 5.479, year: 2007

  7. An integrative analysis of post-translational histone modifications in the marine diatom Phaeodactylum tricornutum

    KAUST Repository

    Veluchamy, Alaguraj; Rastogi, Achal; Lin, Xin; Lombard, Bé rangè re; Murik, Omer; Thomas, Yann; Dingli, Florent; Rivarola, Maximo; Ott, Sandra; Liu, Xinyue; Sun, Yezhou; Rabinowicz, Pablo D.; McCarthy, James; Allen, Andrew E.; Loew, Damarys; Bowler, Chris; Tirichine, Leï la

    2015-01-01

    in global biogeochemical cycles. Dissecting chromatin-mediated regulation of genes in diatoms will help understand the ecological success of these organisms in contemporary oceans. Results: Here, we use high resolution mass spectrometry to identify a full

  8. Protein Glycosylation in Archaea: A Post-Translational Modification to Enhance Extremophilic Protein Stability

    Science.gov (United States)

    2010-01-15

    Analysis of the chemical composition of the Asn-linked polysaccharides decorating many archaeal proteins has revealed the use of a wider variety of sugar...reminiscent of the eukaryal glycan-charged lipid, linked to a variety of monosaccharides , including glucose, mannose, and N-acetylglucosamine (GlcNAc

  9. Functional Characterization of CENP-A Post-Translational Modifications in Chromosome Segregation

    Science.gov (United States)

    2016-09-01

    Yamaguchi S, Oohashi T, Shimada H, Ochiai T, Yoda K, Nomura F. Overexpression and mistargeting of centromere protein-A in human primary colorectal...of cell biology. 2007;176(6):795-805. 9. Tomonaga T, Matsushita K, Yamaguchi S, Oohashi T, Shimada H, Ochiai T, Yoda K, Nomura F. Overexpression and

  10. The roles of MHC class II genes and post-translational modification in celiac disease.

    Science.gov (United States)

    Sollid, Ludvig M

    2017-08-01

    Our increasing understanding of the etiology of celiac disease, previously considered a simple food hypersensitivity disorder caused by an immune response to cereal gluten proteins, challenges established concepts of autoimmunity. HLA is a chief genetic determinant, and certain HLA-DQ allotypes predispose to the disease by presenting posttranslationally modified (deamidated) gluten peptides to CD4 + T cells. The deamidation of gluten peptides is mediated by transglutaminase 2. Strikingly, celiac disease patients generate highly disease-specific autoantibodies to the transglutaminase 2 enzyme. The dual role of transglutaminase 2 in celiac disease is hardly coincidental. This paper reviews the genetic mapping and involvement of MHC class II genes in disease pathogenesis, and discusses the evidence that MHC class II genes, via the involvement of transglutaminase 2, influence the generation of celiac disease-specific autoantibodies.

  11. Near infrared light induces post-translational modifications of human red blood cell proteins.

    Science.gov (United States)

    Walski, Tomasz; Dyrda, Agnieszka; Dzik, Małgorzata; Chludzińska, Ludmiła; Tomków, Tomasz; Mehl, Joanna; Detyna, Jerzy; Gałecka, Katarzyna; Witkiewicz, Wojciech; Komorowska, Małgorzata

    2015-11-01

    There is a growing body of evidence that near infrared (NIR) light exerts beneficial effects on cells. Its usefulness in the treatment of cancer, acute brain injuries, strokes and neurodegenerative disorders has been proposed. The mechanism of the NIR action is probably of photochemical nature, however it is not fully understood. Here, using a relatively simple biological model, human red blood cells (RBCs), and a polychromatic non-polarized light source, we investigate the impact of NIR radiation on the oxygen carrier, hemoglobin (Hb), and anion exchanger (AE1, Band 3). The exposure of intact RBCs to NIR light causes quaternary transitions in Hb, dehydration of proteins and decreases the amount of physiologically inactive methemoglobin, as detected by Raman spectroscopy. These effects are accompanied by a lowering of the intracellular pH (pHi) and changes in the cell membrane topography, as documented by atomic force microscopy (AFM). All those changes are in line with our previous studies where alterations of the membrane fluidity and membrane potential were attributed to NIR action on RBCs. The rate of the above listed changes depends strictly on the dose of NIR light that the cells receive, nonetheless it should not be considered as a thermal effect.

  12. Do post-translational beta cell protein modifications trigger type 1 diabetes?

    DEFF Research Database (Denmark)

    Størling, Joachim; Overgaard, Anne Julie; Brorsson, Caroline Anna

    2013-01-01

    beta cell-specific neo-epitopes. We suggest that the current paradigm of type 1 diabetes as a classical autoimmune disease should be reconsidered since the immune response may not be directed against native beta cell proteins. A modified model for the pathogenetic events taking place in islets leading...... diabetes exists in the published literature. Furthermore, we report that cytokines change the expression levels of several genes encoding proteins involved in PTM processes in human islets, and that there are type 1 diabetes-associated polymorphisms in a number of these. In conclusion, data from...... the literature and presented experimental data support the notion that PTM of beta cell proteins may be involved in triggering beta cell destruction in type 1 diabetes. If the beta cell antigens recognised by the immune system foremost come from modified proteins rather than native ones, the concept of type 1...

  13. Pathogenic leptospires modulate protein expression and post-translational modifications in response to mammalian host signals

    Science.gov (United States)

    Pathogenic species of Leptospira cause leptospirosis, a bacterial zoonotic disease with a global distribution affecting over one million people annually. Reservoir hosts of leptospirosis, including rodents, dogs and cattle, exhibit little to no signs of disease but shed large numbers of organisms in...

  14. Obstructing Androgen Receptor Activation in Prostate Cancer Cells Through Post-translational Modification by NEDD8

    Science.gov (United States)

    2012-11-01

    FACS flow cytometer analysis . In addition, we will measure the steady state protein level of p53, p21, p27, and pRb. In the Jab1 silencing cell...affected by DHT treatment, and the endogenous AR level was not affected by Jab1 silencing. Interestingly, Western blot analysis of immunoprecipitated AR...Avantaggiati, and R. G. Pestell . 2003. Acetylation of androgen receptor enhances coactivator binding and promotes prostate cancer cell growth. Mol

  15. Post-translational modification of host proteins in pathogen-triggered defence signalling in plants

    NARCIS (Netherlands)

    Stulemeijer, I.J.E.; Joosten, M.H.A.J.

    2008-01-01

    Microbial plant pathogens impose a continuous threat to global food production. Similar to animals, an innate immune system allows plants to recognize pathogens and swiftly activate defence. To activate a rapid response, receptor-mediated pathogen perception and subsequent downstream signalling

  16. Exploring the diversity of protein modifications: special bacterial phosphorylation systems

    DEFF Research Database (Denmark)

    Mijakovic, Ivan; Grangeasse, Christophe; Turgay, Kürşad

    2016-01-01

    Protein modifications not only affect protein homeostasis but can also establish new cellular protein functions and are important components of complex cellular signal sensing and transduction networks. Among these post-translational modifications, protein phosphorylation represents the one that ...

  17. Dynamic association with donor cell filopodia and lipid-modification are essential features of Wnt8a during patterning of the zebrafish neuroectoderm.

    Directory of Open Access Journals (Sweden)

    Marta Luz

    Full Text Available BACKGROUND: Wnt proteins are conserved signaling molecules that regulate pattern formation during animal development. Many Wnt proteins are post-translationally modified by addition of lipid adducts. Wnt8a provides a crucial signal for patterning the anteroposterior axis of the developing neural plate in vertebrates. However, it is not clear how this protein propagates from its source, the blastoderm margin, to the target cells in the prospective neural plate, and how lipid-modifications might influence Wnt8a propagation and activity. RESULTS: We have dynamically imaged biologically active, fluorescently tagged Wnt8a in living zebrafish embryos. We find that Wnt8a localizes to membrane-associated, punctate structures in live tissue. In Wnt8a expressing cells, these puncta are found on filopodial cellular processes, from where the protein can be released. In addition, Wnt8a is found colocalized with Frizzled receptor-containing clusters on signal receiving cells. Combining in vitro and in vivo assays, we compare the roles of conserved Wnt8a residues in cell and non-cell-autonomous signaling activity and secretion. Non-signaling Wnt8 variants show these residues can regulate Wnt8a distribution in producing cell membranes and filopodia as well as in the receiving tissue. CONCLUSIONS: Together, our results show that Wnt8a forms dynamic clusters found on filopodial donor cell and on signal receiving cell membranes. Moreover, they demonstrate a differential requirement of conserved residues in Wnt8a protein for distribution in producing cells and receiving tissue and signaling activity during neuroectoderm patterning.

  18. Dynamic association with donor cell filopodia and lipid-modification are essential features of Wnt8a during patterning of the zebrafish neuroectoderm.

    Science.gov (United States)

    Luz, Marta; Spannl-Müller, Stephanie; Özhan, Günes; Kagermeier-Schenk, Birgit; Rhinn, Muriel; Weidinger, Gilbert; Brand, Michael

    2014-01-01

    Wnt proteins are conserved signaling molecules that regulate pattern formation during animal development. Many Wnt proteins are post-translationally modified by addition of lipid adducts. Wnt8a provides a crucial signal for patterning the anteroposterior axis of the developing neural plate in vertebrates. However, it is not clear how this protein propagates from its source, the blastoderm margin, to the target cells in the prospective neural plate, and how lipid-modifications might influence Wnt8a propagation and activity. We have dynamically imaged biologically active, fluorescently tagged Wnt8a in living zebrafish embryos. We find that Wnt8a localizes to membrane-associated, punctate structures in live tissue. In Wnt8a expressing cells, these puncta are found on filopodial cellular processes, from where the protein can be released. In addition, Wnt8a is found colocalized with Frizzled receptor-containing clusters on signal receiving cells. Combining in vitro and in vivo assays, we compare the roles of conserved Wnt8a residues in cell and non-cell-autonomous signaling activity and secretion. Non-signaling Wnt8 variants show these residues can regulate Wnt8a distribution in producing cell membranes and filopodia as well as in the receiving tissue. Together, our results show that Wnt8a forms dynamic clusters found on filopodial donor cell and on signal receiving cell membranes. Moreover, they demonstrate a differential requirement of conserved residues in Wnt8a protein for distribution in producing cells and receiving tissue and signaling activity during neuroectoderm patterning.

  19. Identification of nuclear protein targets for six leukemogenic tyrosine kinases governed by post-translational regulation.

    Directory of Open Access Journals (Sweden)

    Andrew Pierce

    Full Text Available Mutated tyrosine kinases are associated with a number of different haematological malignancies including myeloproliferative disorders, lymphoma and acute myeloid leukaemia. The potential commonalities in the action of six of these leukemogenic proteins on nuclear proteins were investigated using systematic proteomic analysis. The effects on over 3600 nuclear proteins and 1500 phosphopeptide sites were relatively quantified in seven isogenic cell lines. The effects of the kinases were diverse although some commonalities were found. Comparison of the nuclear proteomic data with transcriptome data and cytoplasmic proteomic data indicated that the major changes are due to post-translational mechanisms rather than changes in mRNA or protein distribution. Analysis of the promoter regions of genes whose protein levels changed in response to the kinases showed the most common binding site found was that for NFκB whilst other sites such as those for the glucocorticoid receptor were also found. Glucocorticoid receptor levels and phosphorylation were decreased by all 6 PTKs. Whilst Glucocorticoid receptor action can potentiate NFκB action those proteins where genes have NFκB binding sites were in often regulated post-translationally. However all 6 PTKs showed evidence of NFkB pathway modulation via activation via altered IkB and NFKB levels. Validation of a common change was also undertaken with PMS2, a DNA mismatch repair protein. PMS2 nuclear levels were decreased in response to the expression of all 6 kinases, with no concomitant change in mRNA level or cytosolic protein level. Response to thioguanine, that requires the mismatch repair pathway, was modulated by all 6 oncogenic kinases. In summary common targets for 6 oncogenic PTKs have been found that are regulated by post-translational mechanisms. They represent potential new avenues for therapies but also demonstrate the post-translational regulation is a key target of leukaemogenic kinases.

  20. Modifications of center-surround, spot detection and dot-pattern selective operators

    NARCIS (Netherlands)

    Petkov, Nicolai; Visser, Wicher T.

    2005-01-01

    This paper describes modifications of the models of center-surround and dot-pattern selective cells proposed previously. These modifications concern mainly the normalization of the difference of Gaussians (DoG) function used to model center-surround receptive fields, the normalization of

  1. Novel method for chemical modification and patterning of the SU-8 photoresist

    DEFF Research Database (Denmark)

    Blagoi, Gabriela; Keller, Stephan Urs; Boisen, Anja

    2007-01-01

    the wetting behaviour of SU-8. The resolution limit of the AQ photopatterning method was 20 μm when using an uncollimated light source. AQ modification followed by a reaction with amino groups of Alexa-647 cadaverine and a Biotin-amino derivative proved possible modification and patterning of polymeric...

  2. Photochemical modification and patterning of SU-8 using Anthraquinone photolinkers

    DEFF Research Database (Denmark)

    Blagoi, Gabriela; Keller, Stephan Urs; Persson, Karl Fredrik

    2008-01-01

    -dimensional patterns on a Novolac A derivative polymer (SU-8) and, subsequently, their functionalization with biomolecules. Anthraquinone (AQ) derivatives are used to chemically modify and pattern SU-8 surfaces. Features as small as 20 μm are obtained when using uncollimated light. The X−Y spatial resolution...

  3. Modification of Surface Energy via Direct Laser Ablative Surface Patterning

    Science.gov (United States)

    Wohl, Christopher J., Jr. (Inventor); Belcher, Marcus A. (Inventor); Connell, John W. (Inventor); Hopkins, John W. (Inventor)

    2015-01-01

    Surface energy of a substrate is changed without the need for any template, mask, or additional coating medium applied to the substrate. At least one beam of energy directly ablates a substrate surface to form a predefined topographical pattern at the surface. Each beam of energy has a width of approximately 25 micrometers and an energy of approximately 1-500 microJoules. Features in the topographical pattern have a width of approximately 1-500 micrometers and a height of approximately 1.4-100 micrometers.

  4. Characterization and modification of cavitation pattern in shock wave lithotripsy

    Science.gov (United States)

    Arora, Manish; Ohl, Claus Dieter; Liebler, Marko

    2004-01-01

    The temporal and spatial dynamics of cavitation bubble cloud growth and collapse in extracorporeal shock wave lithotripsy (ESWL) is studied experimentally. The first objective is obtaining reproducible cloud patterns experimentally and comparing them with FDTD-calculations. Second, we describe a method to modify the cavitation pattern by timing two consecutive pressure waves at variable delays. It is found that the spatial and temporal dynamics of the cavitation bubble can be varied in large ranges. The ability to control cavitation dynamics allows discussing strategies for improvement of medical and biological applications of shock waves such as cell membrane poration and stone fragmentation.

  5. Characterization and modification of cavitation pattern in shock wave lithotripsy

    NARCIS (Netherlands)

    Arora, M.; Ohl, C.D.; Liebler, Marko

    2004-01-01

    The temporal and spatial dynamics of cavitation bubble cloud growth and collapse in extracorporeal shock wave lithotripsy (ESWL) is studied experimentally. The first objective is obtaining reproducible cloud patterns experimentally and comparing them with FDTD-calculations. Second, we describe a

  6. Analyzing the evolution of beta-endorphin post-translational processing events: studies on reptiles.

    Science.gov (United States)

    Shoureshi, Pezhman; Baron, Andrea; Szynskie, Laura; Dores, Robert M

    2007-01-01

    In many cartilaginous fishes, most ray-finned fishes, lungfishes, and amphibians, the post-translational processing of POMC includes the monobasic cleavage of beta-endorphin to yield an opioid that is eight to ten amino acids in length. The amino acid motif within the beta-endorphin sequence required for a monobasic cleavage event is -E-R-(S/G)-Q-. Mammals and birds lack this motif and as a result beta-endorphin(1-8) is a not an end-product in either group. Since both mammals and birds were derived from ancestors with reptilian origins, an analysis of beta-endorphin sequences from extant groups of reptiles should provide insights into the manner in which beta-endorphin post-translational processing mechanisms have evolved in amniotes. To this end a POMC cDNA was cloned from the pituitary of the turtle, Chrysemys scripta. The beta-endorphin sequence in this species was compared to other reptile beta-endorphin sequences (i.e., Chinese soft shell turtle and gecko) and to known bird and mammal sequences. This analysis indicated that either the loss of the arginine residue at the cleavage site (the two turtle species, chick, and human) or a substitution at the glutamine position in the consensus sequence (gecko and ostrich) would account for the loss of the monobasic cleavage reaction in that species. Since amphibians are capable of performing the beta-endorphin monobasic reaction, it would appear that the amino acid substitutions that eliminated this post-translational process event in reptilian-related tetrapods must have occurred in the ancestral amniotes.

  7. Regulation of multispanning membrane protein topology via post-translational annealing.

    Science.gov (United States)

    Van Lehn, Reid C; Zhang, Bin; Miller, Thomas F

    2015-09-26

    The canonical mechanism for multispanning membrane protein topogenesis suggests that protein topology is established during cotranslational membrane integration. However, this mechanism is inconsistent with the behavior of EmrE, a dual-topology protein for which the mutation of positively charged loop residues, even close to the C-terminus, leads to dramatic shifts in its topology. We use coarse-grained simulations to investigate the Sec-facilitated membrane integration of EmrE and its mutants on realistic biological timescales. This work reveals a mechanism for regulating membrane-protein topogenesis, in which initially misintegrated configurations of the proteins undergo post-translational annealing to reach fully integrated multispanning topologies. The energetic barriers associated with this post-translational annealing process enforce kinetic pathways that dictate the topology of the fully integrated proteins. The proposed mechanism agrees well with the experimentally observed features of EmrE topogenesis and provides a range of experimentally testable predictions regarding the effect of translocon mutations on membrane protein topogenesis.

  8. Chemically Patterned Inverse Opal Created by a Selective Photolysis Modification Process.

    Science.gov (United States)

    Tian, Tian; Gao, Ning; Gu, Chen; Li, Jian; Wang, Hui; Lan, Yue; Yin, Xianpeng; Li, Guangtao

    2015-09-02

    Anisotropic photonic crystal materials have long been pursued for their broad applications. A novel method for creating chemically patterned inverse opals is proposed here. The patterning technique is based on selective photolysis of a photolabile polymer together with postmodification on released amine groups. The patterning method allows regioselective modification within an inverse opal structure, taking advantage of selective chemical reaction. Moreover, combined with the unique signal self-reporting feature of the photonic crystal, the fabricated structure is capable of various applications, including gradient photonic bandgap and dynamic chemical patterns. The proposed method provides the ability to extend the structural and chemical complexity of the photonic crystal, as well as its potential applications.

  9. Estimation of color modification in digital images by CFA pattern change.

    Science.gov (United States)

    Choi, Chang-Hee; Lee, Hae-Yeoun; Lee, Heung-Kyu

    2013-03-10

    Extensive studies have been carried out for detecting image forgery such as copy-move, re-sampling, blurring, and contrast enhancement. Although color modification is a common forgery technique, there is no reported forensic method for detecting this type of manipulation. In this paper, we propose a novel algorithm for estimating color modification in images acquired from digital cameras when the images are modified. Most commercial digital cameras are equipped with a color filter array (CFA) for acquiring the color information of each pixel. As a result, the images acquired from such digital cameras include a trace from the CFA pattern. This pattern is composed of the basic red green blue (RGB) colors, and it is changed when color modification is carried out on the image. We designed an advanced intermediate value counting method for measuring the change in the CFA pattern and estimating the extent of color modification. The proposed method is verified experimentally by using 10,366 test images. The results confirmed the ability of the proposed method to estimate color modification with high accuracy. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

  10. Ribosomally Synthesized and Post-translationally Modified Peptide Natural Products: New Insights Into the Role of Leader and Core Peptides During Biosynthesis

    Science.gov (United States)

    Yang, Xiao; van der Donk, Wilfred A.

    2013-01-01

    Ribosomally synthesized and post-translationally modified peptides (RiPPs) are a major class of natural products with a high degree of structural diversity and a wide variety of bioactivities. Understanding the biosynthetic machinery of these RiPPs will benefit the discovery and development of new molecules with potential pharmaceutical applications. In this review, we discuss the features of the biosynthetic pathways to different RiPP classes, and propose mechanisms regarding recognition of the precursor peptide by the posttranslational modification enzymes. We propose that the leader peptides function as allosteric regulators that bind the active form of the biosynthetic enzymes in a conformational selection process. We also speculate how enzymes that generate polycyclic products of defined topologies may have been selected for during evolution. PMID:23666908

  11. Phycourobilin in Trichromatic Phycocyanin from Oceanic Cyanobacteria Is Formed Post-translationally by a Phycoerythrobilin Lyase-Isomerase*S⃞

    Science.gov (United States)

    Blot, Nicolas; Wu, Xian-Jun; Thomas, Jean-Claude; Zhang, Juan; Garczarek, Laurence; Böhm, Stephan; Tu, Jun-Ming; Zhou, Ming; Plöscher, Matthias; Eichacker, Lutz; Partensky, Frédéric; Scheer, Hugo; Zhao, Kai-Hong

    2009-01-01

    Most cyanobacteria harvest light with large antenna complexes called phycobilisomes. The diversity of their constituting phycobiliproteins contributes to optimize the photosynthetic capacity of these microorganisms. Phycobiliprotein biosynthesis, which involves several post-translational modifications including covalent attachment of the linear tetrapyrrole chromophores (phycobilins) to apoproteins, begins to be well understood. However, the biosynthetic pathway to the blue-green-absorbing phycourobilin (λmax ∼ 495 nm) remained unknown, although it is the major phycobilin of cyanobacteria living in oceanic areas where blue light penetrates deeply into the water column. We describe a unique trichromatic phycocyanin, R-PC V, extracted from phycobilisomes of Synechococcus sp. strain WH8102. It is evolutionarily remarkable as the only chromoprotein known so far that absorbs the whole wavelength range between 450 and 650 nm. R-PC V carries a phycourobilin chromophore on its α-subunit, and this can be considered an extreme case of adaptation to blue-green light. We also discovered the enzyme, RpcG, responsible for its biosynthesis. This monomeric enzyme catalyzes binding of the green-absorbing phycoerythrobilin at cysteine 84 with concomitant isomerization to phycourobilin. This reaction is analogous to formation of the orange-absorbing phycoviolobilin from the red-absorbing phycocyanobilin that is catalyzed by the lyase-isomerase PecE/F in some freshwater cyanobacteria. The fusion protein, RpcG, and the heterodimeric PecE/F are mutually interchangeable in a heterologous expression system in Escherichia coli. The novel R-PC V likely optimizes rod-core energy transfer in phycobilisomes and thereby adaptation of a major phytoplankton group to the blue-green light prevailing in oceanic waters. PMID:19182270

  12. Post-translational transformation of methionine to aspartate is catalyzed by heme iron and driven by peroxide: a novel subunit-specific mechanism in hemoglobin.

    Science.gov (United States)

    Strader, Michael Brad; Hicks, Wayne A; Kassa, Tigist; Singleton, Eileen; Soman, Jayashree; Olson, John S; Weiss, Mitchell J; Mollan, Todd L; Wilson, Michael T; Alayash, Abdu I

    2014-08-08

    A pathogenic V67M mutation occurs at the E11 helical position within the heme pockets of variant human fetal and adult hemoglobins (Hb). Subsequent post-translational modification of Met to Asp was reported in γ subunits of human fetal Hb Toms River (γ67(E11)Val → Met) and β subunits of adult Hb (HbA) Bristol-Alesha (β67(E11)Val → Met) that were associated with hemolytic anemia. Using kinetic, proteomic, and crystal structural analysis, we were able to show that the Met → Asp transformation involves heme cycling through its oxoferryl state in the recombinant versions of both proteins. The conversion to Met and Asp enhanced the spontaneous autoxidation of the mutants relative to wild-type HbA and human fetal Hb, and the levels of Asp were elevated with increasing levels of hydrogen peroxide (H2O2). Using H2(18)O2, we verified incorporation of (18)O into the Asp carboxyl side chain confirming the role of H2O2 in the oxidation of the Met side chain. Under similar experimental conditions, there was no conversion to Asp at the αMet(E11) position in the corresponding HbA Evans (α62(E11)Val → Met). The crystal structures of the three recombinant Met(E11) mutants revealed similar thioether side chain orientations. However, as in the solution experiments, autoxidation of the Hb mutant crystals leads to electron density maps indicative of Asp(E11) formation in β subunits but not in α subunits. This novel post-translational modification highlights the nonequivalence of human Hb α, β, and γ subunits with respect to redox reactivity and may have direct implications to α/β hemoglobinopathies and design of oxidatively stable Hb-based oxygen therapeutics. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  13. Post-translational processing of synaptophysin in the rat retina is disrupted by diabetes.

    Directory of Open Access Journals (Sweden)

    Travis S D'Cruz

    Full Text Available Synaptophysin, is an abundant presynaptic protein involved in synaptic vesicle recycling and neurotransmitter release. Previous work shows that its content is significantly reduced in the rat retina by streptozotocin (STZ-diabetes. This study tested the hypothesis that STZ-diabetes alters synaptophysin protein turnover and glycosylation in the rat retina. Whole explant retinas from male Sprague-Dawley rats were used in this study. Rats were made diabetic by a single intraperitoneal STZ injection (65 mg/kg body weight in 10 mM sodium citrate, pH 4.5. mRNA translation was measured using a (35S-methionine labeling assay followed by synaptophysin immunoprecipitation and autoradiography. A pulse-chase study was used to determine the depletion of newly synthesized synaptophysin. Depletion of total synaptophysin was determined after treatment with cycloheximide. Mannose rich N-glycosylated synaptophysin was detected by treating retinal lysates with endoglycosidase H followed by immunoblot analysis. Synaptophysin mRNA translation was significantly increased after 1 month (p<0.001 and 2 months (p<0.05 of STZ-diabetes, compared to age-matched controls. Newly synthesized synaptophysin degradation was significantly accelerated in the retina after 1 and 2 months of diabetes compared to controls (p<0.05. Mannose rich glycosylated synaptophysin was significantly increased after 1 month of STZ-diabetes compared to controls (p<0.05.These data suggest that diabetes increases mRNA translation of synaptophysin in the retina, resulting in an accumulation of mannose rich glycosylated synaptophysin, a transient post-translational state of the protein. This diabetes-induced irregularity in post-translational processing could explain the accelerated degradation of retinal synaptophysin in diabetes.

  14. Assessment of modification level of hypoeutectic Al -Si alloys by pattern recognition of cooling curves

    Directory of Open Access Journals (Sweden)

    CHEN Xiang

    2005-11-01

    Full Text Available Most evaluations of modification level are done according to a specific scale based on an merican Foundry Society (AFS standard wall chart as qualitative analysis in Al-Si casting production currently. This method is quite dependent on human experience when making comparisons of the microstructure with the standard chart. And the structures depicted in the AFS chart do not always resemble those seen in actual Al-Si castings. Therefore, this ualitativeanalysis procedure is subjective and can introduce human-caused errors into comparative metallographic analyses. A quantization parameter of the modification level was introduced by setting up the relationship between mean area weighted shape factor of eutectic silicon phase and the modification level using image analysis technology. In order to evaluate the modification level, a new method called "intelligent evaluating of melt quality by pattern recognition of hermal analysis cooling curves" has also been introduced. The results show that silicon modification level can be precisely assessed by comparison of the cooling curve of the melt to be evaluated with the one most similar to it in a database.

  15. Peak-valley-peak pattern of histone modifications delineates active regulatory elements and their directionality

    DEFF Research Database (Denmark)

    Pundhir, Sachin; Bagger, Frederik Otzen; Lauridsen, Felicia Kathrine Bratt

    2016-01-01

    Formation of nucleosome free region (NFR) accompanied by specific histone modifications at flanking nucleosomes is an important prerequisite for enhancer and promoter activity. Due to this process, active regulatory elements often exhibit a distinct shape of histone signal in the form of a peak......-valley-peak (PVP) pattern. However, different features of PVP patterns and their robustness in predicting active regulatory elements have never been systematically analyzed. Here, we present PARE, a novel computational method that systematically analyzes the H3K4me1 or H3K4me3 PVP patterns to predict NFRs. We show...... four ENCODE cell lines and four hematopoietic differentiation stages, we identified several enhancers whose regulatory activity is stage specific and correlates positively with the expression of proximal genes in a particular stage. In conclusion, our results demonstrate that PVP patterns delineate...

  16. Photoreactive molecular layers containing aryl ester units: Preparation, UV patterning and post-exposure modification

    Energy Technology Data Exchange (ETDEWEB)

    Hoefler, Thomas [Institute for Chemistry and Technology of Materials, Graz University of Technology, Stremayrgasse 16, A-8010 Graz (Austria); Track, Anna M. [Institute of Solid State Physics, Graz University of Technology, Petersgasse 16/II, A-8010 Graz (Austria); Institute of Physics, University of Graz, Universitaetsplatz 5, A-8010 Graz (Austria); Pacher, Peter [Institute of Solid State Physics, Graz University of Technology, Petersgasse 16/II, A-8010 Graz (Austria); Shen, Quan [Institute of Physics, Montanuniversitaet Leoben, Franz-Josef-Strasse 18, A-8700 Leoben (Austria); Flesch, Heinz-Georg [Institute of Solid State Physics, Graz University of Technology, Petersgasse 16/II, A-8010 Graz (Austria); Hlawacek, Gregor [Institute of Physics, Montanuniversitaet Leoben, Franz-Josef-Strasse 18, A-8700 Leoben (Austria); Koller, Georg; Ramsey, Michael G. [Institute of Physics, University of Graz, Universitaetsplatz 5, A-8010 Graz (Austria); Schennach, Robert; Resel, Roland [Institute of Solid State Physics, Graz University of Technology, Petersgasse 16/II, A-8010 Graz (Austria); Teichert, Christian [Institute of Physics, Montanuniversitaet Leoben, Franz-Josef-Strasse 18, A-8700 Leoben (Austria); Kern, Wolfgang [Institute of Chemistry of Polymers, Montanuniversitaet Leoben, Franz-Josef-Strasse 18, A-8700 Leoben (Austria); Trimmel, Gregor [Institute for Chemistry and Technology of Materials, Graz University of Technology, Stremayrgasse 16, A-8010 Graz (Austria); Griesser, Thomas, E-mail: thomas.griesser@unileoben.ac.at [Institute for Chemistry and Technology of Materials, Graz University of Technology, Stremayrgasse 16, A-8010 Graz (Austria); Institute of Chemistry of Polymers, Montanuniversitaet Leoben, Franz-Josef-Strasse 18, A-8700 Leoben (Austria)

    2010-01-15

    The photolithographic modification of thin functional silane layers provides a versatile and powerful means of fabricating functionalized patterned surfaces which can be applied for tuning inorganic surface properties and for modern immobilisation techniques. In this contribution we present the synthesis of a new functional trichloro organosilane bearing photoreactive aryl ester groups and its application in thin silane layers on silicon oxide surfaces. Whereas the trichlorosilyl group acts as anchoring unit to the inorganic surface, the aryl ester group undergoes the photo-Fries rearrangement to yield hydroxyketones upon irradiation with UV-light of 254 nm which leads to a change in chemical reactivity of the surface. By a subsequent reaction with perfluorobutyryl chloride, the photogenerated hydroxy groups yield the corresponding perfluorinated ester compound, which allows further tuning of surface properties. The layer formation as well as the photoreaction and post-modification reaction was monitored by FTIR spectroscopy and X-ray photoelectron spectroscopy (XPS). The thickness of the obtained thin layers was determined by X-ray reflectivity (XRR). Photopatterned surfaces were produced using a contact mask during illumination followed by the post-modification reaction. Friction force microscopy (FFM) revealed the contrast between modified and unmodified regions of the patterned samples.

  17. TG2 regulates the heat-shock response by the post-translational modification of HSF1.

    Science.gov (United States)

    Rossin, Federica; Villella, Valeria Rachela; D'Eletto, Manuela; Farrace, Maria Grazia; Esposito, Speranza; Ferrari, Eleonora; Monzani, Romina; Occhigrossi, Luca; Pagliarini, Vittoria; Sette, Claudio; Cozza, Giorgio; Barlev, Nikolai A; Falasca, Laura; Fimia, Gian Maria; Kroemer, Guido; Raia, Valeria; Maiuri, Luigi; Piacentini, Mauro

    2018-05-11

    Heat-shock factor 1 (HSF1) is the master transcription factor that regulates the response to proteotoxic stress by controlling the transcription of many stress-responsive genes including the heat-shock proteins. Here, we show a novel molecular mechanism controlling the activation of HSF1. We demonstrate that transglutaminase type 2 (TG2), dependent on its protein disulphide isomerase activity, triggers the trimerization and activation of HSF1 regulating adaptation to stress and proteostasis impairment. In particular, we find that TG2 loss of function correlates with a defect in the nuclear translocation of HSF1 and in its DNA-binding ability to the HSP70 promoter. We show that the inhibition of TG2 restores the unbalance in HSF1-HSP70 pathway in cystic fibrosis (CF), a human disorder characterized by deregulation of proteostasis. The absence of TG2 leads to an increase of about 40% in CFTR function in a new experimental CF mouse model lacking TG2. Altogether, these results indicate that TG2 plays a key role in the regulation of cellular proteostasis under stressful cellular conditions through the modulation of the heat-shock response. © 2018 The Authors.

  18. Global analysis of myocardial peptides containing cysteines with irreversible sulfinic and sulfonic Acid post-translational modifications

    DEFF Research Database (Denmark)

    Paulech, Jana; Liddy, Kiersten A; Engholm-Keller, Kasper

    2015-01-01

    ) and others (Cys sulfinic [Cys-SO2H] and sulfonic [Cys-SO3H] acids) that are considered "irreversible." We developed an enrichment method to isolate Cys-SO2H/SO3H-containing peptides from complex tissue lysates that is compatible with tandem mass spectrometry (MS/MS). The acidity of these post...

  19. Identification of Disease Relevant Post Translational Modifications of Proteins in Pulmonary Fibrosis as Novel Biochemical Marker Targets

    DEFF Research Database (Denmark)

    Kristensen, Jacob Hull

    elastin and the ELM7 neo-epitope with limited reactivity towards intact elastin. Finally, we tested the assays for clinical relevance in serum from patients diagnosed with IPF or lung cancer and healthy matched controls. Serum EL-NE- and ELM7 fragment levels were significantly elevated in IPF- and lung...... cancer patients compared to matched controls. In conclusion, we have developed two technically stable assays, EL-NE and ELM7, for the quantification of elastin degraded by NE and MMP-7 respectively. Both assays were protease specific. Initial clinical testing suggested clinical relevance of the assays...

  20. Middle-down hybrid chromatography/tandem mass spectrometry workflow for characterization of combinatorial post-translational modifications in histones

    DEFF Research Database (Denmark)

    Sidoli, Simone; Schwämmle, Veit; Ruminowicz, Chrystian

    2014-01-01

    chromatography (WCX-HILIC) interfaced directly to high mass accuracy ESI MS/MS using electron transfer dissociation (ETD). This enabled automated and efficient separation and sequencing of hypermodified histone N-terminal tails for unambiguous localization of combinatorial PTMs. We present Histone Coder and Iso...

  1. Systems Level Analysis of Histone H3 Post-translational Modifications (PTMs) Reveals Features of PTM Crosstalk in Chromatin Regulation

    DEFF Research Database (Denmark)

    Schwämmle, Veit; Sidoli, Simone; Ruminowicz, Chrystian

    2016-01-01

    molecules contain multiple coexisting PTMs, some of which exhibit crosstalk, i.e. coordinated or mutually exclusive activities. Here, we present an integrated experimental and computational systems level molecular characterization of histone PTMs and PTM crosstalk. Using wild type and engineered mouse....... We characterized combinatorial PTM features across the four mESC lines and then applied statistical data analysis to predict crosstalk between histone H3 PTMs. We detected an overrepresentation of positive crosstalk (codependent marks) between adjacent mono-methylated and acetylated marks......, and negative crosstalk (mutually exclusive marks) among most of the seven characterized di- and tri-methylated lysine residues in the H3 tails. We report novel features of PTM interplay involving hitherto poorly characterized arginine methylation and lysine methylation sites, including H3R2me, H3R8me and H3K37...

  2. Middle-down hybrid chromatography/tandem mass spectrometry workflow for characterization of combinatorial post-translational modifications in histones.

    Science.gov (United States)

    Sidoli, Simone; Schwämmle, Veit; Ruminowicz, Chrystian; Hansen, Thomas A; Wu, Xudong; Helin, Kristian; Jensen, Ole N

    2014-10-01

    We present an integrated middle-down proteomics platform for sensitive mapping and quantification of coexisting PTMs in large polypeptides (5-7 kDa). We combined an RP trap column with subsequent weak cation exchange-hydrophilic interaction LC interfaced directly to high mass accuracy ESI MS/MS using electron transfer dissociation. This enabled automated and efficient separation and sequencing of hypermodified histone N-terminal tails for unambiguous localization of combinatorial PTMs. We present Histone Coder and IsoScale software to extract, filter, and analyze MS/MS data, including quantification of cofragmenting isobaric polypeptide species. We characterized histone tails derived from murine embryonic stem cells knockout in suppressor of zeste12 (Suz12(-/-) ) and quantified 256 combinatorial histone marks in histones H3, H4, and H2A. Furthermore, a total of 713 different combinatorial histone marks were identified in purified histone H3. We measured a seven-fold reduction of H3K27me2/me3 (where me2 and me3 are dimethylation and trimethylation, respectively) in Suz12(-) (/) (-) cells and detected significant changes of the relative abundance of 16 other single PTMs of histone H3 and other combinatorial marks. We conclude that the inactivation of Suz12 is associated with changes in the abundance of not only H3K27 methylation but also multiple other PTMs in histone H3 tails. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  3. Regulation of Mammary Stem Cell Quiescence via Post-Translational Modification of DeltaNp63alpha

    Science.gov (United States)

    2012-12-01

    This document is the Annual Summary Report on the training grant awarded to Andrew DeCastro entitled Regulation of Mammary Stem Cell Quiescence via...screen) mediated phosphorylation of deltaNPdelta3 on stem cell behavior and mitotic activity. Task 1 aims to determine the effects of wild-type, phospho...ablative and phospho-mimetic alleles of deltaNP63delta phosphorylation on stem cell behavior in vitro. Thus far, we demonstrate that stem cell enriched

  4. Post-translational modifications of the extracellular matrix are key events in cancer progression: opportunities for biochemical marker development

    DEFF Research Database (Denmark)

    Leeming, D J; Bay-Jensen, A C; Vassiliadis, E

    2011-01-01

    -associated extracellular matrix (ECM) proteins. Furthermore, severe cellular stress and inflammation, caused by cancer, results in generation of PTMs, which will be distributed throughout the ECM. This gives rise to release of protein-specific fragments to the circulation. Here we highlight the importance of remodeling...... of the ECM in cancer and the generation of PTMs, which may be cancer specific and reflect disease progression; thus having potential for biochemical marker development....

  5. Chip-scale pattern modification method for equalizing residual layer thickness in nanoimprint lithography

    Science.gov (United States)

    Youn, Sung-Won; Suzuki, Kenta; Hiroshima, Hiroshi

    2018-06-01

    A software program for modifying a mold design to obtain a uniform residual layer thickness (RLT) distribution has been developed and its validity was verified by UV-nanoimprint lithography (UV-NIL) simulation. First, the effects of granularity (G) on both residual layer uniformity and filling characteristics were characterized. For a constant complementary pattern depth and a granularity that was sufficiently larger than the minimum pattern width, filling time decreased with the decrease in granularity. For a pattern design with a wide density range and an irregular distribution, the choice of a small granularity was not always a good strategy since the etching depth required for a complementary pattern occasionally exceptionally increased with the decrease in granularity. On basis of the results obtained, the automated method was applied to a chip-scale pattern modification. Simulation results showed a marked improvement in residual layer thickness uniformity for a capacity-equalized (CE) mold. For the given conditions, the standard deviation of RLT decreased in the range from 1/3 to 1/5 in accordance with pattern designs.

  6. Patterns of development of unspecific reaction of cells and modification of chemical protection

    International Nuclear Information System (INIS)

    Veksler, A.M.; Korystov, Yu.N.; Kublik, L.N.; Ehjdus, L.Kh.

    1980-01-01

    A study was made of a correlation between radioprotective efficiency of different chemical agents (weak electrolytes) and conditions of treatment. It was demonstrated that the pattern of changes in the protection efficiency, with modification thereof, is similar to that of the development of unspecific reaction and determined by the intracellular concentration of the chemical agents, which, in turn, is function of physicochemical parameters of the substance and pH gradient between cell and medium. With similar intracellular concentration, caffeine-benzoate, thioglicolic acid and caffeine proved to be equally effective, while the protective effect of cysteamine was appreciably higher

  7. Nanoscale patterning of a self-assembled monolayer by modification of the molecule–substrate bond

    Directory of Open Access Journals (Sweden)

    Cai Shen

    2014-03-01

    Full Text Available The intercalation of Cu at the interface of a self-assembled monolayer (SAM and a Au(111/mica substrate by underpotential deposition (UPD is studied as a means of high resolution patterning. A SAM of 2-(4'-methylbiphenyl-4-ylethanethiol (BP2 prepared in a structural phase that renders the Au substrate completely passive against Cu-UPD, is patterned by modification with the tip of a scanning tunneling microscope. The tip-induced defects act as nucleation sites for Cu-UPD. The lateral diffusion of the metal at the SAM–substrate interface and, thus, the pattern dimensions are controlled by the deposition time. Patterning down to the sub-20 nm range is demonstrated. The difference in strength between the S–Au and S–Cu bond is harnessed to develop the latent Cu-UPD image into a patterned binary SAM. Demonstrated by the exchange of BP2 by adamantanethiol (AdSH this is accomplished by a sequence of reductive desorption of BP2 in Cu free areas followed by adsorption of AdSH. The appearance of Au adatom islands upon the thiol exchange suggests that the interfacial structures of BP2 and AdSH SAMs are different.

  8. SMRT Sequencing Revealed Mitogenome Characteristics and Mitogenome-Wide DNA Modification Pattern in Ophiocordyceps sinensis.

    Science.gov (United States)

    Kang, Xincong; Hu, Liqin; Shen, Pengyuan; Li, Rui; Liu, Dongbo

    2017-01-01

    Single molecule, real-time (SMRT) sequencing was used to characterize mitochondrial (mt) genome of Ophiocordyceps sinensis and to analyze the mt genome-wide pattern of epigenetic DNA modification. The complete mt genome of O. sinensis , with a size of 157,539 bp, is the fourth largest Ascomycota mt genome sequenced to date. It contained 14 conserved protein-coding genes (PCGs), 1 intronic protein rps3 , 27 tRNAs and 2 rRNA subunits, which are common characteristics of the known mt genomes in Hypocreales. A phylogenetic tree inferred from 14 PCGs in Pezizomycotina fungi supports O. sinensis as most closely related to Hirsutella rhossiliensis in Ophiocordycipitaceae. A total of 36 sequence sites in rps3 were under positive selection, with dN/dS >1 in the 20 compared fungi. Among them, 16 sites were statistically significant. In addition, the mt genome-wide base modification pattern of O. sinensis was determined in this study, especially DNA methylation. The methylations were located in coding and uncoding regions of mt PCGs in O. sinensis , and might be closely related to the expression of PCGs or the binding affinity of transcription factor A to mtDNA. Consequently, these methylations may affect the enzymatic activity of oxidative phosphorylation and then the mt respiratory rate; or they may influence mt biogenesis. Therefore, methylations in the mitogenome of O. sinensis might be a genetic feature to adapt to the cold and low PO 2 environment at high altitude, where O. sinensis is endemic. This is the first report on epigenetic modifications in a fungal mt genome.

  9. Proliferative response of the murine esophageal epithelium to radiation: Modification by food consumption patterns

    International Nuclear Information System (INIS)

    Burholt, D.R.

    1985-01-01

    The single layer of proliferative epithelial cells of the murine esophagus undergoes a sequence of damage and recovery following cytotoxic insult. The modification of both damage and proliferative recovery by the alteration of animal eating patterns was investigated following thoracic field irradiation through the determination of /sup 3/H-TdR incorporation into the esophagus along with selective counting of labeled nuclei and mitotic figures. Initial radiation-induced damage, as determined by /sup 3/H-TdR incorporation suppression and mitotic delay, is under normal conditions dependent on the time of day of treatment. This circadian sensitivity may be altered by changing the eating pattern of the animal. The proliferative recovery following single dose irradiation is also dependent on food consumption patterns: fasting immediately following treatment and then refeeding 2 days later results in a more rapid proliferative recovery than observed under control eating conditions, while reduced food consumption during the period of proliferative hyperplasia reduces the extent of recovery. During multifraction radiation schedules both the damage produced by a subsequent dose and recovery capacity are influenced by the food consumption pattern between fractions

  10. Chemical modifications of Au/SiO2 template substrates for patterned biofunctional surfaces.

    Science.gov (United States)

    Briand, Elisabeth; Humblot, Vincent; Landoulsi, Jessem; Petronis, Sarunas; Pradier, Claire-Marie; Kasemo, Bengt; Svedhem, Sofia

    2011-01-18

    The aim of this work was to create patterned surfaces for localized and specific biochemical recognition. For this purpose, we have developed a protocol for orthogonal and material-selective surface modifications of microfabricated patterned surfaces composed of SiO(2) areas (100 μm diameter) surrounded by Au. The SiO(2) spots were chemically modified by a sequence of reactions (silanization using an amine-terminated silane (APTES), followed by amine coupling of a biotin analogue and biospecific recognition) to achieve efficient immobilization of streptavidin in a functional form. The surrounding Au was rendered inert to protein adsorption by modification by HS(CH(2))(10)CONH(CH(2))(2)(OCH(2)CH(2))(7)OH (thiol-OEG). The surface modification protocol was developed by testing separately homogeneous SiO(2) and Au surfaces, to obtain the two following results: (i) SiO(2) surfaces which allowed the grafting of streptavidin, and subsequent immobilization of biotinylated antibodies, and (ii) Au surfaces showing almost no affinity for the same streptavidin and antibody solutions. The surface interactions were monitored by quartz crystal microbalance with dissipation monitoring (QCM-D), and chemical analyses were performed by polarization modulation-reflexion absorption infrared spectroscopy (PM-RAIRS) and X-ray photoelectron spectroscopy (XPS) to assess the validity of the initial orthogonal assembly of APTES and thiol-OEG. Eventually, microscopy imaging of the modified Au/SiO(2) patterned substrates validated the specific binding of streptavidin on the SiO(2)/APTES areas, as well as the subsequent binding of biotinylated anti-rIgG and further detection of fluorescent rIgG on the functionalized SiO(2) areas. These results demonstrate a successful protocol for the preparation of patterned biofunctional surfaces, based on microfabricated Au/SiO(2) templates and supported by careful surface analysis. The strong immobilization of the biomolecules resulting from the described

  11. Modification of medullary respiratory-related discharge patterns by behaviors and states of arousal.

    Science.gov (United States)

    Chang, F C

    1992-02-07

    The modulatory influences of behaviors and states of arousal on bulbar respiratory-related unit (RRU) discharge patterns were studied in an unanesthetized, freely behaving guinea pig respiratory model system. When fully instrumented, this model system permits concurrent monitoring and recording of (i) single units from either Bötzinger complex or nucleus para-ambiguus; (ii) electrocorticogram; and, (iii) diaphragmatic EMG. In addition to being used in surveys of RRU discharge patterns in freely behaving states, the model system also offered a unique opportunity in investigating the effects of pentobarbital on RRU discharge patterns before, throughout the course of, and during recovery from anesthesia. In anesthetized preparations, a particular RRU discharge pattern (such as tonic, incrementing or decrementing) typically displayed little, if any notable variation. The most striking development following pentobarbital was a state of progressive bradypnea attributable to a significantly augmented RRU cycle duration, burst duration and an increase in the RRU spike frequencies during anesthesia. In freely behaving states, medullary RRU activities rarely adhered to a fixed, immutable discharge pattern. More specifically, the temporal organization (such as burst duration, cycle duration, and the extent of modulation of within-burst spike frequencies) of RRU discharge patterns regularly showed complex and striking variations, not only with states of arousal (sleep/wakefulness, anesthesia) but also with discrete alterations in electrocorticogram (ECoG) activities and a multitude of on-going behavioral repertoires such as volitional movement, postural modification, phonation, mastication, deglutition, sniffing/exploratory behavior, alerting/startle reflexes. Only during sleep, and on occasions when the animal assumed a motionless, resting posture, could burst patterns of relatively invariable periodicity and uniform temporal attributes be observed. RRU activities during

  12. Chemical rescue of the post-translationally carboxylated lysine mutant of allantoinase and dihydroorotase by metal ions and short-chain carboxylic acids.

    Science.gov (United States)

    Ho, Ya-Yeh; Huang, Yen-Hua; Huang, Cheng-Yang

    2013-04-01

    Bacterial allantoinase (ALLase) and dihydroorotase (DHOase) are members of the cyclic amidohydrolase family. ALLase and DHOase possess similar binuclear metal centers in the active site in which two metals are bridged by a post-translationally carboxylated lysine. In this study, we determined the effects of carboxylated lysine and metal binding on the activities of ALLase and DHOase. Although DHOase is a metalloenzyme, purified DHOase showed high activity without additional metal supplementation in a reaction mixture or bacterial culture. However, unlike DHOase, ALLase had no activity unless some specific metal ions were added to the reaction mixture or culture. Substituting the metal binding sites H59, H61, K146, H186, H242, or D315 with alanine completely abolished the activity of ALLase. However, the K146C, K146D and K146E mutants of ALLase were still active with about 1-6% activity of the wild-type enzyme. These ALLase K146 mutants were found to have 1.4-1.7 mol metal per mole enzyme subunit, which may indicate that they still contained the binuclear metal center in the active site. The activity of the K146A mutant of the ALLase and the K103A mutant of DHOase can be chemically rescued by short-chain carboxylic acids, such as acetic, propionic, and butyric acids, but not by ethanol, propan-1-ol, and imidazole, in the presence of Co2+ or Mn2+ ions. However, the activity was still ~10-fold less than that of wild-type ALLase. Overall, these results indicated that the 20 natural basic amino acid residues were not sufficiently able to play the role of lysine. Accordingly, we proposed that during evolution, the post-translational modification of carboxylated lysine in the cyclic amidohydrolase family was selected for promoting binuclear metal center self-assembly and increasing the nucleophilicity of the hydroxide at the active site for enzyme catalysis. This kind of chemical rescue combined with site-directed mutagenesis may also be used to identify a binuclear metal

  13. Antibodies against post-translationally modified vimentin peptides in patients with rheumatoid arthritis

    Directory of Open Access Journals (Sweden)

    P. A. Kuznetsova

    2017-01-01

    Full Text Available Rheumatoid arthritis (RA is the most common autoimmune rheumatic disease (ARD associated with the production of broad-spectrum antibodies, the detection of which is of important diagnostic and prognostic values. The problems of RA diagnosis are associated with the limited sensitivity of currently used serological markers.Objective: to evaluate the diagnostic informative value of autoantibodies against different post-translationally modified (PTM vimentin peptides in patients with RA and other ARDs.Patients and methods. The frequency of autoantibodies against different isoforms of vimentin was estimated in 144 patients with RA, in 36 patients with other ARDs (ankylosing spondylitis and scleroderma systematica, and in 25 patients of a control group, who had no rheumatic diseases. Antibodies against different PTM vimentin peptides obtained using citrullination, carbamylation/homocitrullination, and acetylation were determined. Anti-citrullinated vimentin (anti-CitVim peptide, anti-carbamylated vimentin (anti-CarVim peptide, and anti-acetylated vimentin (anti-AcVim peptide autoantibodies of IgG and IgA classes were estimated in the serum by enzyme immunoassay.Results. The results of the study showed that IgG and IgA anti-CitVim had the maximum area under the ROC curve (AUC (0.859 and 0.855, respectively. A slightly smaller AUC was seen in IgG anti-CarVim (0.85, IgG anti-AcVim (0.784, and IgA anti-AcVim (0.651. The diagnostic sensitivity and diagnostic specificity were 66.2 and 96.77% for IgG anti-CitVim, 60.56 and 91.94% for IgA anti-CitVim, 91.55 and 53.23% for IgG anti-CarVim, 63.38 and 93.55% for IgG anti-AcVim, and 49.3 and 70.97%, IgA anti-AcVim, respectively. Positivity for IgG anti-CitVim, IgG anti-CarVim, and IgG anti-AcVim, and anti-IgA CitVim was significantly more frequently detected in patients with RA than in those with other ARDs and in the control group (p<0.05. Thus, the identified autoantibodies against modified

  14. Copper circuit patterning on polymer using selective surface modification and electroless plating

    Energy Technology Data Exchange (ETDEWEB)

    Park, Sang Jin [Department of Materials Science and Engineering, Chungnam National University, Daejeon 305-764 (Korea, Republic of); Ko, Tae-Jun [Institute for Multidisciplinary Convergence of Materials, Korea Institute of Science and Technology, Seoul 130-650 (Korea, Republic of); Department of Materials Science and Engineering, Seoul National University, Seoul 151-742 (Korea, Republic of); Yoon, Juil [Department of Mechanical Systems Engineering, Hansung University, Seoul 136-792 (Korea, Republic of); Moon, Myoung-Woon [Institute for Multidisciplinary Convergence of Materials, Korea Institute of Science and Technology, Seoul 130-650 (Korea, Republic of); Oh, Kyu Hwan [Department of Materials Science and Engineering, Seoul National University, Seoul 151-742 (Korea, Republic of); Han, Jun Hyun, E-mail: jhhan@cnu.ac.kr [Department of Materials Science and Engineering, Chungnam National University, Daejeon 305-764 (Korea, Republic of)

    2017-02-28

    Highlights: • A new simple two step method for the pattering of Cu circuits on PET substrate was proposed. • The simple patterning of the high adhesive Cu circuits was achieved by plasma treatment using a patterned mask coated with a catalyst material. • The high adhesive strength of Cu circuits was due to the nanostructure formed by oxygen plasma treatment. - Abstract: We have examined a potential new and simple method for patterning a copper circuit on PET substrate by copper electroless plating, without the pretreatment steps (i.e., sensitization and activation) for electroless plating as well as the etching processes of conventional circuit patterning. A patterned mask coated with a catalyst material, Ag, for the reduction of Cu ions, is placed on a PET substrate. Subsequent oxygen plasma treatment of the PET substrate covered with the mask promotes the selective generation of anisotropic pillar- or hair-like nanostructures coated with co-deposited nanoparticles of the catalyst material on PET. After oxygen plasma treatment, a Cu circuit is well formed just by dipping the plasma-treated PET into a Cu electroless plating solution. By increasing the oxygen gas pressure in the chamber, the height of the nanostructures increases and the Ag catalyst particles are coated on not only the top but also the side surfaces of the nanostructures. Strong mechanical interlocking between the Cu circuit and PET substrate is produced by the large surface area of the nanostructures, and enhances peel strength. Results indicate this new simple two step (plasma surface modification and pretreatment-free electroless plating) method can be used to produce a flexible Cu circuit with good adhesion.

  15. Chemical mechanisms of histone lysine and arginine modifications

    OpenAIRE

    Smith, Brian C.; Denu, John M.

    2008-01-01

    Histone lysine and arginine residues are subject to a wide array of post-translational modifications including methylation, citrullination, acetylation, ubiquitination, and sumoylation. The combinatorial action of these modifications regulates critical DNA processes including replication, repair, and transcription. In addition, enzymes that modify histone lysine and arginine residues have been correlated with a variety of human diseases including arthritis, cancer, heart disease, diabetes, an...

  16. Dietary fat and fiber interact to uniquely modify global histone post-translational epigenetic programming in a rat colon cancer progression model.

    Science.gov (United States)

    Triff, Karen; McLean, Mathew W; Callaway, Evelyn; Goldsby, Jennifer; Ivanov, Ivan; Chapkin, Robert S

    2018-04-16

    Dietary fermentable fiber generates short-chain fatty acids (SCFA), e.g., butyrate, in the colonic lumen which serves as a chemoprotective histone deacetylase inhibitor and/or as an acetylation substrate for histone acetylases. In addition, n-3 polyunsaturated fatty acids (n-3 PUFA) in fish oil can affect the chromatin landscape by acting as ligands for tumor suppressive nuclear receptors. In an effort to gain insight into the global dimension of post-translational modification of histones (including H3K4me3 and H3K9ac) and clarify the chemoprotective impact of dietary bioactive compounds on transcriptional control in a preclinical model of colon cancer, we generated high-resolution genome-wide RNA (RNA-Seq) and "chromatin-state" (H3K4me3-seq and H3K9ac-seq) maps for intestinal (epithelial colonocytes) crypts in rats treated with a colon carcinogen and fed diets containing bioactive (i) fish oil, (ii) fermentable fiber (a rich source of SCFA), (iii) a combination of fish oil plus pectin or (iv) control, devoid of fish oil or pectin. In general, poor correlation was observed between differentially transcribed (DE) and enriched genes (DERs) at multiple epigenetic levels. The combinatorial diet (fish oil + pectin) uniquely affected transcriptional profiles in the intestinal epithelium, e.g., upregulating lipid catabolism and beta-oxidation associated genes. These genes were linked to activated ligand-dependent nuclear receptors associated with n-3 PUFA and were also correlated with the mitochondrial L-carnitine shuttle and the inhibition of lipogenesis. These findings demonstrate that the chemoprotective fish oil + pectin combination diet uniquely induces global histone state modifications linked to the expression of chemoprotective genes. This article is protected by copyright. All rights reserved. © 2018 UICC.

  17. Modification of hemiplegic compensatory gait pattern by symmetry-based motion controller of HAL.

    Science.gov (United States)

    Kawamoto, Hiroaki; Kadone, Hideki; Sakurai, Takeru; Sankai, Yoshiyuki

    2015-01-01

    As one of several characteristics of hemiplegic patients after stroke, compensatory gait caused by affected limb is often seen. The purpose of this research is to apply a symmetry-based controller of a wearable type lower limb robot, Hybrid Assistive Limb (HAL) to hemiplegic patients with compensatory gait, and to investigate improvement of gait symmetry. The controller is designed respectively for swing phase and support phase according to characteristics of hemiplegic gait pattern. The controller during swing phase stores the motion of the unaffected limb and then provides motion support on the affected limb during the subsequent swing using the stored pattern to realize symmetric gait based on spontaneous limb swing. Moreover, the controller during support phase provides motion to extend hip and knee joints to support wearer's body. Clinical tests were conducted in order to assess the modification of gait symmetry. Our case study involved participation of one chronic stroke patient who performs abnormally-compensatory gait for both of the affected and unaffected limbs. As a result, the patient's gait symmetry was improved by providing motion support during the swing phase on the affected side and motion constraint during the support phase on the unaffected side. The study showed promising basis for the effectiveness of the controller for the future clinical study.

  18. The interplay of multiple feedback loops with post-translational kinetics results in bistability of mycobacterial stress response

    International Nuclear Information System (INIS)

    Tiwari, Abhinav; Igoshin, Oleg A; Balázsi, Gábor; Gennaro, Maria Laura

    2010-01-01

    Bacterial persistence is the phenomenon in which a genetically identical fraction of a bacterial population can survive exposure to stress by reduction or cessation of growth. Persistence in mycobacteria has been recently linked to a stress-response network, consisting of the MprA/MprB two-component system and alternative sigma factor σ E . This network contains multiple positive transcriptional feedback loops which may give rise to bistability, making it a good candidate for controlling the mycobacterial persistence switch. To analyze the possibility of bistability, we develop a method that involves decoupling of the network into transcriptional and post-translational interaction modules. As a result we reduce the dimensionality of the dynamical system and independently analyze input–output relations in the two modules to formulate a necessary condition for bistability in terms of their logarithmic gains. We show that neither the positive autoregulation in the MprA/MprB network nor the σ E -mediated transcriptional feedback is sufficient to induce bistability in a biochemically realistic parameter range. Nonetheless, inclusion of the post-translational regulation of σ E by RseA increases the effective cooperativity of the system, resulting in bistability that is robust to parameter variation. We predict that overexpression or deletion of RseA, the key element controlling the ultrasensitive response, can eliminate bistability

  19. Post-translational regulation and trafficking of the granulin-containing protease RD21 of Arabidopsis thaliana.

    Directory of Open Access Journals (Sweden)

    Christian Gu

    Full Text Available RD21-like proteases are ubiquitous, plant-specific papain-like proteases typified by carrying a C-terminal granulin domain. RD21-like proteases are involved in immunity and associated with senescence and various types of biotic and abiotic stresses. Here, we interrogated Arabidopsis RD21 regulation and trafficking by site-directed mutagenesis, agroinfiltration, western blotting, protease activity profiling and protein degradation. Using an introduced N-glycan sensor, deglycosylation experiments and glyco-engineered N. benthamiana plants, we show that RD21 passes through the Golgi where it becomes fucosylated. Our studies demonstrate that RD21 is regulated at three post-translational levels. Prodomain removal is not blocked in the catalytic Cys mutant, indicating that RD21 is activated by a proteolytic cascade. However, RD21 activation in Arabidopsis does not require vacuolar processing enzymes (VPEs or aleurain-like protease AALP. In contrast, granulin domain removal requires the catalytic Cys and His residues and is therefore autocatalytic. Furthermore, SDS can (re-activate latent RD21 in Arabidopsis leaf extracts, indicating the existence of a third layer of post-translational regulation, possibly mediated by endogenous inhibitors. RD21 causes a dominant protease activity in Arabidopsis leaf extracts, responsible for SDS-induced proteome degradation.

  20. Tungstate-induced color-pattern modifications of butterfly wings are independent of stress response and ecdysteroid effect.

    Science.gov (United States)

    Otaki, Joji M; Ogasawara, Tsuyoshi; Yamamoto, Haruhiko

    2005-06-01

    Systemic injections of sodium tungstate, a protein-tyrosine phosphatase (PTPase) inhibitor, to pupae immediately after pupation have been shown to efficiently produce characteristic color-pattern modifications on the wings of many species of butterflies. Here we demonstrated that the tungstate-induced modification pattern was entirely different from other chemically-induced ones in a species of nymphalid butterfly Junonia (Precis) orithya. In this species, the systemic injections of tungstate produced characteristic expansion of black area and shrinkage of white area together with the move of parafocal elements toward the wing base. Overall, pattern boundaries became obscure. In contrast, an entirely different modification pattern, overall darkening of wings, was observed by the injections of stress-inducing chemicals, thapsigargin, ionomycin, or geldanamycin, to pupae under the rearing conditions for the adult summer form. On the ventral wings, this darkening was due to an increase of the proportion of peppered dark scales, which was reminiscent of the natural fall form of this species. Under the same rearing conditions, the injections of ecdysteroid, which is a well-known hormone being responsible for the seasonal polyphenism of nymphalid butterflies, yielded overall expansion of orange area especially around eyespots. Taken together, we conclude that the tungstate-induced modifications are clearly distinguishable from those of stress response and ecdysteroid effect. This conclusion then suggests that the putative PTPase signaling pathway that is sensitive to tungstate uniquely contributes to the wing-wide color-pattern development in butterflies.

  1. Posttranslational modifications in human plasma MBL and human recombinant MBL

    DEFF Research Database (Denmark)

    Jensen, Pia Hønnerup; Laursen, Inga; Matthiesen, Finn

    2007-01-01

    the intact protein in its active conformation. For the first time, positions and occupation frequency of partial hydroxylations and partial glycosylations are reported in MBL. Hydroxylation and glycosylation patterns of both recombinant and plasma derived MBL were determined, using a combination of mass......Mannan-binding lectin (MBL) is a complex serum protein that plays an important role in innate immunity. In addition to assuming several different oligomeric forms, the polypeptide itself is highly heterogeneous. This heterogeneity is due to post-translational modifications, which help to stabilize......(202)) was modified in trace amounts to dehydroalanine, as detected by a 34 Da mass loss. This impairs the formation of a disulphide bond in the carbohydrate recognition domain. The dehydroalanine was identified in similar small amounts in both recombinant and plasma-derived MBL....

  2. Histones and their modifications in ovarian cancer - drivers of disease and therapeutic targets.

    Science.gov (United States)

    Marsh, Deborah J; Shah, Jaynish S; Cole, Alexander J

    2014-01-01

    Epithelial ovarian cancer has the highest mortality of the gynecological malignancies. High grade serous epithelial ovarian cancer (SEOC) is the most common subtype, with the majority of women presenting with advanced disease where 5-year survival is around 25%. Platinum-based chemotherapy in combination with paclitaxel remains the most effective treatment despite platinum therapies being introduced almost 40 years ago. Advances in molecular medicine are underpinning new strategies for the treatment of cancer. Major advances have been made by international initiatives to sequence cancer genomes. For SEOC, with the exception of TP53 that is mutated in virtually 100% of these tumors, there is no other gene mutated at high frequency. There is extensive copy number variation, as well as changes in methylation patterns that will influence gene expression. To date, the role of histones and their post-translational modifications in ovarian cancer is a relatively understudied field. Post-translational histone modifications play major roles in gene expression as they direct the configuration of chromatin and so access by transcription factors. Histone modifications include methylation, acetylation, and monoubiquitination, with involvement of enzymes including histone methyltransferases, histone acetyltransferases/deacetylases, and ubiquitin ligases/deubiquitinases, respectively. Complexes such as the Polycomb repressive complex also play roles in the control of histone modifications and more recently roles for long non-coding RNA and microRNAs are emerging. Epigenomic-based therapies targeting histone modifications are being developed and offer new approaches for the treatment of ovarian cancer. Here, we discuss histone modifications and their aberrant regulation in malignancy and specifically in ovarian cancer. We review current and upcoming histone-based therapies that have the potential to inform and improve treatment strategies for women with ovarian cancer.

  3. Histones and their modifications in ovarian cancer – drivers of disease and therapeutic targets

    Directory of Open Access Journals (Sweden)

    Deborah Joy Marsh

    2014-06-01

    Full Text Available Epithelial ovarian cancer has the highest mortality of the gynecological malignancies. High grade serous epithelial ovarian cancer (SEOC is the most common subtype, with the majority of women presenting with advanced disease where 5 year survival is around 25%. Platinum-based chemotherapy in combination with paclitaxel remains the most effective treatment despite platinum therapies being introduced almost 40 years ago. Advances in molecular medicine are underpinning new strategies for the treatment of cancer. Major advances have been made by international initiatives to sequence cancer genomes. For SEOC, with the exception of TP53 that is mutated in virtually 100% of these tumors, there is no other gene mutated at high frequency. There is extensive copy number variation, as well as changes in methylation patterns that will influence gene expression. To date, the role of histones and their post-translational modifications in ovarian cancer is a relatively understudied field. Post-translational histone modifications play major roles in gene expression as they direct the configuration of chromatin and so access by transcription factors. Histone modifications include methylation, acetylation and monoubiquitination, with involvement of enzymes including histone methyl transferases (HMTases, histone acetyltransferases/deacetylases and ubiquitin ligases/deubiquitinases respectively. Complexes such as the Polycomb Repressive Complex also play roles in the control of histone modifications and more recently roles for long non-coding (lnc RNA and microRNAs (miRNAs are emerging. Epigenomic-based therapies targeting histone modifications are being developed and offer new approaches for the treatment of ovarian cancer. Here we discuss histone modifications and their aberrant regulation in malignancy and specifically in ovarian cancer. We review current and upcoming histone-based therapies that have the potential to inform and improve treatment strategies for

  4. Post-translational control of RIPK3 and MLKL mediated necroptotic cell death [version 1; referees: 4 approved

    Directory of Open Access Journals (Sweden)

    James M. Murphy

    2015-11-01

    Full Text Available Several programmed lytic and necrotic-like cell death mechanisms have now been uncovered, including the recently described receptor interacting protein kinase-3 (RIPK3-mixed lineage kinase domain-like (MLKL-dependent necroptosis pathway. Genetic experiments have shown that programmed necrosis, including necroptosis, can play a pivotal role in regulating host-resistance against microbial infections. Alternatively, excess or unwarranted necroptosis may be pathological in autoimmune and autoinflammatory diseases. This review highlights the recent advances in our understanding of the post-translational control of RIPK3-MLKL necroptotic signaling. We discuss the critical function of phosphorylation in the execution of necroptosis, and highlight the emerging regulatory roles for several ubiquitin ligases and deubiquitinating enzymes. Finally, based on current evidence, we discuss the potential mechanisms by which the essential, and possibly terminal, necroptotic effector, MLKL, triggers the disruption of cellular membranes to cause cell lysis.

  5. Stage-specific histone modification profiles reveal global transitions in the Xenopus embryonic epigenome.

    Directory of Open Access Journals (Sweden)

    Tobias D Schneider

    Full Text Available Vertebrate embryos are derived from a transitory pool of pluripotent cells. By the process of embryonic induction, these precursor cells are assigned to specific fates and differentiation programs. Histone post-translational modifications are thought to play a key role in the establishment and maintenance of stable gene expression patterns underlying these processes. While on gene level histone modifications are known to change during differentiation, very little is known about the quantitative fluctuations in bulk histone modifications during development. To investigate this issue we analysed histones isolated from four different developmental stages of Xenopus laevis by mass spectrometry. In toto, we quantified 59 modification states on core histones H3 and H4 from blastula to tadpole stages. During this developmental period, we observed in general an increase in the unmodified states, and a shift from histone modifications associated with transcriptional activity to transcriptionally repressive histone marks. We also compared these naturally occurring patterns with the histone modifications of murine ES cells, detecting large differences in the methylation patterns of histone H3 lysines 27 and 36 between pluripotent ES cells and pluripotent cells from Xenopus blastulae. By combining all detected modification transitions we could cluster their patterns according to their embryonic origin, defining specific histone modification profiles (HMPs for each developmental stage. To our knowledge, this data set represents the first compendium of covalent histone modifications and their quantitative flux during normogenesis in a vertebrate model organism. The HMPs indicate a stepwise maturation of the embryonic epigenome, which may be causal to the progressing restriction of cellular potency during development.

  6. Modification of Patterned Nanoporous Gold Thin Film Electrodes via Electro-annealing and Electrochemical Etching

    Science.gov (United States)

    Dorofeeva, Tatiana

    Nanostructured materials have had a major impact on various fields, including medicine, catalysis, and energy storage, for the major part due to unique phenomena that arise at nanoscale. For this reason, there is a sustained need for new nanostructured materials, techniques to pattern them, and methods to precisely control their nanostructure. To that end, the primary focus of this dissertation is to demonstrate novel techniques to fabricate and tailor the morphology of a class of nanoporous metals, obtained by a process known as dealloying. In this process, while the less noble constituent of an alloy is chemically dissolved, surface-diffusion of the more noble constituent leads to self-assembly of a bicontinuous ligament network with characteristic porosity of ˜70% and ligament diameter of 10s of nanometers. As a model material produced by dealloying, this work employ nanoporous gold (np-Au), which has attracted significant attention of desirable features, such as high effective surface area, electrical conductivity, well-defined thiol-based surface modification strategies, microfabrication-compatibility, and biocompatibility. The most commonly method used to modify the morphology of np-Au is thermal treatment, where the enhanced diffusivity of the surface atoms leads to ligament (and consequently pore) coarsening. This method, however, is not conducive to modifying the morphology of thin films at specific locations on the film, which is necessary for creating devices that may need to contain different morphologies on a single device. In addition, coarsening attained by thermal treatment also leads to an undesirable reduction in effective surface area. In response to these challenges, this work demonstrates two different techniques that enables in situ modification of np-Au thin film electrodes obtained by sputter-deposition of a precursors silver-rich gold-silver alloy. The first method, referred to as electro-annealing, is achieved by injecting electrical

  7. Albumin modification and fragmentation in renal disease.

    Science.gov (United States)

    Donadio, Carlo; Tognotti, Danika; Donadio, Elena

    2012-02-18

    Albumin is the most important antioxidant substance in plasma and performs many physiological functions. Furthermore, albumin is the major carrier of endogenous molecules and exogenous ligands. This paper reviews the importance of post-translational modifications of albumin and fragments thereof in patients with renal disease. First, current views and controversies on renal handling of proteins, mainly albumin, will be discussed. Post-translational modifications, namely the fragmentation of albumin found with proteomic techniques in nephrotic patients, diabetics, and ESRD patients will be presented and discussed. It is reasonable to hypothesize that proteolytic fragmentation of serum albumin is due to a higher susceptibility to proteases, induced by oxidative stress. The clinical relevance of the fragmentation of albumin has not yet been established. These modifications could affect some physiological functions of albumin and have a patho-physiological role in uremic syndrome. Proteomic analysis of serum allows the identification of over-expressed proteins and can detect post-translational modifications of serum proteins, hitherto hidden, using standard laboratory techniques. Copyright © 2011 Elsevier B.V. All rights reserved.

  8. Nanolithography and nanochemistry: probe-related patterning techniques and chemical modification for nanometer-sized devices

    NARCIS (Netherlands)

    Wouters, D.; Schubert, U.S.

    2004-01-01

    The size regime for devices produced by photolithographic techniques is limited. Therefore, other patterning techniques have been intensively studied to create smaller structures. Scanning-probe-based patterning techniques, such as dip-pen lithography, local force-induced patterning, and local-probe

  9. Programming Post-Translational Control over the Metabolic Labeling of Cellular Proteins with a Noncanonical Amino Acid.

    Science.gov (United States)

    Thomas, Emily E; Pandey, Naresh; Knudsen, Sarah; Ball, Zachary T; Silberg, Jonathan J

    2017-08-18

    Transcriptional control can be used to program cells to label proteins with noncanonical amino acids by regulating the expression of orthogonal aminoacyl tRNA synthetases (aaRSs). However, we cannot yet program cells to control labeling in response to aaRS and ligand binding. To identify aaRSs whose activities can be regulated by interactions with ligands, we used a combinatorial approach to discover fragmented variants of Escherichia coli methionyl tRNA synthetase (MetRS) that require fusion to associating proteins for maximal activity. We found that these split proteins could be leveraged to create ligand-dependent MetRS using two approaches. When a pair of MetRS fragments was fused to FKBP12 and the FKBP-rapamycin binding domain (FRB) of mTOR and mutations were introduced that direct substrate specificity toward azidonorleucine (Anl), Anl metabolic labeling was significantly enhanced in growth medium containing rapamycin, which stabilizes the FKBP12-FRB complex. In addition, fusion of MetRS fragments to the termini of the ligand-binding domain of the estrogen receptor yielded proteins whose Anl metabolic labeling was significantly enhanced when 4-hydroxytamoxifen (4-HT) was added to the growth medium. These findings suggest that split MetRS can be fused to a range of ligand-binding proteins to create aaRSs whose metabolic labeling activities depend upon post-translational interactions with ligands.

  10. GPS-Lipid: a robust tool for the prediction of multiple lipid modification sites

    OpenAIRE

    Xie, Yubin; Zheng, Yueyuan; Li, Hongyu; Luo, Xiaotong; He, Zhihao; Cao, Shuo; Shi, Yi; Zhao, Qi; Xue, Yu; Zuo, Zhixiang; Ren, Jian

    2016-01-01

    As one of the most common post-translational modifications in eukaryotic cells, lipid modification is an important mechanism for the regulation of variety aspects of protein function. Over the last decades, three classes of lipid modifications have been increasingly studied. The co-regulation of these different lipid modifications is beginning to be noticed. However, due to the lack of integrated bioinformatics resources, the studies of co-regulatory mechanisms are still very limited. In this...

  11. USP2-45 Is a Circadian Clock Output Effector Regulating Calcium Absorption at the Post-Translational Level.

    Directory of Open Access Journals (Sweden)

    Daniel Pouly

    Full Text Available The mammalian circadian clock influences most aspects of physiology and behavior through the transcriptional control of a wide variety of genes, mostly in a tissue-specific manner. About 20 clock-controlled genes (CCGs oscillate in virtually all mammalian tissues and are generally considered as core clock components. One of them is Ubiquitin-Specific Protease 2 (Usp2, whose status remains controversial, as it may be a cogwheel regulating the stability or activity of core cogwheels or an output effector. We report here that Usp2 is a clock output effector related to bodily Ca2+ homeostasis, a feature that is conserved across evolution. Drosophila with a whole-body knockdown of the orthologue of Usp2, CG14619 (dUsp2-kd, predominantly die during pupation but are rescued by dietary Ca2+ supplementation. Usp2-KO mice show hyperabsorption of dietary Ca2+ in small intestine, likely due to strong overexpression of the membrane scaffold protein NHERF4, a regulator of the Ca2+ channel TRPV6 mediating dietary Ca2+ uptake. In this tissue, USP2-45 is found in membrane fractions and negatively regulates NHERF4 protein abundance in a rhythmic manner at the protein level. In clock mutant animals (Cry1/Cry2-dKO, rhythmic USP2-45 expression is lost, as well as the one of NHERF4, confirming the inverse relationship between USP2-45 and NHERF4 protein levels. Finally, USP2-45 interacts in vitro with NHERF4 and endogenous Clathrin Heavy Chain. Taken together these data prompt us to define USP2-45 as the first clock output effector acting at the post-translational level at cell membranes and possibly regulating membrane permeability of Ca2+.

  12. Biochemical systems approaches for the analysis of histone modification readout.

    Science.gov (United States)

    Soldi, Monica; Bremang, Michael; Bonaldi, Tiziana

    2014-08-01

    Chromatin is the macromolecular nucleoprotein complex that governs the organization of genetic material in the nucleus of eukaryotic cells. In chromatin, DNA is packed with histone proteins into nucleosomes. Core histones are prototypes of hyper-modified proteins, being decorated by a large number of site-specific reversible and irreversible post-translational modifications (PTMs), which contribute to the maintenance and modulation of chromatin plasticity, gene activation, and a variety of other biological processes and disease states. The observations of the variety, frequency and co-occurrence of histone modifications in distinct patterns at specific genomic loci have led to the idea that hPTMs can create a molecular barcode, read by effector proteins that translate it into a specific transcriptional state, or process, on the underlying DNA. However, despite the fact that this histone-code hypothesis was proposed more than 10 years ago, the molecular details of its working mechanisms are only partially characterized. In particular, two questions deserve specific investigation: how the different modifications associate and synergize into patterns and how these PTM configurations are read and translated by multi-protein complexes into a specific functional outcome on the genome. Mass spectrometry (MS) has emerged as a versatile tool to investigate chromatin biology, useful for both identifying and validating hPTMs, and to dissect the molecular determinants of histone modification readout systems. We review here the MS techniques and the proteomics methods that have been developed to address these fundamental questions in epigenetics research, emphasizing approaches based on the proteomic dissection of distinct native chromatin regions, with a critical evaluation of their present challenges and future potential. This article is part of a Special Issue entitled: Molecular mechanisms of histone modification function. Copyright © 2014 Elsevier B.V. All rights reserved.

  13. Femtosecond laser patterning, synthesis, defect formation, and structural modification of atomic layered materials

    International Nuclear Information System (INIS)

    Yoo, Jae-Hyuck; Kim, Eunpa; Hwang, David J.

    2016-01-01

    This article summarizes recent research on laser-based processing of twodimensional (2D) atomic layered materials, including graphene and transition metal dichalcogenides (TMDCs). Ultrafast lasers offer unique processing routes that take advantage of distinct interaction mechanisms with 2D materials to enable extremely localized energy deposition. Experiments have shown that ablative direct patterning of graphene by ultrafast lasers can achieve resolutions of tens of nanometers, as well as single-step pattern transfer. Ultrafast lasers also induce non-thermal excitation mechanisms that are useful for the thinning of TMDCs to tune the 2D material bandgap. Laser-assisted site-specific doping was recently demonstrated where ultrafast laser radiation under ambient air environment could be used for the direct writing of high-quality graphene patterns on insulating substrates. This article concludes with an outlook towards developing further advanced laser processing with scalability, in situ monitoring strategies and potential applications.

  14. Low-fat dietary pattern and risk of colorectal cancer: the Women's Health Initiative Randomized Controlled Dietary Modification Trial.

    Science.gov (United States)

    Beresford, Shirley A A; Johnson, Karen C; Ritenbaugh, Cheryl; Lasser, Norman L; Snetselaar, Linda G; Black, Henry R; Anderson, Garnet L; Assaf, Annlouise R; Bassford, Tamsen; Bowen, Deborah; Brunner, Robert L; Brzyski, Robert G; Caan, Bette; Chlebowski, Rowan T; Gass, Margery; Harrigan, Rosanne C; Hays, Jennifer; Heber, David; Heiss, Gerardo; Hendrix, Susan L; Howard, Barbara V; Hsia, Judith; Hubbell, F Allan; Jackson, Rebecca D; Kotchen, Jane Morley; Kuller, Lewis H; LaCroix, Andrea Z; Lane, Dorothy S; Langer, Robert D; Lewis, Cora E; Manson, JoAnn E; Margolis, Karen L; Mossavar-Rahmani, Yasmin; Ockene, Judith K; Parker, Linda M; Perri, Michael G; Phillips, Lawrence; Prentice, Ross L; Robbins, John; Rossouw, Jacques E; Sarto, Gloria E; Stefanick, Marcia L; Van Horn, Linda; Vitolins, Mara Z; Wactawski-Wende, Jean; Wallace, Robert B; Whitlock, Evelyn

    2006-02-08

    Observational studies and polyp recurrence trials are not conclusive regarding the effects of a low-fat dietary pattern on risk of colorectal cancer, necessitating a primary prevention trial. To evaluate the effects of a low-fat eating pattern on risk of colorectal cancer in postmenopausal women. The Women's Health Initiative Dietary Modification Trial, a randomized controlled trial conducted in 48,835 postmenopausal women aged 50 to 79 years recruited between 1993 and 1998 from 40 clinical centers throughout the United States. Participants were randomly assigned to the dietary modification intervention (n = 19,541; 40%) or the comparison group (n = 29,294; 60%). The intensive behavioral modification program aimed to motivate and support reductions in dietary fat, to increase consumption of vegetables and fruits, and to increase grain servings by using group sessions, self-monitoring techniques, and other tailored and targeted strategies. Women in the comparison group continued their usual eating pattern. Invasive colorectal cancer incidence. A total of 480 incident cases of invasive colorectal cancer occurred during a mean follow-up of 8.1 (SD, 1.7) years. Intervention group participants significantly reduced their percentage of energy from fat by 10.7% more than did the comparison group at 1 year, and this difference between groups was mostly maintained (8.1% at year 6). Statistically significant increases in vegetable, fruit, and grain servings were also made. Despite these dietary changes, there was no evidence that the intervention reduced the risk of invasive colorectal cancer during the follow-up period. There were 201 women with invasive colorectal cancer (0.13% per year) in the intervention group and 279 (0.12% per year) in the comparison group (hazard ratio, 1.08; 95% confidence interval, 0.90-1.29). Secondary analyses suggested potential interactions with baseline aspirin use and combined estrogen-progestin use status (P = .01 for each). Colorectal

  15. EUV multilayer defect compensation (MDC) by absorber pattern modification: from theory to wafer validation

    Science.gov (United States)

    Pang, Linyong; Hu, Peter; Satake, Masaki; Tolani, Vikram; Peng, Danping; Li, Ying; Chen, Dongxue

    2011-11-01

    According to the ITRS roadmap, mask defects are among the top technical challenges to introduce extreme ultraviolet (EUV) lithography into production. Making a multilayer defect-free extreme ultraviolet (EUV) blank is not possible today, and is unlikely to happen in the next few years. This means that EUV must work with multilayer defects present on the mask. The method proposed by Luminescent is to compensate effects of multilayer defects on images by modifying the absorber patterns. The effect of a multilayer defect is to distort the images of adjacent absorber patterns. Although the defect cannot be repaired, the images may be restored to their desired targets by changing the absorber patterns. This method was first introduced in our paper at BACUS 2010, which described a simple pixel-based compensation algorithm using a fast multilayer model. The fast model made it possible to complete the compensation calculations in seconds, instead of days or weeks required for rigorous Finite Domain Time Difference (FDTD) simulations. Our SPIE 2011 paper introduced an advanced compensation algorithm using the Level Set Method for 2D absorber patterns. In this paper the method is extended to consider process window, and allow repair tool constraints, such as permitting etching but not deposition. The multilayer defect growth model is also enhanced so that the multilayer defect can be "inverted", or recovered from the top layer profile using a calibrated model.

  16. A Canonical DREB2-Type Transcription Factor in Lily Is Post-translationally Regulated and Mediates Heat Stress Response

    Directory of Open Access Journals (Sweden)

    Ze Wu

    2018-03-01

    Full Text Available Based on studies of monocot crops and eudicot model plants, the DREB2 class of AP2-type transcription factor has been shown to play crucial roles in various abiotic stresses, especially in the upstream of the heat stress response; however, research on DREB2s has not been reported in non-gramineous monocot plants. Here, we identified a novel DREB2 (LlDREB2B from lily (Lilium longiflorum, which was homologous to AtDREB2A of Arabidopsis, OsDREB2B of rice, and ZmDREB2A of maize. LlDREB2B was induced by heat, cold, salt, and mannitol stress, and its protein had transcriptional activity, was located in the nucleus, was able to bind to the dehydration-responsive element (DRE, and participated in the heat-responsive pathway of HsfA3. Overexpression of LlDREB2B in Arabidopsis activated expression of downstream genes and improved thermotolerance. LlDREB2B was not regulated by alternative splicing; functional transcripts accumulated under either normal or heat-stress conditions. A potential PEST sequence was predicted in LlDREB2B, but the stability of the LlDREB2B protein was not positively affected when the predicated PEST sequence was deleted. Further analysis revealed that the predicated PEST sequence lacked a SBC or SBC-like motif allowing interaction with BPMs and required for negative regulation. Nevertheless, LlDREB2B was still regulated at the post-translational level by interaction with AtDRIP1 and AtDRIP2 of Arabidopsis. In addition, LlDREB2B also interacted with AtRCD1 and LlRCD1 via a potential RIM motif located at amino acids 215–245. Taken together, our results show that LlDREB2B participated in the establishment of thermotolerance, and its regulation was different from that of the orthologs of gramineous and eudicot plants.

  17. A Canonical DREB2-Type Transcription Factor in Lily Is Post-translationally Regulated and Mediates Heat Stress Response.

    Science.gov (United States)

    Wu, Ze; Liang, Jiahui; Zhang, Shuai; Zhang, Bing; Zhao, Qingcui; Li, Guoqing; Yang, Xi; Wang, Chengpeng; He, Junna; Yi, Mingfang

    2018-01-01

    Based on studies of monocot crops and eudicot model plants, the DREB2 class of AP2-type transcription factor has been shown to play crucial roles in various abiotic stresses, especially in the upstream of the heat stress response; however, research on DREB2s has not been reported in non-gramineous monocot plants. Here, we identified a novel DREB2 (LlDREB2B) from lily ( Lilium longiflorum ), which was homologous to AtDREB2A of Arabidopsis, OsDREB2B of rice, and ZmDREB2A of maize. LlDREB2B was induced by heat, cold, salt, and mannitol stress, and its protein had transcriptional activity, was located in the nucleus, was able to bind to the dehydration-responsive element (DRE), and participated in the heat-responsive pathway of HsfA3. Overexpression of LlDREB2B in Arabidopsis activated expression of downstream genes and improved thermotolerance. LlDREB2B was not regulated by alternative splicing; functional transcripts accumulated under either normal or heat-stress conditions. A potential PEST sequence was predicted in LlDREB2B, but the stability of the LlDREB2B protein was not positively affected when the predicated PEST sequence was deleted. Further analysis revealed that the predicated PEST sequence lacked a SBC or SBC-like motif allowing interaction with BPMs and required for negative regulation. Nevertheless, LlDREB2B was still regulated at the post-translational level by interaction with AtDRIP1 and AtDRIP2 of Arabidopsis. In addition, LlDREB2B also interacted with AtRCD1 and LlRCD1 via a potential RIM motif located at amino acids 215-245. Taken together, our results show that LlDREB2B participated in the establishment of thermotolerance, and its regulation was different from that of the orthologs of gramineous and eudicot plants.

  18. Post-translational inhibition of IP-10 secretion in IEC by probiotic bacteria: impact on chronic inflammation.

    Directory of Open Access Journals (Sweden)

    Gabriele Hoermannsperger

    specific effects of probiotic intervention that correlate with reduced IP-10 protein expression in the native epithelium. Furthermore, we revealed post-translational degradation of IP-10 protein in IEC to be the molecular mechanism underlying the anti-inflammatory effect.

  19. Selective Photophysical Modification on Light-Emitting Polymer Films for Micro- and Nano-Patterning

    Directory of Open Access Journals (Sweden)

    Xinping Zhang

    2016-02-01

    Full Text Available Laser-induced cross-linking in polymeric semiconductors was utilized to achieve micro- and nano-structuring in thin films. Single- and two-photon cross-linking processes led to the reduction in both the refractive index and thickness of the polymer films. The resultant photonic structures combine the features of both relief- and phase-gratings. Selective cross-linking in polymer blend films based on different optical response of different molecular phases enabled “solidification” of the phase-separation scheme, providing a stable template for further photonic structuring. Dielectric and metallic structures are demonstrated for the fabrication methods using cross-linking in polymer films. Selective cross-linking enables direct patterning into polymer films without introducing additional fabrication procedures or additional materials. The diffraction processes of the emission of the patterned polymeric semiconductors may provide enhanced output coupling for light-emitting diodes or distributed feedback for lasers.

  20. Cutting blade dentitions in squaliform sharks form by modification of inherited alternate tooth ordering patterns

    OpenAIRE

    Underwood, Charlie J.; Johanson, Z.; Smith, M.M.

    2016-01-01

    The squaliform sharks represent one of the most speciose shark clades. Many adult squaliforms have blade-like teeth, either on both jaws or restricted to the lower jaw, forming a continuous, serrated blade along the jaw margin. These teeth are replaced as a single unit and successor teeth lack the alternate arrangement present in other elasmobranchs. Micro-CT scans of embryos of squaliforms and a related outgroup (Pristiophoridae) revealed that the squaliform dentition pattern represents a hi...

  1. Cutting blade dentitions in squaliform sharks form by modification of inherited alternate tooth ordering patterns

    Science.gov (United States)

    Underwood, Charlie; Johanson, Zerina; Smith, Moya Meredith

    2016-11-01

    The squaliform sharks represent one of the most speciose shark clades. Many adult squaliforms have blade-like teeth, either on both jaws or restricted to the lower jaw, forming a continuous, serrated blade along the jaw margin. These teeth are replaced as a single unit and successor teeth lack the alternate arrangement present in other elasmobranchs. Micro-CT scans of embryos of squaliforms and a related outgroup (Pristiophoridae) revealed that the squaliform dentition pattern represents a highly modified version of tooth replacement seen in other clades. Teeth of Squalus embryos are arranged in an alternate pattern, with successive tooth rows containing additional teeth added proximally. Asynchronous timing of tooth production along the jaw and tooth loss prior to birth cause teeth to align in oblique sets containing teeth from subsequent rows; these become parallel to the jaw margin during ontogeny, so that adult Squalus has functional tooth rows comprising obliquely stacked teeth of consecutive developmental rows. In more strongly heterodont squaliforms, initial embryonic lower teeth develop into the oblique functional sets seen in adult Squalus, with no requirement to form, and subsequently lose, teeth arranged in an initial alternate pattern.

  2. Modification of parabolic dish antenna pattern using two symmetrically placed circular flat plates

    Science.gov (United States)

    Thorpe, Glen C.

    1987-12-01

    This study aims to formulate a method of predicting the far field pattern of a parabolic dish antenna with two moveable flat plates mounted symmetrically on either side of the feed horn. The approach taken has been to first analyze the radiation pattern of the antenna with the disks at certain heights out from the surface of the dish. To do this the near-field radiation in amplitude and phase was measured over a plane surface in the near-field and the values were then transformed into the far field using a Fast Fourier Transform. Far field pattern values of the antenna were directly measured for each setting of the plates. The results obtained from the Fast Fourier Transform of the near field data were in good agreement with the values obtained by measurement. Finally, an approximate model of the antenna was developed and implemented as a computer program. This model, while relatively unsophisticated, provided some insights into the changes in the near field phase distribution caused by the moveable circular flat plates.

  3. Chromatin proteins and modifications as drug targets

    DEFF Research Database (Denmark)

    Helin, Kristian; Dhanak, Dashyant

    2013-01-01

    A plethora of groundbreaking studies have demonstrated the importance of chromatin-associated proteins and post-translational modifications of histones, proteins and DNA (so-called epigenetic modifications) for transcriptional control and normal development. Disruption of epigenetic control...... is a frequent event in disease, and the first epigenetic-based therapies for cancer treatment have been approved. A generation of new classes of potent and specific inhibitors for several chromatin-associated proteins have shown promise in preclinical trials. Although the biology of epigenetic regulation...

  4. Post-translational protein modifications in type 1 diabetes: a role for the repair enzyme protein-L-isoaspartate (D-aspartate) O-methyltransferase?

    DEFF Research Database (Denmark)

    Wägner, A M; Cloos, P; Bergholdt, R

    2007-01-01

    that recognises and repairs isomerised Asn and Asp residues in proteins. The aim of this study was to assess the role of PIMT in the development of type 1 diabetes. MATERIALS AND METHODS: Immunohistochemical analysis of 59 normal human tissues was performed with a monoclonal PIMT antibody. CGP3466B, which induces...... expression of Pcmt1, was tested on MIN6 and INS1 cells, to assess its effect on Pcmt1 mRNA and PIMT levels (RT-PCR and western blot) and apoptosis. Forty-five diabetes-prone BioBreeding (BB) Ottawa Karlsburg (OK) rats were randomised to receive 0, 14 or 500 microg/kg (denoted as the control, low......-dose and high-dose group, respectively) of CGP3466B from week 5 to week 20. RESULTS: A high level of PIMT protein was detected in beta cells. CGP3466B induced a two- to threefold increase in Pcmt1 mRNA levels and reduced apoptosis by 10% in MIN6 cells. No significant effect was seen on cytokine...

  5. In-Depth Glyco-Peptidomics Approach Reveals Unexpected Diversity of Glycosylated Peptides and Atypical Post-Translational Modifications in Dendroaspis angusticeps Snake Venom.

    Science.gov (United States)

    Degueldre, Michel; Echterbille, Julien; Smargiasso, Nicolas; Damblon, Christian; Gouin, Charlotte; Mourier, Gilles; Gilles, Nicolas; De Pauw, Edwin; Quinton, Loïc

    2017-11-18

    Animal venoms represent a valuable source of bioactive peptides that can be derived into useful pharmacological tools, or even innovative drugs. In this way, the venom of Dendroaspis angusticeps (DA), the Eastern Green Mamba, has been intensively studied during recent years. It mainly contains hundreds of large toxins from 6 to 9 kDa, each displaying several disulfide bridges. These toxins are the main target of venom-based studies due to their valuable activities obtained by selectively targeting membrane receptors, such as ion channels or G-protein coupled receptors. This study aims to demonstrate that the knowledge of venom composition is still limited and that animal venoms contain unexpected diversity and surprises. A previous study has shown that Dendroaspis angusticeps venom contains not only a cocktail of classical toxins, but also small glycosylated peptides. Following this work, a deep exploration of DA glycopeptidome by a dual nano liquid chromatography coupled to electrospray ionization mass spectrometry (nanoLC-ESI-MS) and Matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF-MS) analyses was initiated. This study reveals unsuspected structural diversity of compounds such as 221 glycopeptides, displaying different glycan structures. Sequence alignments underline structural similarities with natriuretic peptides already characterized in Elapidae venoms. Finally, the presence of an S -cysteinylation and hydroxylation of proline on four glycopeptides, never described to date in snake venoms, is also revealed by proteomics and affined by nuclear magnetic resonance (NMR) experiments.

  6. Analysis of the primary structure and post-translational modifications of the Schistosoma mansoni antigen Smp28 by electrospray mass spectrometry

    NARCIS (Netherlands)

    Bouchon, B.; Jaquinod, M.; Klarskov, K.; Trottein, F.; Klein, Michele; Van Dorsselaer, A.; Bischoff, Rainer; Roitsch, C.

    1994-01-01

    The Schistosoma mansoni glutathione-S-transferase with an apparent molecular mass of 28 kDa, Smp28, has a blocked N-terminus which has been elucidated with the aid of the cDNA sequence combined with mass spectrometry and amino acid composition analysis of the N-terminal tryptic peptide. The blocked

  7. α-Defensins Induce a Post-translational Modification of Low Density Lipoprotein (LDL) That Promotes Atherosclerosis at Normal Levels of Plasma Cholesterol.

    Science.gov (United States)

    Abu-Fanne, Rami; Maraga, Emad; Abd-Elrahman, Ihab; Hankin, Aviel; Blum, Galia; Abdeen, Suhair; Hijazi, Nuha; Cines, Douglas B; Higazi, Abd Al-Roof

    2016-02-05

    Approximately one-half of the patients who develop clinical atherosclerosis have normal or only modest elevations in plasma lipids, indicating that additional mechanisms contribute to pathogenesis. In view of increasing evidence that inflammation contributes to atherogenesis, we studied the effect of human neutrophil α-defensins on low density lipoprotein (LDL) trafficking, metabolism, vascular deposition, and atherogenesis using transgenic mice expressing human α-defensins in their polymorphonuclear leukocytes (Def(+/+)). Accelerated Def(+/+) mice developed α-defensin·LDL complexes that accelerate the clearance of LDL from the circulation accompanied by enhanced vascular deposition and retention of LDL, induction of endothelial cathepsins, increased endothelial permeability to LDL, and the development of lipid streaks in the aortic roots when fed a regular diet and at normal plasma levels of LDL. Transplantation of bone marrow from Def(+/+) to WT mice increased LDL clearance, increased vascular permeability, and increased vascular deposition of LDL, whereas transplantation of WT bone marrow to Def(+/+) mice prevented these outcomes. The same outcome was obtained by treating Def(+/+) mice with colchicine to inhibit the release of α-defensins. These studies identify a potential new link between inflammation and the development of atherosclerosis. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  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. Comparative Study of the Life Cycle Dependent Post-Translation Modifications of Protein Synthesis Elongation Factor Tu Present in the Membrane Proteome of Streptomycetes and Mycobacteria

    Czech Academy of Sciences Publication Activity Database

    Holub, Martin; Bezoušková, Silvia; Petráčková, Denisa; Kalachová, Ladislava; Kofroňová, Olga; Benada, Oldřich; Weiser, Jaroslav

    2010-01-01

    Roč. 55, č. 3 (2010), s. 203-210 ISSN 0015-5632 R&D Projects: GA AV ČR IAA600200702; GA AV ČR IAA500200913 Institutional research plan: CEZ:AV0Z50200510 Keywords : ESCHERICHIA-COLI * COELICOLOR A3(2) * OUTER-MEMBRANE Subject RIV: EE - Microbiology, Virology Impact factor: 0.977, year: 2010

  10. In-Depth Glyco-Peptidomics Approach Reveals Unexpected Diversity of Glycosylated Peptides and Atypical Post-Translational Modifications in Dendroaspis angusticeps Snake Venom

    Directory of Open Access Journals (Sweden)

    Michel Degueldre

    2017-11-01

    Full Text Available Animal venoms represent a valuable source of bioactive peptides that can be derived into useful pharmacological tools, or even innovative drugs. In this way, the venom of Dendroaspis angusticeps (DA, the Eastern Green Mamba, has been intensively studied during recent years. It mainly contains hundreds of large toxins from 6 to 9 kDa, each displaying several disulfide bridges. These toxins are the main target of venom-based studies due to their valuable activities obtained by selectively targeting membrane receptors, such as ion channels or G-protein coupled receptors. This study aims to demonstrate that the knowledge of venom composition is still limited and that animal venoms contain unexpected diversity and surprises. A previous study has shown that Dendroaspis angusticeps venom contains not only a cocktail of classical toxins, but also small glycosylated peptides. Following this work, a deep exploration of DA glycopeptidome by a dual nano liquid chromatography coupled to electrospray ionization mass spectrometry (nanoLC-ESI-MS and Matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF-MS analyses was initiated. This study reveals unsuspected structural diversity of compounds such as 221 glycopeptides, displaying different glycan structures. Sequence alignments underline structural similarities with natriuretic peptides already characterized in Elapidae venoms. Finally, the presence of an S-cysteinylation and hydroxylation of proline on four glycopeptides, never described to date in snake venoms, is also revealed by proteomics and affined by nuclear magnetic resonance (NMR experiments.

  11. [Modification of the pattern of fatty acids of erythrocytes’ membranes due to the acetone intoxication].

    Science.gov (United States)

    Momot, T V; Kushnerova, N F; Rakhmanin, Yu A

    Results of the study of the impact of acetone intoxication on the fatty acids pattern of the general lipids of erythrocytes’ membranes in rats are presented. The inhalation exposure of acetone was carried out in the inoculation chamber with the volume of 100 liters. The chamber was designed for the type of B.A. Kurlyandsky with self-contained system of purification and air regeneration and specified parameters of temperature (20-22С) and air humidity. The flow rate of the air and aerosolized acetone passed through the chamber accounted of 10 liters/min. Concentration of acetone in the chamber was sustained at the level of 206 ± 3,9 mg/m that corresponds to maximum permissible concentration for acetone vapor in the air of a working area. The time of exposure was 6 hours per day for 3 weeks in a monotonous mode, excluding weekend, and was based upon specific parameters of environment simulation in industry. The acetone impact was shown to be accompanied by the gain in the quantity of all kinds of saturated fatty acids and the fall of unsaturated fatty acids in general lipids of erythrocytes ’ membranes in rats and in the structure ofphospholipid fractions. In the content of phosphatydilcholine and phosphatydilethanolamine, as a basic structural phospholipids of biological membranes, there was noted the increase in palmitic and stearic acids. In the range offatty acids of the n-6 family the amount of linoleic and arachidonic acids decreased. In the array of fatty acids of the n-3 family the content of linolenic, eicosapentaenoic and docosahexaenoic acids (n-3 family) declined. Redistribution of fatty acids in the erythrocytes membrane towards to such alteration in quantity as the increasing of saturation and decreasing of the unsaturated fatty acids supposes the change of its physical and chemical properties, permeability, lability and complexity of passing erythrocyte via microcircular channels.

  12. Epigenomic landscape modified by histone modification correlated with activation of IGF2 gene

    Science.gov (United States)

    The links of histone post-translational modifications and chromatin structure to cell cycle progression, DNA replication, and overall chromosome functions are very clear. The modulation of genome expression as a consequence of chromatin structural changes is most likely a basic mechanism. The epige...

  13. Exploring oxidative modifications of tyrosine

    DEFF Research Database (Denmark)

    Houée-Lévin, C; Bobrowski, K; Horakova, L

    2015-01-01

    residues are oxidised in vivo with impact on cellular homeostasis and redox signalling pathways. A notable example is tyrosine, which can undergo a number of oxidative post-translational modifications to form 3-hydroxy-tyrosine, tyrosine crosslinks, 3-nitrotyrosine and halogenated tyrosine, with different...... effects on cellular functions. Tyrosine oxidation has been studied extensively in vitro, and this has generated detailed information about the molecular mechanisms that may occur in vivo. An important aspect of studying tyrosine oxidation both in vitro and in biological systems is the ability to monitor...... residues modified and the nature of the modification. These approaches have helped understanding of the consequences of tyrosine oxidation in biological systems, especially its effects on cell signalling and cell dysfunction, linking to roles in disease. There is mounting evidence that tyrosine oxidation...

  14. Abdominoplasty in prune belly syndrome: Modifications in Monfort technique to address variable patterns of abdominal wall weakness.

    Science.gov (United States)

    Smith, Edwin A; Srinivasan, Arun; Scherz, Hal C; Tracey, Anthony J; Broecker, Bruce; Kirsch, Andrew J

    2017-10-01

    Abdominoplasty is an important component of the management of children with prune belly syndrome (PBS). While there are features of the abdominal defect in PBS which are common to all patients, there will be differences unique to each patient that should be taken into consideration in surgical planning. Specifically, we have come to realize that although the Monfort procedure assumes a symmetric pattern of abdominal wall laxity, this symmetry is rarely present. The aim of this report is to describe our modifications and review our outcomes for the Monfort procedure which more completely address correction of the abdominal wall laxity including both common and uncommon features while positioning the umbilicus to a more anatomically correct position (Figure). Sixteen male patients with PBS and one female pseudoprune belly syndrome patient, aged 2-9 years, were treated at our institution between 2003 and 2014. Modifications incorporated into the abdominoplasty procedure for PBS applied to this study group included: 1) use of diagnostic laparoscopy to define the topography of the abdominal wall defect, 2) initial midline rather than elliptical skin incision to defer retailoring of the skin coverage until the final step of the procedure, 3) varying the width of the central plate to correct side to side asymmetry in redundancy, 4) plication of the central plate to reduce vertical redundancy and reposition the umbilicus, and 5) plication of focal areas of fascial weakness, most often in the flank region. All patients have improved abdominal wall contour with a more uniform correction of areas of weakness at a mean follow-up of 5.5 years (range 18 months-11.5 years). All patients and parents indicate that they are very satisfied with the outcome of their procedures without any revisions being performed. This study is descriptive in nature and retrospective, with the patient population treated in a relatively uniform fashion that does not allow direct comparison with other

  15. The impact of chromatin modification on the development of chronic complications in patients with diabetes

    Directory of Open Access Journals (Sweden)

    Małgorzata Wegner

    2015-08-01

    Full Text Available Diabetes is a chronic, metabolic disease. Over 347 million people worldwide have diabetes. Chronic complications (retinopathy, nephropathy or neuropathy are the major dangerous outcome of this disease. Recent studies indicate a significant role of epigenetic regulation in the development of chronic complications in patients with diabetes. Hyperglycemia could cause abnormal regulation of the activity of enzymes participating in the post-translational histone modifications (PTHMs and initiation of changes in patterns of DNA methylation. It leads to modification of chromatin structure. These epigenetic abnormalities result in changes in the expression of genes involved in development of chronic inflammation, such as NF-KAPPAB (nuclear factor kappaB gene, TNFα (tumor necrosis factor a gene, IL6 (interleukin 6 gene or MCP1 (monocyte chemoattractant protein 1 gene. It enhances endothelial cell dysfunction, which plays an important role in development of chronic, diabetic complications. In addition, caused by hyperglycemia epigenetic modifications changes in structure of chromatin explains “metabolic memory”, a phenomenon of presence of pathological pathways related to the prolonged hyperglycemia in the past, despite maintaining good metabolic control later on.

  16. Low-fat dietary pattern and weight change over 7 years: the Women's Health Initiative Dietary Modification Trial.

    Science.gov (United States)

    Howard, Barbara V; Manson, JoAnn E; Stefanick, Marcia L; Beresford, Shirley A; Frank, Gail; Jones, Bobette; Rodabough, Rebecca J; Snetselaar, Linda; Thomson, Cynthia; Tinker, Lesley; Vitolins, Mara; Prentice, Ross

    2006-01-04

    Obesity in the United States has increased dramatically during the past several decades. There is debate about optimum calorie balance for prevention of weight gain, and proponents of some low-carbohydrate diet regimens have suggested that the increasing obesity may be attributed, in part, to low-fat, high-carbohydrate diets. To report data on body weight in a long-term, low-fat diet trial for which the primary end points were breast and colorectal cancer and to examine the relationships between weight changes and changes in dietary components. Randomized intervention trial of 48,835 postmenopausal women in the United States who were of diverse backgrounds and ethnicities and participated in the Women's Health Initiative Dietary Modification Trial; 40% (19,541) were randomized to the intervention and 60% (29,294) to a control group. Study enrollment was between 1993 and 1998, and this analysis includes a mean follow-up of 7.5 years (through August 31, 2004). The intervention included group and individual sessions to promote a decrease in fat intake and increases in vegetable, fruit, and grain consumption and did not include weight loss or caloric restriction goals. The control group received diet-related education materials. Change in body weight from baseline to follow-up. Women in the intervention group lost weight in the first year (mean of 2.2 kg, Pfruit servings, and a nonsignificant trend toward weight loss occurred with increasing intake of fiber. A low-fat eating pattern does not result in weight gain in postmenopausal women. Clinical Trial Registration ClinicalTrials.gov, NCT00000611.

  17. Low-fat dietary pattern and cancer incidence in the Women's Health Initiative Dietary Modification Randomized Controlled Trial.

    Science.gov (United States)

    Prentice, Ross L; Thomson, Cynthia A; Caan, Bette; Hubbell, F Allan; Anderson, Garnet L; Beresford, Shirley A A; Pettinger, Mary; Lane, Dorothy S; Lessin, Lawrence; Yasmeen, Shagufta; Singh, Baljinder; Khandekar, Janardan; Shikany, James M; Satterfield, Suzanne; Chlebowski, Rowan T

    2007-10-17

    The Women's Health Initiative Dietary Modification (DM) Randomized Controlled Trial evaluated the effects of a low-fat dietary pattern on chronic disease incidence, with breast cancer and colorectal cancer as primary outcomes. The trial protocol also listed ovarian cancer and endometrial cancer as outcomes that may be favorably affected by the intervention. A total of 48,835 postmenopausal women were randomly assigned during 1993-1998 to a DM intervention (n = 19,541) or comparison (usual diet; n = 29,294) group and followed up for an average of 8.1 years. The intervention goal was to reduce total fat intake to 20% of energy and to increase consumption of vegetables, fruits, and grains. Cancer outcomes were verified by pathology report review. We used weighted log-rank tests to compare incidence of invasive cancers of the ovary and endometrium, total invasive cancer, and invasive cancers at other sites between the groups. All statistical tests were two-sided. Ovarian cancer risk was lower in the intervention than in the comparison group (P = .03). Although the overall ovarian cancer hazard ratio (HR) was not statistically significantly less than 1.0, the hazard ratio decreased with increasing intervention duration (P(trend) = .01). For the first 4 years, the risk for ovarian cancer was similar in the intervention and control groups (0.52 cases per 1000 person-years in the intervention group versus 0.45 per 1000 person-years in the comparison group; HR = 1.16, 95% confidence interval [CI] = 0.73 to 1.84); over the next 4.1 years, the risk was lower in the intervention group (0.38 cases per 1000 person-years in the intervention group versus 0.64 per 1000 person-years in the comparison group; HR = 0.60, 95% CI = 0.38 to 0.96). Risk of cancer of the endometrium did not differ between the groups (P = .18). The estimated risk of total invasive cancer was slightly lower in the intervention group than in the control group (HR = 0.95, 95% CI = 0.89 to 1.01; P = .10). A low

  18. Precision mapping of coexisting modifications in histone H3 tails from embryonic stem cells by ETD-MS/MS

    DEFF Research Database (Denmark)

    Jung, Hye Ryung; Sidoli, Simone; Haldbo, Simon

    2013-01-01

    Post-translational modifications (PTMs) of histones play a major role in regulating chromatin dynamics and influence processes such as transcription and DNA replication. Here, we report 114 distinct combinations of coexisting PTMs of histone H3 obtained from mouse embryonic stem (ES) cells. Histo...

  19. First report of nonpsychotic self-cannibalism (autophagy), tongue splitting, and scar patterns (scarification) as an extreme form of cultural body modification in a western civilization.

    Science.gov (United States)

    Benecke, M

    1999-09-01

    As part of her current lifestyle, a 28-year-old Caucasian woman routinely injures and allows subsequent healing of her skin and other tissues. Her body modifications include a "split tongue" (a tongue split to the base), which does not interfere with speaking and eating. Other modifications include large scarification patterns produced by branding and cutting. This woman has been known to eat parts of her skin that were previously cut out of her body. She also performs "needle play" by allowing medical syringe needles to be lodged temporarily under her skin. The patient had a normal childhood, is currently employed full-time as an office manager, and is psychologically stable. Although one other case of self-induced penoscrotal hypospadias is known, this is the only report of extensive, nonpsychotic, autodestructive behavior. However, this may not be the case in the future as an increasing number of young individuals have become interested in body modifications.

  20. Functional O-GlcNAc modifications: Implications in molecular regulation and pathophysiology

    Science.gov (United States)

    Wells, Lance

    2016-01-01

    O-linked β-N-acetylglucosamine (O-GlcNAc) is a regulatory post-translational modification of intracellular proteins. The dynamic and inducible cycling of the modification is governed by O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA) in response to UDP-GlcNAc levels in the hexosamine biosynthetic pathway (HBP). Due to its reliance on glucose flux and substrate availability, a major focus in the field has been on how O-GlcNAc contributes to metabolic disease. For years this post-translational modification has been known to modify thousands of proteins implicated in various disorders, but direct functional connections have until recently remained elusive. New research is beginning to reveal the specific mechanisms through which O-GlcNAc influences cell dynamics and disease pathology including clear examples of O-GlcNAc modification at a specific site on a given protein altering its biological functions. The following review intends to focus primarily on studies in the last half decade linking O-GlcNAc modification of proteins with chromatin-directed gene regulation, developmental processes, and several metabolically related disorders including Alzheimer’s, heart disease and cancer. These studies illustrate the emerging importance of this post-translational modification in biological processes and multiple pathophysiologies. PMID:24524620

  1. Identification and Interrogation of Combinatorial Histone Modifications

    Directory of Open Access Journals (Sweden)

    Kelly R Karch

    2013-12-01

    Full Text Available Histone proteins are dynamically modified to mediate a variety of cellular processes including gene transcription, DNA damage repair, and apoptosis. Regulation of these processes occurs through the recruitment of non-histone proteins to chromatin by specific combinations of histone post-translational modifications (PTMs. Mass spectrometry has emerged as an essential tool to discover and quantify histone PTMs both within and between samples in an unbiased manner. Developments in mass spectrometry that allow for characterization of large histone peptides or intact protein has made it possible to determine which modifications occur simultaneously on a single histone polypeptide. A variety of techniques from biochemistry, biophysics, and chemical biology have been employed to determine the biological relevance of discovered combinatorial codes. This review first describes advancements in the field of mass spectrometry that have facilitated histone PTM analysis and then covers notable approaches to probe the biological relevance of these modifications in their nucleosomal context.

  2. An ion-current mutant of Paramecium tetraurelia with defects in the primary structure and post-translational N-methylation of calmodulin

    International Nuclear Information System (INIS)

    Wallen-Friedman, M.A.

    1988-01-01

    My work on pantophobiac A 2 (pntA 2 ), a behavioral mutant of Paramecium tetraurelia, suggest that the Ca ++ -binding protein calmodulin (CaM), and post-translation N-methylation of CaM, are important for Ca ++ -related ion-current function. Calmodulin from wild-type Paramecium has two sites of lysine-N-methylation. Both of these sites are almost fully methylated in vivo; thus wild-type calmodulin is a poor substrate for N-methylation in vitro. In contrast, pntA/ 2 CaM can be heavily N-methylated in vitro, suggesting that the mutant calmodulin is under-methylated in vivo. Amino-acid composition analysis showed that CaM lysine 115 is undermethylated in pntA 2 . Once pntA 2 CaM is N-methylated, the [methyl- 3 H] group does not turn over in either wild-type or pntA 2 cytoplasmic fractions. The methylating enzymes in pntA 2 high-speed supernatant fractions are active, but may be less robust than those of the wild type, suggesting a possible control of these enzymes by CaM

  3. The Wheat NAC Transcription Factor TaNAC2L Is Regulated at the Transcriptional and Post-Translational Levels and Promotes Heat Stress Tolerance in Transgenic Arabidopsis.

    Science.gov (United States)

    Guo, Weiwei; Zhang, Jinxia; Zhang, Ning; Xin, Mingming; Peng, Huiru; Hu, Zhaorong; Ni, Zhongfu; Du, Jinkun

    2015-01-01

    Heat stress poses a serious threat to global crop production. In efforts that aim to mitigate the adverse effects of heat stress on crops, a variety of genetic tools are being used to develop plants with improved thermotolerance. The characterization of important regulators of heat stress tolerance provides essential information for this aim. In this study, we examine the wheat (Triticum aestivum) NAC transcription factor gene TaNAC2L. High temperature induced TaNAC2L expression in wheat and overexpression of TaNAC2L in Arabidopsis thaliana enhanced acquired heat tolerance without causing obvious alterations in phenotype compared with wild type under normal conditions. TaNAC2L overexpression also activated the expression of heat-related genes in the transgenic Arabidopsis plants, suggesting that TaNAC2L may improve heat tolerance by regulating the expression of stress-responsive genes. Notably, TaNAC2L is also regulated at the post-translational level and might be degraded via a proteasome-mediated pathway. Thus, this wheat transcription factor may have potential uses in enhancing thermotolerance in crops.

  4. The Wheat NAC Transcription Factor TaNAC2L Is Regulated at the Transcriptional and Post-Translational Levels and Promotes Heat Stress Tolerance in Transgenic Arabidopsis.

    Directory of Open Access Journals (Sweden)

    Weiwei Guo

    Full Text Available Heat stress poses a serious threat to global crop production. In efforts that aim to mitigate the adverse effects of heat stress on crops, a variety of genetic tools are being used to develop plants with improved thermotolerance. The characterization of important regulators of heat stress tolerance provides essential information for this aim. In this study, we examine the wheat (Triticum aestivum NAC transcription factor gene TaNAC2L. High temperature induced TaNAC2L expression in wheat and overexpression of TaNAC2L in Arabidopsis thaliana enhanced acquired heat tolerance without causing obvious alterations in phenotype compared with wild type under normal conditions. TaNAC2L overexpression also activated the expression of heat-related genes in the transgenic Arabidopsis plants, suggesting that TaNAC2L may improve heat tolerance by regulating the expression of stress-responsive genes. Notably, TaNAC2L is also regulated at the post-translational level and might be degraded via a proteasome-mediated pathway. Thus, this wheat transcription factor may have potential uses in enhancing thermotolerance in crops.

  5. Conserved Residues Lys57 and Lys401 of Protein Disulfide Isomerase Maintain an Active Site Conformation for Optimal Activity: Implications for Post-Translational Regulation

    Directory of Open Access Journals (Sweden)

    Cody Caba

    2018-02-01

    Full Text Available Despite its study since the 1960's, very little is known about the post-translational regulation of the multiple catalytic activities performed by protein disulfide isomerase (PDI, the primary protein folding catalyst of the cell. This work identifies a functional role for the highly conserved CxxC-flanking residues Lys57 and Lys401 of human PDI in vitro. Mutagenesis studies have revealed these residues as modulating the oxidoreductase activity of PDI in a pH-dependent manner. Non-conservative amino acid substitutions resulted in enzyme variants upwards of 7-fold less efficient. This attenuated activity was found to translate into a 2-fold reduction of the rate of electron shuttling between PDI and the intraluminal endoplasmic reticulum oxidase, ERO1α, suggesting a functional significance to oxidative protein folding. In light of this, the possibility of lysine acetylation at residues Lys57 and Lys401 was assessed by in vitro treatment using acetylsalicylic acid (aspirin. A total of 28 acetyllysine residues were identified, including acLys57 and acLys401. The kinetic behavior of the acetylated protein form nearly mimicked that obtained with a K57/401Q double substitution variant providing an indication that acetylation of the active site-flanking lysine residues can act to reversibly modulate PDI activity.

  6. Conserved Residues Lys57 and Lys401 of Protein Disulfide Isomerase Maintain an Active Site Conformation for Optimal Activity: Implications for Post-Translational Regulation.

    Science.gov (United States)

    Caba, Cody; Ali Khan, Hyder; Auld, Janeen; Ushioda, Ryo; Araki, Kazutaka; Nagata, Kazuhiro; Mutus, Bulent

    2018-01-01

    Despite its study since the 1960's, very little is known about the post-translational regulation of the multiple catalytic activities performed by protein disulfide isomerase (PDI), the primary protein folding catalyst of the cell. This work identifies a functional role for the highly conserved CxxC-flanking residues Lys 57 and Lys 401 of human PDI in vitro . Mutagenesis studies have revealed these residues as modulating the oxidoreductase activity of PDI in a pH-dependent manner. Non-conservative amino acid substitutions resulted in enzyme variants upwards of 7-fold less efficient. This attenuated activity was found to translate into a 2-fold reduction of the rate of electron shuttling between PDI and the intraluminal endoplasmic reticulum oxidase, ERO1α, suggesting a functional significance to oxidative protein folding. In light of this, the possibility of lysine acetylation at residues Lys 57 and Lys 401 was assessed by in vitro treatment using acetylsalicylic acid (aspirin). A total of 28 acetyllysine residues were identified, including acLys 57 and acLys 401 . The kinetic behavior of the acetylated protein form nearly mimicked that obtained with a K57/401Q double substitution variant providing an indication that acetylation of the active site-flanking lysine residues can act to reversibly modulate PDI activity.

  7. Interaction proteins of invertase and invertase inhibitor in cold-stored potato tubers suggested a protein complex underlying post-translational regulation of invertase.

    Science.gov (United States)

    Lin, Yuan; Liu, Jun; Liu, Xun; Ou, Yongbin; Li, Meng; Zhang, Huiling; Song, Botao; Xie, Conghua

    2013-12-01

    The activity of vacuolar invertase (VI) is vital to potato cold-induced sweetening (CIS). A post-translational regulation of VI activity has been proposed which involves invertase inhibitor (VIH), but the mechanism for the interaction between VI and VIH has not been fully understood. To identify the potential partners of VI and VIH, two cDNA libraries were respectively constructed from CIS-resistant wild potato species Solanum berthaultii and CIS-sensitive potato cultivar AC035-01 for the yeast two-hybrid analysis. The StvacINV1 (one of the potato VIs) and StInvInh2B (one of the potato VIHs), previously identified to be associated with potato CIS, were used as baits to screen the two libraries. Through positive selection and sequencing, 27 potential target proteins of StvacINV1 and eight of StInvInh2B were clarified. The Kunitz-type protein inhibitors were captured by StvacINV1 in both libraries and the interaction between them was confirmed by bimolecular fluorescence complementation assay in tobacco cells, reinforcing a fundamental interaction between VI and VIH. Notably, a sucrose non-fermenting-1-related protein kinase 1 was captured by both the baits, suggesting that a protein complex could be necessary for fine turning of the invertase activity. The target proteins clarified in present research provide a route to elucidate the mechanism by which the VI activity can be subtly modulated. Copyright © 2013 Elsevier Masson SAS. All rights reserved.

  8. Standardization and quality control in quantifying non-enzymatic oxidative protein modifications in relation to ageing and disease: Why is it important and why is it hard?

    DEFF Research Database (Denmark)

    Nedić, Olgica; Rogowska-Wrzesinska, Adelina; Rattan, Suresh

    2015-01-01

    Post-translational modifications (PTM) of proteins determine the activity, stability, specificity, transportability and lifespan of a protein. Some PTM are highly specific and regulated involving various enzymatic pathways, but there are other non-enzymatic PTM (nePTM), which occur stochastically...

  9. Designing and Producing Modified, New-to-Nature Peptides with Antimicrobial Activity by Use of a Combination of Various Lantibiotic Modification Enzymes

    NARCIS (Netherlands)

    van Heel, Auke J.; Mu, Dongdong; Montalban-Lopez, Manuel; Hendriks, Djoke; Kuipers, Oscar P.

    Lanthipeptides are peptides that contain several post-translationally modified amino acid residues and commonly show considerable antimicrobial activity. After translation, the amino acid residues of these peptides are modified by a distinct set of modification enzymes. This process results in

  10. Crosstalk between histone modifications maintains the developmental pattern of gene expression on a tissue-specific locus.

    Science.gov (United States)

    Hosey, Alison M; Chaturvedi, Chandra-Prakash; Brand, Marjorie

    2010-05-16

    Genome wide studies have provided a wealth of information related to histone modifications. Particular modifications, which can encompass both broad and discrete regions, are associated with certain genomic elements and gene expression status. Here we focus on how studies on the beta-globin gene cluster can complement the genome wide effort through the thorough dissection of histone modifying protein crosstalk. The beta-globin locus serves as a model system to study both regulation of gene expression driven at a distance by enhancers and mechanisms of developmental switching of clustered genes. We investigate recent studies, which uncover that histone methyltransferases, recruited at the beta-globin enhancer, control gene expression by long range propagation on chromatin. Specifically, we focus on how seemingly antagonistic complexes, such as those including MLL2, G9a and UTX, can cooperate to functionally regulate developmentally controlled gene expression. Finally, we speculate on the mechanisms of chromatin modifying complex propagation on genomic domains.

  11. High Energy Conversion Efficiency with 3-D Micro-Patterned Photoanode for Enhancement Diffusivity and Modification of Photon Distribution in Dye-Sensitized Solar Cells.

    Science.gov (United States)

    Yun, Min Ju; Sim, Yeon Hyang; Cha, Seung I; Seo, Seon Hee; Lee, Dong Y

    2017-11-08

    Dye sensitize solar cells (DSSCs) have been considered as the promising alternatives silicon based solar cell with their characteristics including high efficiency under weak illumination and insensitive power output to incident angle. Therefore, many researches have been studied to improve the energy conversion efficiency of DSSCs. However the efficiency of DSSCs are still trapped at the around 10%. In this study, micro-scale hexagonal shape patterned photoanode have proposed to modify light distribution of photon. In the patterned electrode, the appearance efficiency have been obtained from 7.1% to 7.8% considered active area and the efficiency of 12.7% have been obtained based on the photoanode area. Enhancing diffusion of electrons and modification of photon distribution utilizing the morphology of the electrode are major factors to improving the performance of patterned electrode. Also, finite element method analyses of photon distributions were conducted to estimate morphological effect that influence on the photon distribution and current density. From our proposed study, it is expecting that patterned electrode is one of the solution to overcome the stagnant efficiency and one of the optimized geometry of electrode to modify photon distribution. Process of inter-patterning in photoanode has been minimized.

  12. Ezh2 regulates transcriptional and post-translational expression of T-bet and promotes Th1 cell responses mediating aplastic anemia in mice1

    Science.gov (United States)

    Tong, Qing; He, Shan; Xie, Fang; Mochizuki, Kazuhiro; Liu, Yongnian; Mochizuki, Izumi; Meng, Lijun; Sun, Hongxing; Zhang, Yanyun; Guo, Yajun; Hexner, Elizabeth; Zhang, Yi

    2014-01-01

    Acquired aplastic anemia (AA) is a potentially fatal bone marrow (BM) failure syndrome. IFN-γ-producing T helper (Th)1 CD4+ T cells mediate the immune destruction of hematopoietic cells, and are central to the pathogenesis. However, the molecular events that control the development of BM-destructive Th1 cells remain largely unknown. Ezh2 is a chromatin-modifying enzyme that regulates multiple cellular processes primarily by silencing gene expression. We recently reported that Ezh2 is crucial for inflammatory T cell responses after allogeneic BM transplantation. To elucidate whether Ezh2 mediates pathogenic Th1 responses in AA and the mechanism of Ezh2 action in regulating Th1 cells, we studied the effects of Ezh2 inhibition in CD4+ T cells using a mouse model of human AA. Conditionally deleting Ezh2 in mature T cells dramatically reduced the production of BM-destructive Th1 cells in vivo, decreased BM-infiltrating Th1 cells, and rescued mice from BM failure. Ezh2 inhibition resulted in significant decrease in the expression of Tbx21 and Stat4 (which encode transcription factors T-bet and STAT4, respectively). Introduction of T-bet but not STAT4 into Ezh2-deficient T cells fully rescued their differentiation into Th1 cells mediating AA. Ezh2 bound to the Tbx21 promoter in Th1 cells, and directly activated Tbx21 transcription. Unexpectedly, Ezh2 was also required to prevent proteasome-mediated degradation of T-bet protein in Th1 cells. Our results identify T-bet as the transcriptional and post-translational Ezh2 target that acts together to generate BM-destructive Th1 cells, and highlight the therapeutic potential of Ezh2 inhibition in reducing AA and other autoimmune diseases. PMID:24760151

  13. Post-translational amino acid racemization in the frog skin peptide deltorphin I in the secretion granules of cutaneous serous glands.

    Science.gov (United States)

    Auvynet, Constance; Seddiki, Nabila; Dunia, Irene; Nicolas, Pierre; Amiche, Mohamed; Lacombe, Claire

    2006-01-01

    The dermal glands of the South American hylid frog Phyllomedusa bicolor synthesize and expel huge amounts of cationic, alpha-helical, 24- to 33-residue antimicrobial peptides, the dermaseptins B. These glands also produce a wide array of peptides that are similar to mammalian hormones and neuropeptides, including a heptapeptide opioid containing a D-amino acid, deltorphin I (Tyr-DAla-Phe-Asp-Val-Val-Gly NH2). Its biological activity is due to the racemization of L-Ala2 to D-Ala. The dermaseptins B and deltorphins are all derived from a single family of precursor polypeptides that have an N-terminal preprosequence that is remarkably well conserved, although the progenitor sequences giving rise to mature opioid or antimicrobial peptides are markedly different. Monoclonal and polyclonal antibodies were used to examine the cellular and ultrastructural distributions of deltorphin I and dermaseptin B in the serous glands by immunofluoresence confocal microscopy and immunogold-electron microscopy. Preprodeltorphin I and preprodermaseptins B are sorted into the regulated pathway of secretion, where they are processed to give the mature products. Deltorphin I, [l-Ala2]-deltorphin I and dermaseptin B are all stored together in secretion granules which accumulate in the cytoplasm of all serous glands. We conclude that the L- to D-amino acid isomerization of the deltorphin I occurs in the secretory granules as a post-translational event. Thus the specificity of isomerization depends on the presence of structural and/or conformational determinants in the peptide N-terminus surrounding the isomerization site.

  14. Post-Translational Regulation of Polycystin-2 Protein Expression as a Novel Mechanism of Cholangiocyte Reaction and Repair from Biliary Damage

    Science.gov (United States)

    Spirli, Carlo; Villani, Ambra; Mariotti, Valeria; Fabris, Luca; Fiorotto, Romina; Strazzabosco, Mario

    2015-01-01

    Polycystin-2 (PC2 /TRPP2), a member of the transient receptor potential channels (TRP) family, is a non-selective calcium channel. Mutations in PC2/TRPP2 are associated with Polycystic Liver Diseases. PC2-defective cholangiocytes shows increased production of cAMP, PKA-dependent activation of the ERK1/2 pathway, HIF1α-mediated VEGF production, and stimulation of cyst growth and progression. Activation of the ERK/HIF1α/VEGF pathway in cholangiocytes plays a key role during repair from biliary damage. We hypothesized that PC2 levels are modulated during biliary damage/repair, resulting in activation of the ERK/HIF1α/VEGF pathway. Results PC2 protein expression, but not its gene expression, was significantly reduced in mouse livers with biliary damage (Mdr2−/−-KO, bile duct ligation, DDC-treatment). Treatment of colangiocytes with pro-inflammatory cytokines, nitric oxide (NO) donors and ER stressors), increased ERK1/2 phosphorylation, HIF1α transcriptional activity, secretion of VEGF, VEGFR2 phosphorylation and downregulated PC2 protein expression without affecting PC2 gene expression. Expression of Herp and NEK, ubiquitin-like proteins that promote proteosomal PC2 degradation was increased. Pre-treatment with the proteasome inhibitor MG-132 restored the expression of PC2 in cells treated with cytokines but not in cells treated with NO donors or with ER stressors. In these conditions, PC2 degradation was instead inhibited by interfering with the autophagy pathway. Treatment of DDC-mice and of Mdr2−/−-mice with the proteasome inhibitor bortezomib, restored PC2 expression and significantly reduced the ductular reaction, fibrosis and p-ERK1/2. In conclusion, in response to biliary damage, PC2 expression is modulated post-translationally by the proteasome or the autophagy pathways. PC2-dowregulation is associated with activation of ERK1/2 and increase of HIF1α-mediated VEGF secretion. Treatments able to restore PC2 expression and to reduce ductular reaction

  15. Habitat Modification by the Leaf-Cutter Ant, Atta cephalotes, and Patterns of Leaf-Litter Arthropod Communities.

    Science.gov (United States)

    Wells, Rachel L; Murphy, Serena K; Moran, Matthew D

    2017-12-08

    Ecosystem engineers are profoundly important in many biological communities. A Neotropical taxonomic group considered to have engineering effects is the Formicidae (ants). Leaf-cutter ants (LCAs), in particular, which form extensive colonies of millions of individuals, can be important ecosystem engineers in these environments. While the effects of LCAs on plant community structure and soil chemistry are well-studied, their effects on consumers are poorly understood. Therefore, we examined the indirect effects of the LCA Atta cephalotes L. on the leaf-litter arthropod community. We compared abundance and diversity patterns at ant nests to areas distant from nests, utilizing both a factorial design and gradient analysis for both nocturnal and diurnal arthropods. We found that arthropod abundance and diversity was significantly lower for multiple taxonomic groups and trophic levels near leaf-cutter nests, and this pattern was strongest at night. Exceptions to this pattern included two morphospecies of Collembola that were more abundant on nests, suggesting some specialization for these species. For the gradient analysis, abundance increased exponentially for most groups of arthropods. However, for the dominant arthropod species, the amphipod Cerrorchestia hyloraina Lindeman, a quadratic function was the best fit curvilinear model for abundance. It appeared that C. hyloraina had maximal abundance at the transition between nest site and less disturbed forest. These results indicate that LCA activity has a strong effect on the leaf-litter arthropod community, adding to spatial heterogeneity within neotropical forests. These effects may translate into changes in important ecological processes such as nutrient cycling and food web function. © The Authors 2017. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  16. Modification of epigenetic patterns in low birth weight children: importance of hypomethylation of the ACE gene promoter.

    Science.gov (United States)

    Rangel, Marina; dos Santos, Jéssica Cassilla; Ortiz, Paula Helena Lima; Hirata, Mario; Jasiulionis, Miriam Galvonas; Araujo, Ronaldo C; Ierardi, Daniela Filippini; Franco, Maria do Carmo

    2014-01-01

    There is a growing body of evidence that epigenetic alterations are involved in the pathological mechanisms of many chronic disorders linked to fetal programming. Angiotensin-converting enzyme (ACE) appears as one candidate gene that brings new insights into the epigenetic control and later development of diseases. In this view, we have postulated that epigenetic modifications in the ACE gene might show different interactions between birth weight (BW), blood pressure levels, plasma ACE activity and ACE I/D polymorphism. To explore this hypothesis, we performed a cross-sectional study to evaluate the DNA methylation of 3 CpG sites using pyrosequencing within the ACE gene promoter of peripheral blood leukocytes from 45 LBW children compared with 70 NBW children. Our results have revealed that LBW children have lower methylation levels (PACE activity (P = 0.001). Adjusting for prematurity, gender, age, body mass index, and family history of cardiovascular disease did not alter these findings. We have also performed analyses of individual CpG sites. The frequency of DNA methylation was significantly different at two CpG sites (site 1: nucleotide position +555; and site 3: nucleotide position +563). In addition, we have found a significant inverse correlation between degree of DNA methylation and both ACE activity (PACE gene promoter is associated with LBW in 6 to 12 year-old children. The magnitude of these epigenetic changes appears to be clinically important, which is supported by the observation that discrete changes in DNA methylation can affect systolic blood pressure and ACE protein activity levels.

  17. Selective hydrophilic modification of Parylene C films: a new approach to cell micro-patterning for synthetic biology applications

    International Nuclear Information System (INIS)

    Trantidou, T; Toumazou, C; Prodromakis, T; Rao, C; Barrett, H; Camelliti, P; Pinto, K; Yacoub, M H; Terracciano, C M; Athanasiou, T

    2014-01-01

    We demonstrate a simple, accurate and versatile method to manipulate Parylene C, a material widely known for its high biocompatibility, and transform it to a substrate that can effectively control the cellular microenvironment and consequently affect the morphology and function of the cells in vitro. The Parylene C scaffolds are fabricated by selectively increasing the material's surface water affinity through lithography and oxygen plasma treatment, providing free bonds for attachment of hydrophilic biomolecules. The micro-engineered constructs were tested as culture scaffolds for rat ventricular fibroblasts and neonatal myocytes (NRVM), toward modeling the unique anisotropic architecture of native cardiac tissue. The scaffolds induced the patterning of extracellular matrix compounds and therefore of the cells, which demonstrated substantial alignment compared to typical unstructured cultures. Ca 2+  cycling properties of the NRVM measured at rates of stimulation 0.5–2 Hz were significantly modified with a shorter time to peak and time to 90% decay, and a larger fluorescence amplitude (p < 0.001). The proposed technique is compatible with standard cell culturing protocols and exhibits long-term pattern durability. Moreover, it allows the integration of monitoring modalities into the micro-engineered substrates for a comprehensive interrogation of physiological parameters. (paper)

  18. Low-fat Dietary Pattern and Pancreatic Cancer Risk in the Women's Health Initiative Dietary Modification Randomized Controlled Trial.

    Science.gov (United States)

    Jiao, Li; Chen, Liang; White, Donna L; Tinker, Lesley; Chlebowski, Rowan T; Van Horn, Linda V; Richardson, Peter; Lane, Dorothy; Sangi-Haghpeykar, Haleh; El-Serag, Hashem B

    2018-01-01

    Observational studies suggest that diet may influence pancreatic cancer risk. We investigated the effect of a low-fat dietary intervention on pancreatic cancer incidence. The Women's Health Initiative Dietary Modification (WHI-DM) trial is a randomized controlled trial conducted in 48 835 postmenopausal women age 50 to 79 years in the United States between 1993 and 1998. Women were randomly assigned to the intervention group (n = 19 541), with the goal of reducing total fat intake and increasing intake of vegetables, fruits, and grains, or to the usual diet comparison group (n = 29 294). The intervention concluded in March 2005. We evaluated the effect of the intervention on pancreatic cancer incidence with the follow-up through 2014 using the log-rank test and multivariable Cox proportional hazards regression model. All statistical tests were two-sided. In intention-to-treat analyses including 46 200 women, 92 vs 165 pancreatic cancer cases were ascertained in the intervention vs the comparison group (P = .23). The multivariable hazard ratio (HR) of pancreatic cancer was 0.86 (95% confidence interval [CI] = 0.67 to 1.11). Risk was statistically significantly reduced among women with baseline body mass indexes (BMIs) of 25 kg/m2 or higher (HR = 0.71, 95% CI = 0.53 to 0.96), but not among women with BMIs of less than 25 kg/m2 (HR = 1.62, 95% CI = 0.97 to 2.71, Pinteraction = .01). A low-fat dietary intervention was associated with reduced pancreatic cancer incidence in women who were overweight or obese in the WHI-DM trial. Caution needs to be taken in interpreting the findings based on subgroup analyses. © The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  19. Selective hydrophilic modification of Parylene C films: a new approach to cell micro-patterning for synthetic biology applications.

    Science.gov (United States)

    Trantidou, T; Rao, C; Barrett, H; Camelliti, P; Pinto, K; Yacoub, M H; Athanasiou, T; Toumazou, C; Terracciano, C M; Prodromakis, T

    2014-06-01

    We demonstrate a simple, accurate and versatile method to manipulate Parylene C, a material widely known for its high biocompatibility, and transform it to a substrate that can effectively control the cellular microenvironment and consequently affect the morphology and function of the cells in vitro. The Parylene C scaffolds are fabricated by selectively increasing the material's surface water affinity through lithography and oxygen plasma treatment, providing free bonds for attachment of hydrophilic biomolecules. The micro-engineered constructs were tested as culture scaffolds for rat ventricular fibroblasts and neonatal myocytes (NRVM), toward modeling the unique anisotropic architecture of native cardiac tissue. The scaffolds induced the patterning of extracellular matrix compounds and therefore of the cells, which demonstrated substantial alignment compared to typical unstructured cultures. Ca(2+) cycling properties of the NRVM measured at rates of stimulation 0.5-2 Hz were significantly modified with a shorter time to peak and time to 90% decay, and a larger fluorescence amplitude (p micro-engineered substrates for a comprehensive interrogation of physiological parameters.

  20. Biochemical and physiological modifications in tissues of Sardina pilchardus: spatial and temporal patterns as a baseline for biomonitoring studies

    Directory of Open Access Journals (Sweden)

    Bruno Silva Nunes

    2015-02-01

    Full Text Available Sardina pilchardus is a marine species common in the North Atlantic Ocean, and is subjected to diffuse anthropogenic chemical contamination and seasonal fluctuations in biotic and abiotic parameters that may alter its physiology and condition. Biological material is easily available through commercial fisheries, which could facilitate its use as a bioindicator species. The aim of the present work was to address its potential inclusion in biomonitoring studies, considering a combinatory approach through the use of enzymatic biomarkers and somatic indices, by assessing spatial and temporal patterns in a metapopulation along the west coast of Portugal. Our results showed significant variability of the biochemical and physiological profile of the fish, mainly concordant between sampling sites. Large differences for most markers were found across periods of the year, showing the importance of seasonality, which was mostly related to the reproductive cycle. Hence, environmental scientists should acknowledge seasonality as a strong driving force for physiological adaptations, influencing biochemical markers that are normally used to identify effects of chemical contamination. The here-obtained set of data suggests that S. pilchardus may be successfully included in oceanic biomonitoring studies, when one considers that the contribution of seasonal factors may exceed the influence of eventual anthropogenic contamination.

  1. ICRPfinder: a fast pattern design algorithm for coding sequences and its application in finding potential restriction enzyme recognition sites

    Directory of Open Access Journals (Sweden)

    Stafford Phillip

    2009-09-01

    Full Text Available Abstract Background Restriction enzymes can produce easily definable segments from DNA sequences by using a variety of cut patterns. There are, however, no software tools that can aid in gene building -- that is, modifying wild-type DNA sequences to express the same wild-type amino acid sequences but with enhanced codons, specific cut sites, unique post-translational modifications, and other engineered-in components for recombinant applications. A fast DNA pattern design algorithm, ICRPfinder, is provided in this paper and applied to find or create potential recognition sites in target coding sequences. Results ICRPfinder is applied to find or create restriction enzyme recognition sites by introducing silent mutations. The algorithm is shown capable of mapping existing cut-sites but importantly it also can generate specified new unique cut-sites within a specified region that are guaranteed not to be present elsewhere in the DNA sequence. Conclusion ICRPfinder is a powerful tool for finding or creating specific DNA patterns in a given target coding sequence. ICRPfinder finds or creates patterns, which can include restriction enzyme recognition sites, without changing the translated protein sequence. ICRPfinder is a browser-based JavaScript application and it can run on any platform, in on-line or off-line mode.

  2. Sorption-reduction coupled gold recovery process boosted by Pycnoporus sanguineus biomass: Uptake pattern and performance enhancement via biomass surface modification.

    Science.gov (United States)

    Shi, Chaohong; Zhu, Nengwu; Kang, Naixin; Wu, Pingxiao; Zhang, Xiaoping; Zhang, Yanhong

    2017-09-01

    Biorecovery is emerging as a promising process to retrieve gold from secondary resources. The present study aimed to explore the uptake pattern of Pycnoporus sanguineus biomass for gold, identify the effective functional groups in gold recovery process, and thus further intensify the process via microbial surface modification. Results showed that P. sanguineus biomass could effectively recover gold with the formation of highly crystal AuNPs without any exogeneous electron donor. Under the conditions of various initial gold concentrations (1.0, 2.0, and 3.0 mM), biomass dosage of 2.0 g/L, solution pH value of 4.0, and incubation temperature of 30°C, the uptake equilibrium established after 4, 8, and 12 h, respectively. The uptake process could be well described by pseudo-second order kinetics model (R 2  = 0.9988) and Langmuir isotherm model (R 2  = 0.9958). The maximum uptake capacity of P. sanguineus reached as high as 358.69 mg/g. Further analysis indicated that amino, carboxyl and hydroxyl groups positively contributed to the uptake process. Among them, amino group significantly favored the uptake of gold during recovery process. When P. sanguineus biomass was modified by introduction of amino group, the gold uptake process was successfully intensified by shortening the uptake period and enhancing the uptake capacity. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 33:1314-1322, 2017. © 2017 American Institute of Chemical Engineers.

  3. The Human Thioredoxin System: Modifications and Clinical Applications

    Directory of Open Access Journals (Sweden)

    Seyed Isaac Hashemy

    2011-03-01

    Full Text Available The thioredoxin system, comprising thioredoxin (Trx, thioredoxin reductase (TrxR and NADPH, is one of the major cellular antioxidant systems, implicated in a large and growing number of biological functions. Trx acts as an oxidoreductase via a highly conserved dithiol/disulfide motif located in the active site (-Trp-Cys-Gly-Pro-Cys-Lys-. Different factors are involved in the regulation of Trx activity, including its expression level, localization, protein-protein interactions, post-translational modifications and some chemical inhibitors. Mammalian TrxRs are selenoproteins which have a –Cys-Val-Asn-Val-Gly-Cys- N-terminal active site, as well as a C-terminal selenium-containing active site. Besides two Cys-residues in the redox-regulatory domain of cytosolic Trx (Trx1, human Trx1 has three additional Cys-residues. Post-translational modifications of human Trx1 which are involved in the regulation of its activity can happen via modification of Cys-residues including thiol oxidation, glutathionylation and S-nitrosylation or via modification of other amino acid residues such as nitration of Tyr-49. Because of the numerous functions of the thioredoxin system, its inhibition (mainly happens via the targeting TrxR can result in major cellular consequences, which are potentially pro-oxidant in nature, leading to cell death via necrosis or apoptosis if overexpression of Trx and other antioxidative enzymes can not recuperate cell response. Considering this feature, several anticancer drugs have been used which can inhibit TrxR. Elevated levels of Trx and/or TrxR have been reported in many different human malignancies, positively correlated with aggressive tumor growth and poor prognosis. Moreover, anti-oxidative and anti-apoptotic effects of Trx are reasons to study its clinical application as a drug.

  4. Fatty acid modification of Wnt1 and Wnt3a at serine is prerequisite for lipidation at cysteine and is essential for Wnt signalling

    Czech Academy of Sciences Publication Activity Database

    Doubravská, Lenka; Krausová, Michaela; Gradl, D.; Vojtěchová, Martina; Tůmová, Lucie; Lukáš, Jan; Valenta, Tomáš; Pospíchalová, Vendula; Fafílek, Bohumil; Plachý, Jiří; Sebesta, O.; Kořínek, Vladimír

    2011-01-01

    Roč. 23, č. 5 (2011), s. 837-848 ISSN 0898-6568 R&D Projects: GA ČR(CZ) GA204/07/1567; GA MŠk 1M0506 Institutional research plan: CEZ:AV0Z50520514 Keywords : Wnt signaling * post-translational modification * acylation Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 4.058, year: 2011

  5. Regulation of p53 by reversible post-transcriptional and post-translational mechanisms in liver and skeletal muscle of an anoxia tolerant turtle, Trachemys scripta elegans.

    Science.gov (United States)

    Zhang, Jing; Biggar, Kyle K; Storey, Kenneth B

    2013-01-15

    The red-eared slider turtle (Trachemys scripta elegans) exhibits well-developed natural anoxia tolerance that depends on multiple biochemical adaptations, including anoxia-induced hypometabolism. We hypothesized that signaling by the p53 protein could aid in establishing the hypometabolic state by arresting the cell cycle, protecting against DNA damage as well as altering pathways of energy metabolism. Immunoblotting was used to evaluate the regulation and post-transcriptional modifications of p53 in liver and skeletal muscle of red-eared slider turtles subjected to 5h or 20h of anoxic submergence. Tissue specific regulation of p53 was observed with the liver showing a more rapid activation of p53 in response to anoxia as well as differential expression of seven serine phosphorylation and two lysine acetylation sites when compared with skeletal muscle. Protein expression of MDM2, a major p53 inhibitor, was also examined but did not change during anoxia. Reverse-transcriptase PCR was used to assess transcript levels of selected p53 target genes (14-3-3σ, Gadd45α and Pgm) and one microRNA (miR-34a); results showed down-regulation of Pgm and up-regulation of the other three. These findings show an activation of p53 in response to anoxia exposure and suggest an important role for the p53 stress response pathway in regulating natural anoxia tolerance and hypometabolism in a vertebrate facultative anaerobe. Copyright © 2012 Elsevier B.V. All rights reserved.

  6. Grainsize Patterns and Bed Evolution of the Rhone River (France): A Present-day Snapshot Following a Century and a Half of Human Modifications

    Science.gov (United States)

    Michal, T.; Parrot, E.; Piegay, H.

    2014-12-01

    Over the past 150 years the Rhône River has been heavily altered by human infrastructures. The first wave (1860 - 1930) of modifications consisted of dikes and groynes designed to narrow the channel and promote incision in order to facilitate navigation. A second period (1948 - 1986) involved the construction of a series of canals and dams for hydroelectricity production. These works bypass multiple reaches of the original channel and drastically reduce the discharge and sediment load reaching them. A comprehensive study underway is aimed at describing the present-day morphology of the Rhone along its 512 km length from its source at Lake Geneva to its sink at the Mediterranean Sea and quantifying the role of management works in the evolution to its current state. Grainsize distributions and armour ratios were determined using a combination of Wolman counts on bars and in shallow channels and dredge samples collected from a boat in navigable reaches. Long profiles were constructed from historical bathymetric maps and bathymetric data collected between 1950 - 2010. Differential long profiles highlighting changes in bed elevation due to sediment storage and erosion were analyzed for three different periods: post-channelization, post-dam construction, and a recent period of major floods. Results show a complex discontinuous pattern in grainsize associated with hydraulic discontinuities imposed by dams. The D50 for bypass reaches is 45 mm compared to a D50 of 34 mm in the non-bypass reaches. The lower D50 as well as a finer tailed distribution in non-bypass reaches reflects fining associated with storage upstream of dams. Armour ratios are on average around 2 but are notably higher for reaches in the middle section of the Rhone. The average incision rate was 1.8 cm/yr for the period of post-channelization and 1.2 cm/yr following dam construction, suggesting the post-dam Rhone was already partially armoured due to incision associated with channelization preceding dam

  7. Distinct ontogenic patterns of overt and latent DGAT activities of rat liver microsomes.

    Science.gov (United States)

    Waterman, Ian J; Price, Nigel T; Zammit, Victor A

    2002-09-01

    We have studied the ontogeny of the two functional diacylglycerol acyltransferase (DGAT) activities (overt and latent) during postnatal development in rat liver. We find that the ontogenic patterns of the two are highly distinct. Overt DGAT shows a transient rise in activity up to day 4 postnatally, after which it declines until weaning; thereafter, it increases steadily to reach high adult values that may contribute to the high rates of turnover of cytosolic triacylglycerol (TAG). By contrast, latent DGAT activity increases continuously during the suckling period but falls sharply upon weaning onto chow but not onto a high-fat diet. Rates of TAG secretion by hepatocytes are higher than in the adult during the first 7 days after birth, and are largely dependent on the mobilization of the abundant intrahepatocyte TAG as a source of acyl moieties. When the hepatic steatosis is cleared (after day 7) the TAG secretion rate declines by 80% to reach adult values. Quantification of the content of mRNA for the DGAT1 and DGAT2 genes does not show correlation with either of the DGAT activities. We conclude that post-translational modification may play an important role in the overt and latent distribution of DGAT activity in the liver microsomal membrane.

  8. Fast Multi-blind Modification Search through Tandem Mass Spectrometry*

    Science.gov (United States)

    Na, Seungjin; Bandeira, Nuno; Paek, Eunok

    2012-01-01

    With great biological interest in post-translational modifications (PTMs), various approaches have been introduced to identify PTMs using MS/MS. Recent developments for PTM identification have focused on an unrestrictive approach that searches MS/MS spectra for all known and possibly even unknown types of PTMs at once. However, the resulting expanded search space requires much longer search time and also increases the number of false positives (incorrect identifications) and false negatives (missed true identifications), thus creating a bottleneck in high throughput analysis. Here we introduce MODa, a novel “multi-blind” spectral alignment algorithm that allows for fast unrestrictive PTM searches with no limitation on the number of modifications per peptide while featuring over an order of magnitude speedup in relation to existing approaches. We demonstrate the sensitivity of MODa on human shotgun proteomics data where it reveals multiple mutations, a wide range of modifications (including glycosylation), and evidence for several putative novel modifications. Based on the reported findings, we argue that the efficiency and sensitivity of MODa make it the first unrestrictive search tool with the potential to fully replace conventional restrictive identification of proteomics mass spectrometry data. PMID:22186716

  9. Histone modifications influence mediator interactions with chromatin

    Science.gov (United States)

    Zhu, Xuefeng; Zhang, Yongqiang; Bjornsdottir, Gudrun; Liu, Zhongle; Quan, Amy; Costanzo, Michael; Dávila López, Marcela; Westholm, Jakub Orzechowski; Ronne, Hans; Boone, Charles; Gustafsson, Claes M.; Myers, Lawrence C.

    2011-01-01

    The Mediator complex transmits activation signals from DNA bound transcription factors to the core transcription machinery. Genome wide localization studies have demonstrated that Mediator occupancy not only correlates with high levels of transcription, but that the complex also is present at transcriptionally silenced locations. We provide evidence that Mediator localization is guided by an interaction with histone tails, and that this interaction is regulated by their post-translational modifications. A quantitative, high-density genetic interaction map revealed links between Mediator components and factors affecting chromatin structure, especially histone deacetylases. Peptide binding assays demonstrated that pure wild-type Mediator forms stable complexes with the tails of Histone H3 and H4. These binding assays also showed Mediator—histone H4 peptide interactions are specifically inhibited by acetylation of the histone H4 lysine 16, a residue critical in transcriptional silencing. Finally, these findings were validated by tiling array analysis that revealed a broad correlation between Mediator and nucleosome occupancy in vivo, but a negative correlation between Mediator and nucleosomes acetylated at histone H4 lysine 16. Our studies show that chromatin structure and the acetylation state of histones are intimately connected to Mediator localization. PMID:21742760

  10. The effect of surface modifications of carbon nanotubes on the electrical properties of inkjet-printed SWNT/PEDOT-PSS composite line patterns

    International Nuclear Information System (INIS)

    Najeeb, C K; Lee, Jae-Hyeok; Chang, Jingbo; Kim, Jae-Ho

    2010-01-01

    We prepared nanocomposite inks of poly(3,4-ethylenedioxythiophene)-poly(styrenesulfonate) (PEDOT-PSS) filled with single-walled carbon nanotubes (SWNTs) purified by acidic treatment, carboxylated by chemical oxidation and carboxyl-functionalized nanotubes physically modified with a natural gum, gum arabic. Inkjet printing of line patterns with a feature size of 100 μm width and lengths ranging from 1 to 5 cm was performed on glass substrates with a piezoelectric inkjet printer. The carboxyl-functionalized SWNT-based composite demonstrated a significant decrease (fourfold) of electrical resistance for the line patterns compared to that with a purified CNT-based composite due to improved dispersability of nanotubes in the polymer matrix. The use of gum arabic for the dispersion of carboxyl-functionalized nanotubes demonstrated a further drastic decrease (18-fold) of the resistance compared with a purified CNT-based composite owing to the formation of an extended continuous network within the line pattern. The inkjet-printed conductive patterns can be applied in various fields, such as flexible high speed transistors, high efficiency solar cells and transparent electrodes.

  11. Microtubule proteins and their post-translational forms in the cerebrospinal fluid of patients with paraparesis associated with HTLV-I infection and in SH-SY5Y cells: An in vitro model of HTLV-I-induced disease

    Directory of Open Access Journals (Sweden)

    HORACIO MALDONADO

    2008-01-01

    Full Text Available HTLV-I-associated myelopathy/tropical spastic paraparesis (HAM/TSP is characterized by axonal degeneration of the corticospinal tracts. The specific requirements for transport of proteins and organelles to the distal part of the long axon are crucial in the corticospinal tracts. Microtubule dysfunction could be involved in this disease, configuring an axonal transport disease. We measured tubulin and its post-translational modified forms (acetylated and tyrosinated in CSF of patients and controls, as well as tau and its phosphorylated forms. There were no significant differences in the contents of tubulin and acetyl-tubulin between patients and controls; tyrosyl-tubulin was not detected. In HAM/TSP, tau levéis were significantly reduced, while the ratio of pT181/total tau was higher in patients than in controls, this being completely different from what is reported in other neurodegenerative diseases. Phosphorylation at T181 was also confirmed by Mass Spectrometry analysis. Western Blotting with monospecific polyclonal antibodies against pS199, pT205, pT231, pS262, pS356, pS396, pS404 and pS422 did not show differences in phosphorylation in these residues between patients and controls. Treating human SH-SY5Y neuroblastoma cells, a well-known in vitro neurite retraction model, with culture supernatant of MT-2 cells (HTLV-I infected cell line that secretes the viral Tax protein we observed neurite retraction and an increase in tau phosphorylation at T181. A disruption of normal phosphorylation of tau protein in T181 could result in its dysfunction, contributing to axonal damage.

  12. Modification site localization scoring integrated into a search engine.

    Science.gov (United States)

    Baker, Peter R; Trinidad, Jonathan C; Chalkley, Robert J

    2011-07-01

    Large proteomic data sets identifying hundreds or thousands of modified peptides are becoming increasingly common in the literature. Several methods for assessing the reliability of peptide identifications both at the individual peptide or data set level have become established. However, tools for measuring the confidence of modification site assignments are sparse and are not often employed. A few tools for estimating phosphorylation site assignment reliabilities have been developed, but these are not integral to a search engine, so require a particular search engine output for a second step of processing. They may also require use of a particular fragmentation method and are mostly only applicable for phosphorylation analysis, rather than post-translational modifications analysis in general. In this study, we present the performance of site assignment scoring that is directly integrated into the search engine Protein Prospector, which allows site assignment reliability to be automatically reported for all modifications present in an identified peptide. It clearly indicates when a site assignment is ambiguous (and if so, between which residues), and reports an assignment score that can be translated into a reliability measure for individual site assignments.

  13. Cell patterning without chemical surface modification: Cell cell interactions between printed bovine aortic endothelial cells (BAEC) on a homogeneous cell-adherent hydrogel

    Science.gov (United States)

    Chen, C. Y.; Barron, J. A.; Ringeisen, B. R.

    2006-10-01

    Cell printing offers the unique ability to directly deposit one or multiple cell types directly onto a surface without the need to chemically pre-treat the surface with lithographic methods. We utilize biological laser printing (BioLP ™) to form patterns of bovine aortic endothelial cells (BAECs) onto a homogeneous cell adherent hydrogel surface. These normal cells are shown to retain near-100% viability post-printing. In order to determine whether BAECs encountered shear and/or heat stress during printing, immunocytochemical staining experiments were performed to detect potential expression of heat shock proteins (HSP) by the deposited cells. Printed BAECs expressed HSP at levels similar to negative control cells, indicating that the BioLP process does not expose cells to damaging levels of stress. However, HSP expression was slightly higher at the highest laser energy studied, suggesting more stress was present under these extreme conditions. Printed BAECs also showed preferential asymmetric growth and migration towards each other and away from the originally printed pattern, demonstrating a retained ability for the cells to communicate post-printing.

  14. Low-Fat Dietary Pattern Intervention and Health-Related Quality of Life: The Women's Health Initiative Randomized Controlled Dietary Modification Trial.

    Science.gov (United States)

    Assaf, Annlouise R; Beresford, Shirley A A; Risica, Patricia Markham; Aragaki, Aaron; Brunner, Robert L; Bowen, Deborah J; Naughton, Michelle; Rosal, Milagros C; Snetselaar, Linda; Wenger, Nanette

    2016-02-01

    Intensive dietary intervention programs may lead to benefits in vitality and other components of health quality. The Women's Health Initiative Dietary Modification (DM) intervention includes a large randomized controlled trial of an intensive intervention. To evaluate whether the intervention is associated with improved health-related quality of life (HRQoL) subscales, overall self-reported health, depression symptoms, cognitive functioning, and sleep quality. This randomized controlled trial was analyzed as intent to treat. Between 1993 and 1998, 48,835 women aged 50 to 79 years were recruited by 40 clinical centers across the United States. Eligibility included having fat intake at baseline ≥32% of total calories, and excluded women with any prior colorectal or breast cancer, recent other cancers, type 1 diabetes, or medical conditions with predicted survival <3 years. Goals were to reduce calories from fat to 20%, increase vegetables and fruit to 5+ servings, and increase grain servings to 6+ servings a day. During the first year, 18 group sessions were held, with quarterly sessions thereafter. The RAND 36-Item Health Survey was used to assess HRQoL at baseline, Year 1, and close-out (about 8 years postrandomization), and estimate differential HRQoL subscale change scores. Mean change in HRQoL scores (Year 1 minus baseline) were compared by randomization group using linear models. At 1 year, there was a differential change between intervention and comparison group of 1.7 units (95% CI 1.5, 2.0) in general health associated with the intervention. DM intervention improved physical functioning by 2.0 units (95% CI 1.7, 2.3), vitality by 1.9 units (95% CI 1.6, 2.2), and global quality of life by 0.09 units (95% CI 0.07, 0.12). With the exception of global quality of life, these effects were significantly modified by body mass index at baseline. DM intervention was associated with small, but significant improvements in three HRQoL subscales: general health

  15. PLMD: An updated data resource of protein lysine modifications.

    Science.gov (United States)

    Xu, Haodong; Zhou, Jiaqi; Lin, Shaofeng; Deng, Wankun; Zhang, Ying; Xue, Yu

    2017-05-20

    Post-translational modifications (PTMs) occurring at protein lysine residues, or protein lysine modifications (PLMs), play critical roles in regulating biological processes. Due to the explosive expansion of the amount of PLM substrates and the discovery of novel PLM types, here we greatly updated our previous studies, and presented a much more integrative resource of protein lysine modification database (PLMD). In PLMD, we totally collected and integrated 284,780 modification events in 53,501 proteins across 176 eukaryotes and prokaryotes for up to 20 types of PLMs, including ubiquitination, acetylation, sumoylation, methylation, succinylation, malonylation, glutarylation, glycation, formylation, hydroxylation, butyrylation, propionylation, crotonylation, pupylation, neddylation, 2-hydroxyisobutyrylation, phosphoglycerylation, carboxylation, lipoylation and biotinylation. Using the data set, a motif-based analysis was performed for each PLM type, and the results demonstrated that different PLM types preferentially recognize distinct sequence motifs for the modifications. Moreover, various PLMs synergistically orchestrate specific cellular biological processes by mutual crosstalks with each other, and we totally found 65,297 PLM events involved in 90 types of PLM co-occurrences on the same lysine residues. Finally, various options were provided for accessing the data, while original references and other annotations were also present for each PLM substrate. Taken together, we anticipated the PLMD database can serve as a useful resource for further researches of PLMs. PLMD 3.0 was implemented in PHP + MySQL and freely available at http://plmd.biocuckoo.org. Copyright © 2017 Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, and Genetics Society of China. Published by Elsevier Ltd. All rights reserved.

  16. Modification of the trapped field in bulk high-temperature superconductors as a result of the drilling of a pattern of artificial columnar holes

    International Nuclear Information System (INIS)

    Lousberg, Gregory P; Vanderbemden, Ph; Vanderheyden, B; Fagnard, J-F; Ausloos, M

    2010-01-01

    The trapped magnetic field is examined in bulk high-temperature superconductors that are artificially drilled along their c-axis. The influence of the hole pattern on the magnetization is studied and compared by means of numerical models and Hall probe mapping techniques. To this aim, we consider two bulk YBCO samples with a rectangular cross-section that are drilled each by six holes arranged either on a rectangular lattice (sample I) or on a centered rectangular lattice (sample II). For the numerical analysis, three different models are considered for calculating the trapped flux: (i), a two-dimensional (2D) Bean model neglecting demagnetizing effects and flux creep, (ii), a 2D finite-element model neglecting demagnetizing effects but incorporating magnetic relaxation in the form of an E - J power law, and, (iii), a 3D finite element analysis that takes into account both the finite height of the sample and flux creep effects. For the experimental analysis, the trapped magnetic flux density is measured above the sample surface by Hall probe mapping performed before and after the drilling process. The maximum trapped flux density in the drilled samples is found to be smaller than that in the plain samples. The smallest magnetization drop is found for sample II, with the centered rectangular lattice. This result is confirmed by the numerical models. In each sample, the relative drops that are calculated independently with the three different models are in good agreement. As observed experimentally, the magnetization drop calculated in the sample II is the smallest one and its relative value is comparable to the measured one. By contrast, the measured magnetization drop in sample (1) is much larger than that predicted by the simulations, most likely because of a change of the microstructure during the drilling process.

  17. Characterization and stability of transthyretin isoforms in cerebrospinal fluid examined by immunoprecipitation and high-resolution mass spectrometry of intact protein

    DEFF Research Database (Denmark)

    Poulsen, Keld; Bahl, Justyna M C; Tanassi, Julia T

    2012-01-01

    Post-translational modifications (PTMs) contribute significantly to the complexity of proteins. PTMs may vary in certain patterns according to diseases and microenviroments making them potential markers for pathological processes. Human transthyretin (TTR) is a transporter of thyroxine and retino...

  18. Effect of the UV modification of α-crystallin on its ability to suppress nonspecific aggregation

    International Nuclear Information System (INIS)

    Ellozy, A.R.; Ceger, Patricia; Wang, R.H.; Dillon, James

    1996-01-01

    Recent studies have shown that structural modifications of α-crystallin during lens aging decrease it's effectiveness as a molecular chaperone. Some of these post-translational modifications have been linked to UV radiation, and this study was undertaken to investigate the effect of UV irradiation on the ability of α-crystallin to suppress nonspecific aggregation. The effect of 3-hydroxykynurenine (3-HK) was also investigated as a model for its glucoside (3-HKG), a main lens chromophore that has been linked to photochemical changes in the human lens. Alpha- and γ-crystallin solutions (1 mg/mL, 1:0.125 wt/wt) were photolyzed (transmission above 295nm) for various time intervals. Thermal denaturation of γ-crystallin with or without α-crystallin was carried out at 70 o C and increases in light scattering were measured at 360 nm. We found that (1) irradiation of γ-crystallin increased its susceptibility to heat-induced scattering. The addition of α-crystallin protects it against thermal denaturation, although its ability to do so decreases the longer γ-crystallin is irradiated and (2) irradiation of α-crystallin decreases its ability to suppress nonspecific aggregation and the presence of 3-HK during irradiation decreases its further. Our results indicate that post-translational modifications of α-crystallin due to UV irradiation affect the sites and mechanisms by which it interacts with γ-crystallin. The kinetics of γ-crystallin unfolding during thermal denaturation were also analyzed. We found that a simple two state model applied for nonirradiated γ-crystallin. This model does not hold when γ-crystallin is irradiated in the prescence or absence of α-crystallin. In these cases, two step or multistep mechanisms are more likely. (Author)

  19. Chromatin modifications and the DNA damage response to ionizing radiation

    International Nuclear Information System (INIS)

    Kumar, Rakesh; Horikoshi, Nobuo; Singh, Mayank; Gupta, Arun; Misra, Hari S.; Albuquerque, Kevin; Hunt, Clayton R.; Pandita, Tej K.

    2013-01-01

    In order to survive, cells have evolved highly effective repair mechanisms to deal with the potentially lethal DNA damage produced by exposure to endogenous as well as exogenous agents. Ionizing radiation exposure induces highly lethal DNA damage, especially DNA double-strand breaks (DSBs), that is sensed by the cellular machinery and then subsequently repaired by either of two different DSB repair mechanisms: (1) non-homologous end joining, which re-ligates the broken ends of the DNA and (2) homologous recombination, that employs an undamaged identical DNA sequence as a template, to maintain the fidelity of DNA repair. Repair of DSBs must occur within the natural context of the cellular DNA which, along with specific proteins, is organized to form chromatin, the overall structure of which can impede DNA damage site access by repair proteins. The chromatin complex is a dynamic structure and is known to change as required for ongoing cellular processes such as gene transcription or DNA replication. Similarly, during the process of DNA damage sensing and repair, chromatin needs to undergo several changes in order to facilitate accessibility of the repair machinery. Cells utilize several factors to modify the chromatin in order to locally open up the structure to reveal the underlying DNA sequence but post-translational modification of the histone components is one of the primary mechanisms. In this review, we will summarize chromatin modifications by the respective chromatin modifying factors that occur during the DNA damage response.

  20. Discovery of a Chemical Modification by Citric Acid in a Recombinant Monoclonal Antibody

    Science.gov (United States)

    2015-01-01

    Recombinant therapeutic monoclonal antibodies exhibit a high degree of heterogeneity that can arise from various post-translational modifications. The formulation for a protein product is to maintain a specific pH and to minimize further modifications. Generally Recognized as Safe (GRAS), citric acid is commonly used for formulation to maintain a pH at a range between 3 and 6 and is generally considered chemically inert. However, as we reported herein, citric acid covalently modified a recombinant monoclonal antibody (IgG1) in a phosphate/citrate-buffered formulation at pH 5.2 and led to the formation of so-called “acidic species” that showed mass increases of 174 and 156 Da, respectively. Peptide mapping revealed that the modification occurred at the N-terminus of the light chain. Three additional antibodies also showed the same modification but displayed different susceptibilities of the N-termini of the light chain, heavy chain, or both. Thus, ostensibly unreactive excipients under certain conditions may increase heterogeneity and acidic species in formulated recombinant monoclonal antibodies. By analogy, other molecules (e.g., succinic acid) with two or more carboxylic acid groups and capable of forming an anhydride may exhibit similar reactivities. Altogether, our findings again reminded us that it is prudent to consider formulations as a potential source for chemical modifications and product heterogeneity. PMID:25136741

  1. Evaluation of Proteomic Search Engines for the Analysis of Histone Modifications

    Science.gov (United States)

    2015-01-01

    Identification of histone post-translational modifications (PTMs) is challenging for proteomics search engines. Including many histone PTMs in one search increases the number of candidate peptides dramatically, leading to low search speed and fewer identified spectra. To evaluate database search engines on identifying histone PTMs, we present a method in which one kind of modification is searched each time, for example, unmodified, individually modified, and multimodified, each search result is filtered with false discovery rate less than 1%, and the identifications of multiple search engines are combined to obtain confident results. We apply this method for eight search engines on histone data sets. We find that two search engines, pFind and Mascot, identify most of the confident results at a reasonable speed, so we recommend using them to identify histone modifications. During the evaluation, we also find some important aspects for the analysis of histone modifications. Our evaluation of different search engines on identifying histone modifications will hopefully help those who are hoping to enter the histone proteomics field. The mass spectrometry proteomics data have been deposited to the ProteomeXchange Consortium with the data set identifier PXD001118. PMID:25167464

  2. Evaluation of proteomic search engines for the analysis of histone modifications.

    Science.gov (United States)

    Yuan, Zuo-Fei; Lin, Shu; Molden, Rosalynn C; Garcia, Benjamin A

    2014-10-03

    Identification of histone post-translational modifications (PTMs) is challenging for proteomics search engines. Including many histone PTMs in one search increases the number of candidate peptides dramatically, leading to low search speed and fewer identified spectra. To evaluate database search engines on identifying histone PTMs, we present a method in which one kind of modification is searched each time, for example, unmodified, individually modified, and multimodified, each search result is filtered with false discovery rate less than 1%, and the identifications of multiple search engines are combined to obtain confident results. We apply this method for eight search engines on histone data sets. We find that two search engines, pFind and Mascot, identify most of the confident results at a reasonable speed, so we recommend using them to identify histone modifications. During the evaluation, we also find some important aspects for the analysis of histone modifications. Our evaluation of different search engines on identifying histone modifications will hopefully help those who are hoping to enter the histone proteomics field. The mass spectrometry proteomics data have been deposited to the ProteomeXchange Consortium with the data set identifier PXD001118.

  3. O-GlcNAc modification: why so intimately associated with phosphorylation?

    Directory of Open Access Journals (Sweden)

    Ande Sudharsana R

    2011-01-01

    Full Text Available Abstract Post-translational modification of proteins at serine and threonine side chains by β-N-acetylglucosamine (O-GlcNAc mediated by the enzyme β-N-acetylglucosamine transferase has been emerging as a fundamental regulatory mechanism encompassing a wide range of proteins involved in cell division, metabolism, transcription and cell signaling. Furthermore, an extensive interplay between O-GlcNAc modification and serine/threonine phosphorylation in a variety of proteins has been reported to exist. However, our understanding of the regulatory mechanisms involved in O-GlcNAc modification and its interplay with serine/threonine phosphorylation in proteins is still elusive. Recent success in the mapping of O-GlcNAc modification sites in proteins as a result of technological advancement in mass spectrometry have revealed two important clues which may be inherently connected to the regulation of O-GlcNAc modification and its interplay with phosphorylation in proteins. First, almost all O-GlcNAc modified proteins are known phospho proteins. Second, the prevalence of tyrosine phosphorylation among O-GlcNAc modified proteins is exceptionally higher (~68% than its normal occurrence (~2% alone. We hypothesize that phosphorylation may be a requisite for O-GlcNAc modification and tyrosine phosphorylation plays a role in the interplay between O-GlcNAc modification and serine/threonine phosphorylation in proteins. In other words, the interplay between O-GlcNAc modification and phosphorylation is not limited to serine/threonine phosphorylation but also includes tyrosine phosphorylation. Our hypothesis provides an opportunity to understand the underlying mechanism involved in O-GlcNAc modification and its interplay with serine/threonine phosphorylation in proteins. Furthermore, implication of our hypothesis extends to tyrosine kinase signaling.

  4. Characterizing Patterns of MILCON Project Contract Modifications

    Science.gov (United States)

    2007-12-01

    overtime and over manning), the amount of change, absenteeism and turnover, manpower increase over planned levels, and the processing time from the...S., & Vandenberg, P. J. (1999). The impact of change orders on mechanical construction labour efficiency. Construction Management & Economics, 17(6

  5. Drug Addiction and DNA Modifications.

    Science.gov (United States)

    Brown, Amber N; Feng, Jian

    2017-01-01

    Drug addiction is a complex disorder which can be influenced by both genetic and environmental factors. Research has shown that epigenetic modifications can translate environmental signals into changes in gene expression, suggesting that epigenetic changes may underlie the causes and possibly treatment of substance use disorders. This chapter will focus on epigenetic modifications to DNA, which include DNA methylation and several recently defined additional DNA epigenetic changes. We will discuss the functions of DNA modifications and methods for detecting them, followed by a description of the research investigating the function and consequences of drug-induced changes in DNA methylation patterns. Understanding these epigenetic changes may provide us translational tools for the diagnosis and treatment of addiction in the future.

  6. The Impact of Commonly Used Alkylating Agents on Artifactual Peptide Modification.

    Science.gov (United States)

    Hains, Peter G; Robinson, Phillip J

    2017-09-01

    Iodoacetamide is by far the most commonly used agent for alkylation of cysteine during sample preparation for proteomics. An alternative, 2-chloroacetamide, has recently been suggested to reduce the alkylation of residues other than cysteine, such as the N-terminus, Asp, Glu, Lys, Ser, Thr, and Tyr. Here we show that although 2-chloroacetamide reduces the level of off-target alkylation, it exhibits a range of adverse effects. The most significant of these is methionine oxidation, which increases to a maximum of 40% of all Met-containing peptides, compared with 2-5% with iodoacetamide. Increases were also observed for mono- and dioxidized tryptophan. No additional differences between the alkylating reagents were observed for a range of other post-translational modifications and digestion parameters. The deleterious effects were observed for 2-chloroacetamide from three separate suppliers. The adverse impact of 2-chloroacetamide on methionine oxidation suggests that it is not the ideal alkylating reagent for proteomics.

  7. Structure predictions of two Bauhinia variegata lectins reveal patterns of C-terminal properties in single chain legume lectins.

    Science.gov (United States)

    Moreira, Gustavo M S G; Conceição, Fabricio R; McBride, Alan J A; Pinto, Luciano da S

    2013-01-01

    Bauhinia variegata lectins (BVL-I and BVL-II) are single chain lectins isolated from the plant Bauhinia variegata. Single chain lectins undergo post-translational processing on its N-terminal and C-terminal regions, which determines their physiological targeting, carbohydrate binding activity and pattern of quaternary association. These two lectins are isoforms, BVL-I being highly glycosylated, and thus far, it has not been possible to determine their structures. The present study used prediction and validation algorithms to elucidate the likely structures of BVL-I and -II. The program Bhageerath-H was chosen from among three different structure prediction programs due to its better overall reliability. In order to predict the C-terminal region cleavage sites, other lectins known to have this modification were analysed and three rules were created: (1) the first amino acid of the excised peptide is small or hydrophobic; (2) the cleavage occurs after an acid, polar, or hydrophobic residue, but not after a basic one; and (3) the cleavage spot is located 5-8 residues after a conserved Leu amino acid. These rules predicted that BVL-I and -II would have fifteen C-terminal residues cleaved, and this was confirmed experimentally by Edman degradation sequencing of BVL-I. Furthermore, the C-terminal analyses predicted that only BVL-II underwent α-helical folding in this region, similar to that seen in SBA and DBL. Conversely, BVL-I and -II contained four conserved regions of a GS-I association, providing evidence of a previously undescribed X4+unusual oligomerisation between the truncated BVL-I and the intact BVL-II. This is the first report on the structural analysis of lectins from Bauhinia spp. and therefore is important for the characterisation C-terminal cleavage and patterns of quaternary association of single chain lectins.

  8. Differential N-glycan patterns identified in lung adenocarcinoma by N-glycan profiling of formalin-fixed paraffin-embedded (FFPE) tissue sections.

    Science.gov (United States)

    Wang, Xiaoning; Deng, Zaian; Huang, Chuncui; Zhu, Tong; Lou, Jiatao; Wang, Lin; Li, Yan

    2018-02-10

    N-glycan profiling is a powerful approach for analyzing the functional relationship between N-glycosylation and cancer. Current methods rely on either serum or fresh tissue samples; however, N-glycan patterns may differ between serum and tissue, as the proteins of serum originate from a variety of tissues. Furthermore, fresh tissue samples are difficult to ship and store. Here, we used a profiling method based on formalin-fixed paraffin-embedded (FFPE) tissue sections from lung adenocarcinoma patients. We found that our method was highly reproducible. We identified 58 N-glycan compositions from lung adenocarcinoma FFPE samples, 51 of which were further used for MS n -based structure prediction. We show that high mannose type N-glycans are upregulated, while sialylated N-glycans are downregulated in our FFPE lung adenocarcinoma samples, compared to the control samples. Our receiver operating characteristic (ROC) curve analysis shows that high mannose type and sialylated N-glycans are useful discriminators to distinguish between lung adenocarcinoma and control tissue. Together, our results indicate that expression levels of specific N-glycans correlate well with lung adenocarcinoma, and strongly suggest that our FFPE-based method will be useful for N-glycan profiling of cancer tissues. Glycosylation is one of the most important post-translational protein modifications, and is associated with several physiopathological processes, including carcinogenesis. In this study, we tested the feasibility of using formalin-fixed paraffin-embedded (FFPE) tissue sections to identify changes in N-glycan patterns and identified the differentially expressed N-glycans of lung adenocarcinoma. Our study shows that the FFPE-based N-glycan profiling method is useful for clinical diagnosis as well as identification of potential biomarkers, and our data expand current knowledge of differential N-glycan patterns of lung adenocarcinoma. Copyright © 2017 Elsevier B.V. All rights reserved.

  9. Bioorthogonal Chemistry for the Isolation and Study of Newly Synthesized Histones and Their Modifications.

    Science.gov (United States)

    Arnaudo, Anna M; Link, A James; Garcia, Benjamin A

    2016-03-18

    The nucleosome is an octamer containing DNA wrapped around one histone H3-H4 tetramer and two histone H2A-H2B dimers. Within the nucleosome, histones are decorated with post-translational modifications. Previous studies indicate that the H3-H4 tetramer is conserved during DNA replication, suggesting that old tetramers serve as a template for the modification of newly synthesized tetramers. Here, we present a method that merges bioorthogonal chemistry with mass spectrometry for the study of modifications on newly synthesized histones in mammalian cells. HeLa S3 cells are dually labeled with the methionine analog azidohomoalanine and heavy (13)C6,(15)N4 isotope labeled arginine. Heavy amino acid labeling marks newly synthesized histones while azidohomoalanine incorporation allows for their isolation using bioorthogonal ligation. Labeled mononucleosomes were covalently linked via a copper catalyzed reaction to a FLAG-GGR-alkyne peptide, immunoprecipitated, and subjected to mass spectrometry for quantitative modification analysis. Mononucleosomes containing new histones were successfully isolated using this approach. Additionally, the development of this method highlights the potential deleterious effects of azidohomoalanine labeling on protein PTMs and cell cycle progression, which should be considered for future studies utilizing bioorthogonal labeling strategies in mammalian cells.

  10. Site-selective protein-modification chemistry for basic biology and drug development.

    Science.gov (United States)

    Krall, Nikolaus; da Cruz, Filipa P; Boutureira, Omar; Bernardes, Gonçalo J L

    2016-02-01

    Nature has produced intricate machinery to covalently diversify the structure of proteins after their synthesis in the ribosome. In an attempt to mimic nature, chemists have developed a large set of reactions that enable post-expression modification of proteins at pre-determined sites. These reactions are now used to selectively install particular modifications on proteins for many biological and therapeutic applications. For example, they provide an opportunity to install post-translational modifications on proteins to determine their exact biological roles. Labelling of proteins in live cells with fluorescent dyes allows protein uptake and intracellular trafficking to be tracked and also enables physiological parameters to be measured optically. Through the conjugation of potent cytotoxicants to antibodies, novel anti-cancer drugs with improved efficacy and reduced side effects may be obtained. In this Perspective, we highlight the most exciting current and future applications of chemical site-selective protein modification and consider which hurdles still need to be overcome for more widespread use.

  11. Regulation of Wnt/β-catenin signaling by posttranslational modifications

    Science.gov (United States)

    2014-01-01

    The canonical Wnt signaling pathway (or Wnt/β-catenin pathway) plays a pivotal role in embryonic development and adult homeostasis; deregulation of the Wnt pathway contributes to the initiation and progression of human diseases including cancer. Despite its importance in human biology and disease, how regulation of the Wnt/β-catenin pathway is achieved remains largely undefined. Increasing evidence suggests that post-translational modifications (PTMs) of Wnt pathway components are essential for the activation of the Wnt/β-catenin pathway. PTMs create a highly dynamic relay system that responds to Wnt stimulation without requiring de novo protein synthesis and offer a platform for non-Wnt pathway components to be involved in the regulation of Wnt signaling, hence providing alternative opportunities for targeting the Wnt pathway. This review highlights the current status of PTM-mediated regulation of the Wnt/β-catenin pathway with a focus on factors involved in Wnt-mediated stabilization of β-catenin. PMID:24594309

  12. Overexpression of Catalase Diminishes Oxidative Cysteine Modifications of Cardiac Proteins.

    Directory of Open Access Journals (Sweden)

    Chunxiang Yao

    Full Text Available Reactive protein cysteine thiolates are instrumental in redox regulation. Oxidants, such as hydrogen peroxide (H2O2, react with thiolates to form oxidative post-translational modifications, enabling physiological redox signaling. Cardiac disease and aging are associated with oxidative stress which can impair redox signaling by altering essential cysteine thiolates. We previously found that cardiac-specific overexpression of catalase (Cat, an enzyme that detoxifies excess H2O2, protected from oxidative stress and delayed cardiac aging in mice. Using redox proteomics and systems biology, we sought to identify the cysteines that could play a key role in cardiac disease and aging. With a 'Tandem Mass Tag' (TMT labeling strategy and mass spectrometry, we investigated differential reversible cysteine oxidation in the cardiac proteome of wild type and Cat transgenic (Tg mice. Reversible cysteine oxidation was measured as thiol occupancy, the ratio of total available versus reversibly oxidized cysteine thiols. Catalase overexpression globally decreased thiol occupancy by ≥1.3 fold in 82 proteins, including numerous mitochondrial and contractile proteins. Systems biology analysis assigned the majority of proteins with differentially modified thiols in Cat Tg mice to pathways of aging and cardiac disease, including cellular stress response, proteostasis, and apoptosis. In addition, Cat Tg mice exhibited diminished protein glutathione adducts and decreased H2O2 production from mitochondrial complex I and II, suggesting improved function of cardiac mitochondria. In conclusion, our data suggest that catalase may alleviate cardiac disease and aging by moderating global protein cysteine thiol oxidation.

  13. Methylglyoxal-induced modification causes aggregation of myoglobin

    Science.gov (United States)

    Banerjee, Sauradipta; Maity, Subhajit; Chakraborti, Abhay Sankar

    2016-02-01

    Post-translational modification of proteins by Maillard reaction, known as glycation, is thought to be the root cause of different complications, particularly in diabetes mellitus and age-related disorders. Methylglyoxal (MG), a reactive α-oxoaldehyde, increases in diabetic condition and reacts with proteins to form advanced glycation end products (AGEs) following Maillard-like reaction. We have investigated the in vitro effect of MG (200 μM) on the monomeric heme protein myoglobin (Mb) (100 μM) in a time-dependent manner (7 to 18 days incubation at 25 °C). MG induces significant structural alterations of the heme protein, including heme loss, changes in tryptophan fluorescence, decrease of α-helicity with increased β-sheet content etc. These changes occur gradually with increased period of incubation. Incubation of Mb with MG for 7 days results in formation of the AGE adducts: carboxyethyllysine at Lys-16, carboxymethyllysine at Lys-87 and carboxyethyllysine or pyrraline-carboxymethyllysine at Lys-133. On increasing the period of incubation up to 14 days, additional AGEs namely, carboxyethyllysine at Lys-42 and hydroimidazolone or argpyrimidine at Arg-31 and Arg-139 have been detected. MG also induces aggregation of Mb, which is clearly evident with longer period of incubation (18 days), and appears to have amyloid nature. MG-derived AGEs may thus have an important role as the precursors of protein aggregation, which, in turn, may be associated with physiological complications.

  14. Behavior Modification in Coaching.

    Science.gov (United States)

    Lynch, Annette Rutt; Stillman, Stephen M.

    1979-01-01

    An example of behavior modification used in athletic coaching is presented. The case study involves a member of a women's basketball team and details the use of behavior modification for both weight reduction and skill improvement. (JMF)

  15. Epigenetic Modifications: Therapeutic Potential in Cancer

    Directory of Open Access Journals (Sweden)

    Manisha Sachan

    2015-08-01

    Full Text Available Epigenetic modifications and alterations in chromatin structure and function contribute to the cumulative changes observed as normal cells undergo malignant transformation. These modifications and enzymes (DNA methyltransferases, histone deacetylases, histone methyltransferases, and demethylases related to them have been deeply studied to develop new drugs, epigenome-targeted therapies and new diagnostic tools. Epigenetic modifiers aim to restore normal epigenetic modification patterns through the inhibition of epigenetic modifier enzymes. Four of them (azacitidine, decitabine, vorinostat and romidepsin are approved by the U.S. Food and Drug Administration. This article provides an overview about the known functional roles of epigenetic enzymes in cancer development.

  16. Disruption of histone modification and CARM1 recruitment by arsenic represses transcription at glucocorticoid receptor-regulated promoters.

    Science.gov (United States)

    Barr, Fiona D; Krohmer, Lori J; Hamilton, Joshua W; Sheldon, Lynn A

    2009-08-26

    Chronic exposure to inorganic arsenic (iAs) found in the environment is one of the most significant and widespread environmental health risks in the U.S. and throughout the world. It is associated with a broad range of health effects from cancer to diabetes as well as reproductive and developmental anomalies. This diversity of diseases can also result from disruption of metabolic and other cellular processes regulated by steroid hormone receptors via aberrant transcriptional regulation. Significantly, exposure to iAs inhibits steroid hormone-mediated gene activation. iAs exposure is associated with disease, but is also used therapeutically to treat specific cancers complicating an understanding of iAs action. Transcriptional activation by steroid hormone receptors is accompanied by changes in histone and non-histone protein post-translational modification (PTM) that result from the enzymatic activity of coactivator and corepressor proteins such as GRIP1 and CARM1. This study addresses how iAs represses steroid receptor-regulated gene transcription. PTMs on histones H3 and H4 at the glucocorticoid receptor (GR)-activated mouse mammary tumor virus (MMTV) promoter were identified by chromatin immunoprecipitation analysis following exposure to steroid hormone+/-iAs. Histone H3K18 and H3R17 amino acid residues had significantly different patterns of PTMs after treatment with iAs. Promoter interaction of the coactivator CARM1 was disrupted, but the interaction of GRIP1, a p160 coactivator through which CARM1 interacts with a promoter, was intact. Over-expression of CARM1 was able to fully restore and GRIP1 partially restored iAs-repressed transcription indicating that these coactivators are functionally associated with iAs-mediated transcriptional repression. Both are essential for robust transcription at steroid hormone regulated genes and both are associated with disease when inappropriately expressed. We postulate that iAs effects on CARM1 and GRIP1 may underlie some

  17. N-terminal modifications of cellular proteins: The enzymes involved, their substrate specificities and biological effects

    Science.gov (United States)

    Varland, Sylvia; Osberg, Camilla; Arnesen, Thomas

    2015-01-01

    The vast majority of eukaryotic proteins are N-terminally modified by one or more processing enzymes. Enzymes acting on the very first amino acid of a polypeptide include different peptidases, transferases, and ligases. Methionine aminopeptidases excise the initiator methionine leaving the nascent polypeptide with a newly exposed amino acid that may be further modified. N-terminal acetyl-, methyl-, myristoyl-, and palmitoyltransferases may attach an acetyl, methyl, myristoyl, or palmitoyl group, respectively, to the α-amino group of the target protein N-terminus. With the action of ubiquitin ligases, one or several ubiquitin molecules are transferred, and hence, constitute the N-terminal modification. Modifications at protein N-termini represent an important contribution to proteomic diversity and complexity, and are essential for protein regulation and cellular signaling. Consequently, dysregulation of the N-terminal modifying enzymes is implicated in human diseases. We here review the different protein N-terminal modifications occurring co- or post-translationally with emphasis on the responsible enzymes and their substrate specificities. PMID:25914051

  18. Role of Transcription Factor Modifications in the Pathogenesis of Insulin Resistance

    Directory of Open Access Journals (Sweden)

    Mi-Young Kim

    2012-01-01

    Full Text Available Non-alcoholic fatty liver disease (NAFLD is characterized by fat accumulation in the liver not due to alcohol abuse. NAFLD is accompanied by variety of symptoms related to metabolic syndrome. Although the metabolic link between NAFLD and insulin resistance is not fully understood, it is clear that NAFLD is one of the main cause of insulin resistance. NAFLD is shown to affect the functions of other organs, including pancreas, adipose tissue, muscle and inflammatory systems. Currently efforts are being made to understand molecular mechanism of interrelationship between NAFLD and insulin resistance at the transcriptional level with specific focus on post-translational modification (PTM of transcription factors. PTM of transcription factors plays a key role in controlling numerous biological events, including cellular energy metabolism, cell-cycle progression, and organ development. Cell type- and tissue-specific reversible modifications include lysine acetylation, methylation, ubiquitination, and SUMOylation. Moreover, phosphorylation and O-GlcNAcylation on serine and threonine residues have been shown to affect protein stability, subcellular distribution, DNA-binding affinity, and transcriptional activity. PTMs of transcription factors involved in insulin-sensitive tissues confer specific adaptive mechanisms in response to internal or external stimuli. Our understanding of the interplay between these modifications and their effects on transcriptional regulation is growing. Here, we summarize the diverse roles of PTMs in insulin-sensitive tissues and their involvement in the pathogenesis of insulin resistance.

  19. Glutamine methylation in histone H2A is an RNA-polymerase-I-dedicated modification

    Science.gov (United States)

    Tessarz, Peter; Santos-Rosa, Helena; Robson, Sam C.; Sylvestersen, Kathrine B.; Nelson, Christopher J.; Nielsen, Michael L.; Kouzarides, Tony

    2014-01-01

    Nucleosomes are decorated with numerous post-translational modifications capable of influencing many DNA processes. Here we describe a new class of histone modification, methylation of glutamine, occurring on yeast histone H2A at position 105 (Q105) and human H2A at Q104. We identify Nop1 as the methyltransferase in yeast and demonstrate that fibrillarin is the orthologue enzyme in human cells. Glutamine methylation of H2A is restricted to the nucleolus. Global analysis in yeast, using an H2AQ105me-specific antibody, shows that this modification is exclusively enriched over the 35S ribosomal DNA transcriptional unit. We show that the Q105 residue is part of the binding site for the histone chaperone FACT (facilitator of chromatin transcription) complex. Methylation of Q105 or its substitution to alanine disrupts binding to FACT in vitro. A yeast strain mutated at Q105 shows reduced histone incorporation and increased transcription at the ribosomal DNA locus. These features are phenocopied by mutations in FACT complex components. Together these data identify glutamine methylation of H2A as the first histone epigenetic mark dedicated to a specific RNA polymerase and define its function as a regulator of FACT interaction with nucleosomes.

  20. Characterization of the E.coli proteome and its modifications during growth and ethanol stress

    Directory of Open Access Journals (Sweden)

    Boumediene eSoufi

    2015-02-01

    Full Text Available We set out to provide a resource to the microbiology community especially with respect to systems biology based endeavors. To this end, we generated a comprehensive dataset monitoring the changes in protein expression, copy number, and post translational modifications in a systematic fashion during growth and ethanol stress in E.coli. We utilized high-resolution mass spectrometry combined with the Super-SILAC approach. In a single experiment, we have identified over 2,300 proteins, which represent approximately 88% of the estimated expressed proteome of E. coli and estimated protein copy numbers using the Intensity Based Absolute Quantitation (IBAQ. The dynamic range of protein expression spanned up to six orders of magnitude, with the highest protein copy per cell estimated at approximately 300,000. We focused on the proteome dynamics involved during stationary phase growth. A global up-regulation of proteins related to stress response was detected in later stages of growth. We observed the down-regulation of the methyl directed mismatch repair system containing MutS and MutL of E. coli growing in long term growth cultures, confirming that higher incidence of mutations presents an important mechanism in the increase in genetic diversity and stationary phase survival in E.coli. During ethanol stress, known markers such as alcohol dehydrogenase and aldehyde dehydrogenase were induced, further validating the dataset. Finally, we performed unbiased protein modification detection and revealed changes of many known and unknown protein modifications in both experimental conditions.

  1. Permit application modifications

    Energy Technology Data Exchange (ETDEWEB)

    NONE

    1995-11-01

    This document contains the Permit Application Modifications for the Y-12 Industrial Landfill V site on the Oak Ridge Reservation. These modifications include the assessment of stability of the proposed Landfill V under static and loading conditions. Analyses performed include the general slope stability, veneer stability of the bottom liner and cover system, and a liquefaction potential assessment of the foundation soils.

  2. The management of modifications

    International Nuclear Information System (INIS)

    Bernard, C.

    1992-01-01

    Description of the management methods of modifications at EDF. To maintain safety standards of the nuclear power station the 'Direction de l'Equipment' and the 'Direction du Parc en Exploitation' have jointly fixed the modalities of management for all modifications and recorded them in a 'Practical Guide'

  3. Permit application modifications

    International Nuclear Information System (INIS)

    1995-11-01

    This document contains the Permit Application Modifications for the Y-12 Industrial Landfill V site on the Oak Ridge Reservation. These modifications include the assessment of stability of the proposed Landfill V under static and loading conditions. Analyses performed include the general slope stability, veneer stability of the bottom liner and cover system, and a liquefaction potential assessment of the foundation soils

  4. Prediction of protein modification sites of pyrrolidone carboxylic acid using mRMR feature selection and analysis.

    Directory of Open Access Journals (Sweden)

    Lu-Lu Zheng

    Full Text Available Pyrrolidone carboxylic acid (PCA is formed during a common post-translational modification (PTM of extracellular and multi-pass membrane proteins. In this study, we developed a new predictor to predict the modification sites of PCA based on maximum relevance minimum redundancy (mRMR and incremental feature selection (IFS. We incorporated 727 features that belonged to 7 kinds of protein properties to predict the modification sites, including sequence conservation, residual disorder, amino acid factor, secondary structure and solvent accessibility, gain/loss of amino acid during evolution, propensity of amino acid to be conserved at protein-protein interface and protein surface, and deviation of side chain carbon atom number. Among these 727 features, 244 features were selected by mRMR and IFS as the optimized features for the prediction, with which the prediction model achieved a maximum of MCC of 0.7812. Feature analysis showed that all feature types contributed to the modification process. Further site-specific feature analysis showed that the features derived from PCA's surrounding sites contributed more to the determination of PCA sites than other sites. The detailed feature analysis in this paper might provide important clues for understanding the mechanism of the PCA formation and guide relevant experimental validations.

  5. Modifications to POISSON

    International Nuclear Information System (INIS)

    Harwood, L.H.

    1981-01-01

    At MSU we have used the POISSON family of programs extensively for magnetic field calculations. In the presently super-saturated computer situation, reducing the run time for the program is imperative. Thus, a series of modifications have been made to POISSON to speed up convergence. Two of the modifications aim at having the first guess solution as close as possible to the final solution. The other two aim at increasing the convergence rate. In this discussion, a working knowledge of POISSON is assumed. The amount of new code and expected time saving for each modification is discussed

  6. A central role for ubiquitination within a circadian clock protein modification code

    Directory of Open Access Journals (Sweden)

    Katarina eStojkovic

    2014-08-01

    Full Text Available Circadian rhythms, endogenous cycles of about 24 h in physiology, are generated by a master clock located in the suprachiasmatic nucleus of the hypothalamus and other clocks located in the brain and peripheral tissues. Circadian disruption is known to increase the incidence of various illnesses, such as mental disorders, metabolic syndrome and cancer. At the molecular level, periodicity is established by a set of clock genes via autoregulatory translation-transcription feedback loops. This clock mechanism is regulated by post-translational modifications such as phosphorylation and ubiquitination, which set the pace of the clock. Ubiquitination in particular has been found to regulate the stability of core clock components, but also other clock protein functions. Mutation of genes encoding ubiquitin ligases can cause either elongation or shortening of the endogenous circadian period. Recent research has also started to uncover roles for deubiquitination in the molecular clockwork. Here we review the role of the ubiquitin pathway in regulating the circadian clock and we propose that ubiquitination is a key element in a clock protein modification code that orchestrates clock mechanisms and circadian behavior over the daily cycle.

  7. Proteomic analysis of the cyanobacterium of the Azolla symbiosis: identity, adaptation, and NifH modification.

    Science.gov (United States)

    Ekman, Martin; Tollbäck, Petter; Bergman, Birgitta

    2008-01-01

    Cyanobacteria are able to form stable nitrogen-fixing symbioses with diverse eukaryotes. To extend our understanding of adaptations imposed by plant hosts, two-dimensional gel electrophoresis and mass spectrometry (MS) were used for comparative protein expression profiling of a cyanobacterium (cyanobiont) dwelling in leaf cavities of the water-fern Azolla filiculoides. Homology-based protein identification using peptide mass fingerprinting [matrix-assisted laser desorption ionization-time of flight (MALDI-TOF-MS)], tandem MS analyses, and sequence homology searches resulted in an identification success rate of 79% of proteins analysed in the unsequenced cyanobiont. Compared with a free-living strain, processes related to energy production, nitrogen and carbon metabolism, and stress-related functions were up-regulated in the cyanobiont while photosynthesis and metabolic turnover rates were down-regulated, stressing a slow heterotrophic mode of growth, as well as high heterocyst frequencies and nitrogen-fixing capacities. The first molecular data set on the nature of the NifH post-translational modification in cyanobacteria was also obtained: peptide mass spectra of the protein demonstrated the presence of a 300-400 Da protein modification localized to a specific 13 amino acid sequence, within the part of the protein that is ADP-ribosylated in other bacteria and close to the active site of nitrogenase. Furthermore, the distribution of the highest scoring database hits for the identified proteins points to the possibility of using proteomic data in taxonomy.

  8. Infection Reveals a Modification of SIRT2 Critical for Chromatin Association

    Directory of Open Access Journals (Sweden)

    Jorge M. Pereira

    2018-04-01

    Full Text Available Summary: Sirtuin 2 is a nicotinamide-adenine-dinucleotide-dependent deacetylase that regulates cell processes such as carcinogenesis, cell cycle, DNA damage, and infection. Subcellular localization of SIRT2 is crucial for its function but is poorly understood. Infection with the bacterial pathogen Listeria monocytogenes, which relocalizes SIRT2 from the cytoplasm to the chromatin, provides an ideal stimulus for the molecular study of this process. In this report, we provide a map of SIRT2 post-translational modification sites and focus on serine 25 phosphorylation. We show that infection specifically induces dephosphorylation of S25, an event essential for SIRT2 chromatin association. Furthermore, we identify a nuclear complex formed by the phosphatases PPM1A and PPM1B, with SIRT2 essential for controlling H3K18 deacetylation and SIRT2-mediated gene repression during infection and necessary for a productive Listeria infection. This study reveals a molecular mechanism regulating SIRT2 function and localization, paving the way for understanding other SIRT2-regulated cellular processes. : Sirtuins are enzymes critical for various processes, including genomic stability, metabolism, and aging. Through study of Listeria monocytogenes, a bacterial pathogen that exploits SIRT2 for productive infection, Pereira et al. uncover a SIRT2 modification necessary for chromatin association and function. Keywords: chromatin, sirtuin, Listeria monocytogenes, phosphorylation, PPM1, histone acetylation, H3K18, infection, subcellular localization

  9. Accommodating for plant modifications

    International Nuclear Information System (INIS)

    Weirich, P.H.

    1977-01-01

    Modification to a nuclear power plant may have different causes: 1) new instructions by the authorities; 2) changes of the marginal conditions on the construction site; 3) progress in the technological development. - Examples from different plants are supposed to demonstrate how such changes influence the planning or the construction and how they are integrated in the process of preparation. A distinction can be made between modifications before the completion of the submission of the order, during the phase of preparatory planning and during the construction phase. Of great importance are especially modifications made after the beginning of the construction works, since, in general, there is little scope for technical modifications and since consequences for the time schedule are to be expected. (orig.) [de

  10. Structural dynamic modification

    Indian Academy of Sciences (India)

    and stiffness matrices) andaor modal parameters, in order to acquire some ... For the above reasons, another modification approach is presented here ... The data necessary to solve the direct problem are dynamic behaviour of the original.

  11. Contrasting patterns of evolutionary constraint and novelty revealed by comparative sperm proteomic analysis in Lepidoptera.

    Science.gov (United States)

    Whittington, Emma; Forsythe, Desiree; Borziak, Kirill; Karr, Timothy L; Walters, James R; Dorus, Steve

    2017-12-02

    Rapid evolution is a hallmark of reproductive genetic systems and arises through the combined processes of sequence divergence, gene gain and loss, and changes in gene and protein expression. While studies aiming to disentangle the molecular ramifications of these processes are progressing, we still know little about the genetic basis of evolutionary transitions in reproductive systems. Here we conduct the first comparative analysis of sperm proteomes in Lepidoptera, a group that exhibits dichotomous spermatogenesis, in which males produce a functional fertilization-competent sperm (eupyrene) and an incompetent sperm morph lacking nuclear DNA (apyrene). Through the integrated application of evolutionary proteomics and genomics, we characterize the genomic patterns potentially associated with the origination and evolution of this unique spermatogenic process and assess the importance of genetic novelty in Lepidopteran sperm biology. Comparison of the newly characterized Monarch butterfly (Danaus plexippus) sperm proteome to those of the Carolina sphinx moth (Manduca sexta) and the fruit fly (Drosophila melanogaster) demonstrated conservation at the level of protein abundance and post-translational modification within Lepidoptera. In contrast, comparative genomic analyses across insects reveals significant divergence at two levels that differentiate the genetic architecture of sperm in Lepidoptera from other insects. First, a significant reduction in orthology among Monarch sperm genes relative to the remainder of the genome in non-Lepidopteran insect species was observed. Second, a substantial number of sperm proteins were found to be specific to Lepidoptera, in that they lack detectable homology to the genomes of more distantly related insects. Lastly, the functional importance of Lepidoptera specific sperm proteins is broadly supported by their increased abundance relative to proteins conserved across insects. Our results identify a burst of genetic novelty

  12. Pragmatic Graph Rewriting Modifications

    OpenAIRE

    Rodgers, Peter; Vidal, Natalia

    1999-01-01

    We present new pragmatic constructs for easing programming in visual graph rewriting programming languages. The first is a modification to the rewriting process for nodes the host graph, where nodes specified as 'Once Only' in the LHS of a rewrite match at most once with a corresponding node in the host graph. This reduces the previously common use of tags to indicate the progress of matching in the graph. The second modification controls the application of LHS graphs, where those specified a...

  13. Modification of JRR-2

    International Nuclear Information System (INIS)

    Miyasaka, Yasuhiko

    1978-01-01

    This report gives an outline of some of the main modifications carried out around the Reactor Core on the Research Reactor JRR-2, at the Tokai Research Establishment of JAERI. The JRR-2 was shut down in December 1973, to improve it in heavy water leakage from the metal packing between core tank and support ring, corrosion of the lower shielding plug, and fault in the control-rod mechanism. Main modifications were a standing seal weld at the support ring to stop heavy water leakage, replacement of the reactor top shield and improvement of the helium system. The control-rod assemblies and the refueling devices were replaced by the newly designed ones also. In addition to the modification plan, the irradiated air exhaust system was improved to reduce radioactive argon gas release through the stack. Works were completed successfully in September 1975. But a light water leakage occurred at the stand pipe below the light water tank on November 11, 1975, which was repaired in about 4 months. When considering the operation of above 5,000 hours after the modification, however, the quality of the modification work may be said to be quite satisfactory. The present report in which works to the completion are described may be valuable as a record of reactor modification which is a new experience at JAERI. (auth.)

  14. Human Rights and Behavior Modification

    Science.gov (United States)

    Roos, Philip

    1974-01-01

    Criticisms of behavior modification, which charge that it violates ethical and legal principles, are discussed and reasons are presented to explain behavior modification's susceptibility to attack. (GW)

  15. Functional significance of O-GlcNAc modification in regulating neuronal properties.

    Science.gov (United States)

    Hwang, Hongik; Rhim, Hyewhon

    2018-03-01

    Post-translational modifications (PTMs) covalently modify proteins and diversify protein functions. Along with protein phosphorylation, another common PTM is the addition of O-linked β-N-acetylglucosamine (O-GlcNAc) to serine and/or threonine residues. O-GlcNAc modification is similar to phosphorylation in that it occurs to serine and threonine residues and cycles on and off with a similar time scale. However, a striking difference is that the addition and removal of the O-GlcNAc moiety on all substrates are mediated by the two enzymes regardless of proteins, O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA), respectively. O-GlcNAcylation can interact or potentially compete with phosphorylation on serine and threonine residues, and thus serves as an important molecular mechanism to modulate protein functions and activation. However, it has been challenging to address the role of O-GlcNAc modification in regulating protein functions at the molecular level due to the lack of convenient tools to determine the sites and degrees of O-GlcNAcylation. Studies in this field have only begun to expand significantly thanks to the recent advances in detection and manipulation methods such as quantitative proteomics and highly selective small-molecule inhibitors for OGT and OGA. Interestingly, multiple brain regions, especially hippocampus, express high levels of both OGT and OGA, and a number of neuron-specific proteins have been reported to undergo O-GlcNAcylation. This review aims to discuss the recent updates concerning the impacts of O-GlcNAc modification on neuronal functions at multiple levels ranging from intrinsic neuronal properties to synaptic plasticity and animal behaviors. Copyright © 2017 Elsevier Ltd. All rights reserved.

  16. mRNA Display Selection of a High-Affinity, Modification-Specific Phospho-IκBα-Binding Fibronectin

    Science.gov (United States)

    Olson, C. Anders; Liao, Hsiang-I; Sun, Ren; Roberts, Richard W.

    2009-01-01

    The complexity of the human proteome is greatly expanded by post-translational modifications. New tools capable of recognizing these modifications in a sequence-specific fashion provide a route to purify these modified proteins, to alter protein trafficking, and to visualize signal transduction in real time. Here, we have evolved novel, modification-specific ligands that target phosphorylated IκBα. To do this, we employed mRNA display-based in vitro selection using a 30-trillion-member protein library based on the fibronectin type III domain. The selection yielded one fibronectin molecule, 10C17C25, that binds a phospho-IκBα peptide with Kd = 18 nM and is over 1000-fold specific compared to the nonphosphorylated peptide. 10C17C25 specifically recognizes endogenous phosphorylated IκBα from mammalian cell extract and stabilizes phospho-IκBα in vivo. We also incorporated 10C17C25 into a FRET indicator that detects IκB kinase (IKK) activity in vitro, demonstrating the utility of selecting designed adaptors for kinase activity sensors. PMID:18590330

  17. mRNA display selection of a high-affinity, modification-specific phospho-IkappaBalpha-binding fibronectin.

    Science.gov (United States)

    Olson, C Anders; Liao, Hsiang-I; Sun, Ren; Roberts, Richard W

    2008-08-15

    The complexity of the human proteome is greatly expanded by post-translational modifications. New tools capable of recognizing these modifications in a sequence-specific fashion provide a route to purify these modified proteins, to alter protein trafficking, and to visualize signal transduction in real time. Here, we have evolved novel, modification-specific ligands that target phosphorylated IkappaBalpha. To do this, we employed mRNA display-based in vitro selection using a 30-trillion-member protein library based on the fibronectin type III domain. The selection yielded one fibronectin molecule, 10C17C25, that binds a phospho-IkappaBalpha peptide with K d = 18 nM and is over 1000-fold specific compared to the nonphosphorylated peptide. 10C17C25 specifically recognizes endogenous phosphorylated IkappaBalpha from mammalian cell extract and stabilizes phospho-IkappaBalpha in vivo. We also incorporated 10C17C25 into a FRET indicator that detects IkappaB kinase (IKK) activity in vitro, demonstrating the utility of selecting designed adaptors for kinase activity sensors.

  18. Human protein secretory pathway genes are expressed in a tissue-specific pattern to match processing demands of the secretome

    DEFF Research Database (Denmark)

    Feizi, Amir; Gatto, Francesco; Uhlén, Mathias

    2017-01-01

    Protein secretory pathway in eukaryal cells is responsible for delivering functional secretory proteins. The dysfunction of this pathway causes a range of important human diseases from congenital disorders to cancer. Despite the piled-up knowledge on the molecular biology and biochemistry level...... in specific gene families of the secretory pathway. We also inspected the potential functional link between detected extreme genes and the corresponding tissues enriched secretome. As a result, the detected extreme genes showed correlation with the enrichment of the nature and number of specific post......-translational modifications in each tissue's secretome. Our findings conciliate both the housekeeping and tissue-specific nature of the protein secretory pathway, which we attribute to a fine-tuned regulation of defined gene families to support the diversity of secreted proteins and their modifications....

  19. Predicting post-translational lysine acetylation using support vector machines

    DEFF Research Database (Denmark)

    Gnad, Florian; Ren, Shubin; Choudhary, Chunaram

    2010-01-01

    spectrometry to identify 3600 lysine acetylation sites on 1750 human proteins covering most of the previously annotated sites and providing the most comprehensive acetylome so far. This dataset should provide an excellent source to train support vector machines (SVMs) allowing the high accuracy in silico...

  20. Post-translational regulation of Oct4 transcriptional activity.

    Directory of Open Access Journals (Sweden)

    Jonathan P Saxe

    Full Text Available Oct4 is a key component of the molecular circuitry which regulates embryonic stem cell proliferation and differentiation. It is essential for maintenance of undifferentiated, pluripotent cell populations, and accomplishes these tasks by binding DNA in multiple heterodimer and homodimer configurations. Very little is known about how formation of these complexes is regulated, or the mechanisms through which Oct4 proteins respond to complex extracellular stimuli which regulate pluripotency. Here, we provide evidence for a phosphorylation-based mechanism which regulates specific Oct4 homodimer conformations. Point mutations of a putative phosphorylation site can specifically abrogate transcriptional activity of a specific homodimer assembly, with little effect on other configurations. Moreover, we performed bioinformatic predictions to identify a subset of Oct4 target genes which may be regulated by this specific assembly, and show that altering Oct4 protein levels affects transcription of Oct4 target genes which are regulated by this assembly but not others. Finally, we identified several signaling pathways which may mediate this phosphorylation and act in combination to regulate Oct4 transcriptional activity and protein stability. These results provide a mechanism for rapid and reversible alteration of Oct4 transactivation potential in response to extracellular signals.

  1. The PTEN protein: cellular localization and post-translational regulation.

    Science.gov (United States)

    Leslie, Nick R; Kriplani, Nisha; Hermida, Miguel A; Alvarez-Garcia, Virginia; Wise, Helen M

    2016-02-01

    The phosphatase and tensin homologue deleted on chromosome 10 (PTEN) phosphatase dephosphorylates PIP3, the lipid product of the class I PI 3-kinases, and suppresses the growth and proliferation of many cell types. It has been heavily studied, in large part due to its status as a tumour suppressor, the loss of function of which is observed through diverse mechanisms in many tumour types. Here we present a concise review of our understanding of the PTEN protein and highlight recent advances, particularly in our understanding of its localization and regulation by ubiquitination and SUMOylation. © 2016 Authors; published by Portland Press Limited.

  2. SUCROSE SYNTHASE: ELUCIDATION OF COMPLEX POST-TRANSLATIONAL REGULATORY MECHANISMS

    Energy Technology Data Exchange (ETDEWEB)

    Steven C. Huber

    2009-05-12

    Studies have focused on the enzyme sucrose synthase, which plays an important role in the metabolism of sucrose in seeds and tubers. There are three isoforms of SUS in maize, referred to as SUS1, SUS-SH1, and SUS2. SUS is generally considered to be tetrameric protein but recent evidence suggests that SUS can also occur as a dimeric protein. The formation of tetrameric SUS is regulated by sucrose concentration in vitro and this could also be an important factor in the cellular localization of the protein. We found that high sucrose concentrations, which promote tetramer formation, also inhibit the binding of SUS1 to actin filaments in vitro. Previously, high sucrose concentrations were shown to promote SUS association with the plasma membrane. The specific regions of the SUS molecule involved in oligomerization are not known, but we identified a region of the SUS1 moelcule by bioinformatic analysis that was predicted to form a coiled coil. We demonstrated that this sequence could, in fact, self-associate as predicted for a coiled coil, but truncation analysis with the full-length recombinant protein suggested that it was not responsible for formation of dimers or tetramers. However, the coiled coil may function in binding of other proteins to SUS1. Overall, sugar availability may differentially influence the binding of SUS to cellular structures, and these effects may be mediated by changes in the oligomeric nature of the enzyme.

  3. Progressive methylation of ageing histones by Dot1 functions as a timer

    NARCIS (Netherlands)

    De Vos, Dirk; Frederiks, Floor; Terweij, Marit; van Welsem, Tibor; Verzijlbergen, Kitty F.; Iachina, Ekaterina; de Graaf, Erik L.; Altelaar, A. F. Maarten; Oudgenoeg, Gideon; Heck, Albert J. R.; Krijgsveldz, Jeroen; Bakker, Barbara M.; van Leeuwen, Fred

    Post-translational modifications of histone proteins have a crucial role in regulating gene expression. If efficiently re-established after chromosome duplication, histone modifications could help propagate gene expression patterns in dividing cells by epigenetic mechanisms. We used an integrated

  4. Mining Proteomic Data to Expose Protein Modifications in Methanosarcina mazei strain Gö1

    Directory of Open Access Journals (Sweden)

    Deborah eLeon

    2015-03-01

    Full Text Available Proteomic tools identify constituents of complex mixtures, often delivering long lists of identified proteins. The high-throughput methods excel at matching tandem mass spectrometry data to spectra predicted from sequence databases. Unassigned mass spectra are ignored, but could, in principle, provide valuable information on unanticipated modifications and improve protein annotations while consuming limited quantities of material. Strategies to mine information from these discards are presented, along with discussion of features that, when present, provide strong support for modifications. In this study we mined LC-MS/MS datasets of proteolytically-digested concanavalin A pull down fractions from Methanosarcina mazei Gö1 cell lysates. Analyses identified 154 proteins. Many of the observed proteins displayed post-translationally modified forms, including O-formylated and methyl-esterified segments that appear biologically relevant (i.e., not artifacts of sample handling. Interesting cleavages and modifications (e.g., S-cyanylation and trimethylation were observed near catalytic sites of methanogenesis enzymes. Of 31 Methanosarcina protein N-termini recovered by concanavalin A binding or from a previous study, only M. mazei S-layer protein MM1976 and its M. acetivorans C2A orthologue, MA0829, underwent signal peptide excision. Experimental results contrast with predictions from algorithms SignalP 3.0 and Exprot, which were found to over-predict the presence of signal peptides. Proteins MM0002, MM0716, MM1364, and MM1976 were found to be glycosylated, and employing chromatography tailored specifically for glycopeptides will likely reveal more.This study supplements limited, existing experimental datasets of mature archaeal N-termini, including presence or absence of signal peptides, translation initiation sites, and other processing. Methanosarcina surface and membrane proteins are richly modified.

  5. Enzymatic modification of starch

    DEFF Research Database (Denmark)

    Jensen, Susanne Langgård

    In the food industry approaches for using bioengineering are investigated as alternatives to conventional chemical and physical starch modification techniques in development of starches with specific properties. Enzyme-assisted post-harvest modification is an interesting approach to this, since...... it is considered a clean and energy saving technology. This thesis aimed to investigate the effect of using reaction conditions, simulating an industrial process, for enzymatic treatment of starch with branching enzyme (BE) from Rhodothermus obamensis. Thus treatements were conducted at 70°C using very high...... substrate concentration (30-40% dry matter (DM)) and high enzyme activity (750-2250 BE units (BEU)/g sample). Starches from various botanical sources, representing a broad range of properties, were used as substrates. The effects of the used conditions on the BE-reaction were evaluated by characterization...

  6. The Listeria monocytogenes Bile Stimulon under Acidic Conditions Is Characterized by Strain-Specific Patterns and the Upregulation of Motility, Cell Wall Modification Functions, and the PrfA Regulon

    Science.gov (United States)

    Guariglia-Oropeza, Veronica; Orsi, Renato H.; Guldimann, Claudia; Wiedmann, Martin; Boor, Kathryn J.

    2018-01-01

    Listeria monocytogenes uses a variety of transcriptional regulation strategies to adapt to the extra-host environment, the gastrointestinal tract, and the intracellular host environment. While the alternative sigma factor SigB has been proposed to be a key transcriptional regulator that facilitates L. monocytogenes adaptation to the gastrointestinal environment, the L. monocytogenes' transcriptional response to bile exposure is not well-understood. RNA-seq characterization of the bile stimulon was performed in two L. monocytogenes strains representing lineages I and II. Exposure to bile at pH 5.5 elicited a large transcriptomic response with ~16 and 23% of genes showing differential transcription in 10403S and H7858, respectively. The bile stimulon includes genes involved in motility and cell wall modification mechanisms, as well as genes in the PrfA regulon, which likely facilitate survival during the gastrointestinal stages of infection that follow bile exposure. The fact that bile exposure induced the PrfA regulon, but did not induce further upregulation of the SigB regulon (beyond that expected by exposure to pH 5.5), suggests a model where at the earlier stages of gastrointestinal infection (e.g., acid exposure in the stomach), SigB-dependent gene expression plays an important role. Subsequent exposure to bile induces the PrfA regulon, potentially priming L. monocytogenes for subsequent intracellular infection stages. Some members of the bile stimulon showed lineage- or strain-specific distribution when 27 Listeria genomes were analyzed. Even though sigB null mutants showed increased sensitivity to bile, the SigB regulon was not found to be upregulated in response to bile beyond levels expected by exposure to pH 5.5. Comparison of wildtype and corresponding ΔsigB strains newly identified 26 SigB-dependent genes, all with upstream putative SigB-dependent promoters. PMID:29467736

  7. Geometrically Consistent Mesh Modification

    KAUST Repository

    Bonito, A.

    2010-01-01

    A new paradigm of adaptivity is to execute refinement, coarsening, and smoothing of meshes on manifolds with incomplete information about their geometry and yet preserve position and curvature accuracy. We refer to this collectively as geometrically consistent (GC) mesh modification. We discuss the concept of discrete GC, show the failure of naive approaches, and propose and analyze a simple algorithm that is GC and accuracy preserving. © 2010 Society for Industrial and Applied Mathematics.

  8. Radiation modification of materials

    International Nuclear Information System (INIS)

    Pikaev, A.K.

    1987-01-01

    Industrial and radiation chemical processes of material modification based on cross-linking of polymers as a result of radiation are considered. Among them are production of cables and rods with irradiated modified insulation, production of hardened and thermo-shrinkaging polymer products (films, tubes, fashioned products), production of radiation cross-linked polyethylene foam, technology of radiation vulcanization of elastomers. Attention is paid to radiation plants on the basis of γ-sources and electron acceleratos as well as to radiation conditions

  9. Behavior Modification in the Classroom

    Science.gov (United States)

    Whitman, Mryon; Whitman, Joan

    1971-01-01

    This article presents the theoretical rationale for behavior modification, principally through its comparison with traditional psychotherapies, and suggests some behavior modification techniques for the classroom management of maladaptive behavior. (Author)

  10. Biomaterials modification by ion beam

    International Nuclear Information System (INIS)

    Zhang Tonghe; Yi Zhongzhen; Zhang Xu; Wu Yuguang

    2001-01-01

    Ion beam technology is one of best ways for the modification of biomaterials. The results of ion beam modification of biomaterials are given. The method and results of improved biocompatibility are indicated by ion beam technology. The future development of ion beam modification of biomaterials is discussed

  11. Epigenetic modifications by Trithorax group proteins during early embryogenesis: do members of Trx-G function as maternal effect genes?

    Science.gov (United States)

    Andreu-Vieyra, Claudia; Matzuk, Martin M

    2007-02-01

    Maternal effect genes encode transcripts that are expressed during oogenesis. These gene products are stored in the oocyte and become functional during resumption of meiosis and zygote genome activation, and in embryonic stem cells. To date, a few maternal effect genes have been identified in mammals. Epigenetic modifications have been shown to be important during early embryonic development and involve DNA methylation and post-translational modification of core histones. During development, two families of proteins have been shown to be involved in epigenetic changes: Trithorax group (Trx-G) and Polycomb group (Pc-G) proteins. Trx-G proteins function as transcriptional activators and have been shown to accumulate in the oocyte. Deletion of Trx-G members using conventional knockout technology results in embryonic lethality in the majority of the cases analysed to date. Recent studies using conditional knockout mice have revealed that at least one family member is necessary for zygote genome activation. We propose that other Trx-G members may also regulate embryonic genome activation and that the use of oocyte-specific deletor mouse lines will help clarify their roles in this process.

  12. Combinatorial modification of human histone H4 quantitated by two-dimensional liquid chromatography coupled with top down mass spectrometry.

    Science.gov (United States)

    Pesavento, James J; Bullock, Courtney R; LeDuc, Richard D; Mizzen, Craig A; Kelleher, Neil L

    2008-05-30

    Quantitative proteomics has focused heavily on correlating protein abundances, ratios, and dynamics by developing methods that are protein expression-centric (e.g. isotope coded affinity tag, isobaric tag for relative and absolute quantification, etc.). These methods effectively detect changes in protein abundance but fail to provide a comprehensive perspective of the diversity of proteins such as histones, which are regulated by post-translational modifications. Here, we report the characterization of modified forms of HeLa cell histone H4 with a dynamic range >10(4) using a strictly Top Down mass spectrometric approach coupled with two dimensions of liquid chromatography. This enhanced dynamic range enabled the precise characterization and quantitation of 42 forms uniquely modified by combinations of methylation and acetylation, including those with trimethylated Lys-20, monomethylated Arg-3, and the novel dimethylated Arg-3 (each <1% of all H4 forms). Quantitative analyses revealed distinct trends in acetylation site occupancy depending on Lys-20 methylation state. Because both modifications are dynamically regulated through the cell cycle, we simultaneously investigated acetylation and methylation kinetics through three cell cycle phases and used these data to statistically assess the robustness of our quantitative analysis. This work represents the most comprehensive analysis of histone H4 forms present in human cells reported to date.

  13. Genetic modification and genetic determinism

    Science.gov (United States)

    Resnik, David B; Vorhaus, Daniel B

    2006-01-01

    In this article we examine four objections to the genetic modification of human beings: the freedom argument, the giftedness argument, the authenticity argument, and the uniqueness argument. We then demonstrate that each of these arguments against genetic modification assumes a strong version of genetic determinism. Since these strong deterministic assumptions are false, the arguments against genetic modification, which assume and depend upon these assumptions, are therefore unsound. Serious discussion of the morality of genetic modification, and the development of sound science policy, should be driven by arguments that address the actual consequences of genetic modification for individuals and society, not by ones propped up by false or misleading biological assumptions. PMID:16800884

  14. The continual fuel management modification in Qinshan project II

    International Nuclear Information System (INIS)

    Ye Guodong; Pan Zefei; Zhang Xingtian

    2010-01-01

    The fuel management strategy is the basis of the nuclear power plants. The performance of the fuel management strategy affects the plants' safety and economy indicators directly. The paper summarizes all the modifications on the fuel management work in Qinshan Project II since the plant was established. It includes the surveillance system of physics tests, fetching in high performance fuel assemblies, reloading pattern optimization, and the modifications of the final safety analysis report. At the same time, it evaluates the benefit of the modifications in the few years. The experience in this paper is much helpful and could be implemented on the same type plants. (authors)

  15. Mutually exclusive STAT1 modifications identified by Ubc9/substrate dimerization-dependent SUMOylation.

    Science.gov (United States)

    Zimnik, Susan; Gaestel, Matthias; Niedenthal, Rainer

    2009-03-01

    Post-translational modifications control the physiological activity of the signal transducer and activator of transcription STAT1. While phosphorylation at tyrosine Y701 is a prerequisite for STAT1 dimerization, its SUMOylation represses the transcriptional activity. Recently, we have demonstrated that SUMOylation at lysine K703 inhibits the phosphorylation of nearby localized Y701 of STAT1. Here, we analysed the influence of phosphorylation of Y701 on SUMOylation of K703 in vivo. For that reason, an Ubc9/substrate dimerization-dependent SUMOylation (USDDS) system was developed, which consists of fusions of the SUMOylation substrate and of the SUMO-conjugating enzyme Ubc9 to the chemically activatable heterodimerization domains FKBP and FRB, respectively. When FKBP fusion proteins of STAT1, p53, CRSP9, FOS, CSNK2B, HES1, TCF21 and MYF6 are coexpressed with Ubc9-FRB, treatment of HEK293 cells with the rapamycin-related dimerizer compound AP21967 induces SUMOylation of these proteins in vivo. For STAT1-FKBP and p53-FKBP we show that this SUMOylation takes place at their specific SUMOylation sites in vivo. Using USDDS, we then demonstrate that STAT1 phosphorylation at Y701 induced by interferon-beta treatment inhibits SUMOylation of K703 in vivo. Thus, pY701 and SUMO-K703 of STAT1 represent mutually exclusive modifications, which prevent signal integration at this molecule and probably ensure the existence of differentially modified subpopulations of STAT1 necessary for its regulated nuclear cytoplasmic activation/inactivation cycle.

  16. Epigenetic and Posttranslational Modifications in Light Signal Transduction and the Circadian Clock in Neurospora crassa

    Directory of Open Access Journals (Sweden)

    Marco Proietto

    2015-07-01

    Full Text Available Blue light, a key abiotic signal, regulates a wide variety of physiological processes in many organisms. One of these phenomena is the circadian rhythm presents in organisms sensitive to the phase-setting effects of blue light and under control of the daily alternation of light and dark. Circadian clocks consist of autoregulatory alternating negative and positive feedback loops intimately connected with the cellular metabolism and biochemical processes. Neurospora crassa provides an excellent model for studying the molecular mechanisms involved in these phenomena. The White Collar Complex (WCC, a blue-light receptor and transcription factor of the circadian oscillator, and Frequency (FRQ, the circadian clock pacemaker, are at the core of the Neurospora circadian system. The eukaryotic circadian clock relies on transcriptional/translational feedback loops: some proteins rhythmically repress their own synthesis by inhibiting the activity of their transcriptional factors, generating self-sustained oscillations over a period of about 24 h. One of the basic mechanisms that perpetuate self-sustained oscillations is post translation modification (PTM. The acronym PTM generically indicates the addition of acetyl, methyl, sumoyl, or phosphoric groups to various types of proteins. The protein can be regulatory or enzymatic or a component of the chromatin. PTMs influence protein stability, interaction, localization, activity, and chromatin packaging. Chromatin modification and PTMs have been implicated in regulating circadian clock function in Neurospora. Research into the epigenetic control of transcription factors such as WCC has yielded new insights into the temporal modulation of light-dependent gene transcription. Here we report on epigenetic and protein PTMs in the regulation of the Neurospora crassa circadian clock. We also present a model that illustrates the molecular mechanisms at the basis of the blue light control of the circadian clock.

  17. Covalent protein modification with ISG15 via a conserved cysteine in the hinge region.

    Directory of Open Access Journals (Sweden)

    Veronika N Bade

    Full Text Available The ubiquitin-like protein ISG15 (interferon-stimulated gene of 15 kDa is strongly induced by type I interferons and displays antiviral activity. As other ubiquitin-like proteins (Ubls, ISG15 is post-translationally conjugated to substrate proteins by an isopeptide bond between the C-terminal glycine of ISG15 and the side chains of lysine residues in the substrates (ISGylation. ISG15 consists of two ubiquitin-like domains that are separated by a hinge region. In many orthologs, this region contains a single highly reactive cysteine residue. Several hundred potential substrates for ISGylation have been identified but only a few of them have been rigorously verified. In order to investigate the modification of several ISG15 substrates, we have purified ISG15 conjugates from cell extracts by metal-chelate affinity purification and immunoprecipitations. We found that the levels of proteins modified by human ISG15 can be decreased by the addition of reducing agents. With the help of thiol blocking reagents, a mutational analysis and miRNA mediated knock-down of ISG15 expression, we revealed that this modification occurs in living cells via a disulphide bridge between the substrates and Cys78 in the hinge region of ISG15. While the ISG15 activating enzyme UBE1L is conjugated by ISG15 in the classical way, we show that the ubiquitin conjugating enzyme Ubc13 can either be classically conjugated by ISG15 or can form a disulphide bridge with ISG15 at the active site cysteine 87. The latter modification would interfere with its function as ubiquitin conjugating enzyme. However, we found no evidence for an ISG15 modification of the dynamin-like GTPases MxA and hGBP1. These findings indicate that the analysis of potential substrates for ISG15 conjugation must be performed with great care to distinguish between the two types of modification since many assays such as immunoprecipitation or metal-chelate affinity purification are performed with little or no

  18. Enzymatic Modification of Sphingomyelin

    DEFF Research Database (Denmark)

    Due to its major role in maintaining the water-retaining properties of the epidermis, ceramide is of great commercial potential in cosmetic and pharmaceuticals such as hair and skin care products. Currently, chemical synthesis of ceramide is a costly process, and developments of alternative cost......-efficient, high yield production methods are of great interest. In the present study, the potential of producing ceramide through the enzymatic hydrolysis of sphingomyelin have been studied. sphingomyelin is a ubiquitous membrane-lipid and rich in dairy products or by-products. It has been verified...... that sphingomyelin modification gives a feasible approach to the potential production of ceramide. The reaction system has been improved through system evaluation and the optimization of several important factors, and phospholipase C from Clostridium perfringens shows higher activity towards the hydrolysis reaction...

  19. Patterned Colloidal Photonic Crystals.

    Science.gov (United States)

    Hou, Jue; Li, Mingzhu; Song, Yanlin

    2018-03-01

    Colloidal photonic crystals (PCs) have been well developed because they are easy to prepare, cost-effective, and versatile with regards to modification and functionalization. Patterned colloidal PCs contribute a novel approach to constructing high-performance PC devices with unique structures and specific functions. In this review, an overview of the strategies for fabricating patterned colloidal PCs, including patterned substrate-induced assembly, inkjet printing, and selective immobilization and modification, is presented. The advantages of patterned PC devices are also discussed in detail, for example, improved detection sensitivity and response speed of the sensors, control over the flow direction and wicking rate of microfluidic channels, recognition of cross-reactive molecules through an array-patterned microchip, fabrication of display devices with tunable patterns, well-arranged RGB units, and wide viewing-angles, and the ability to construct anti-counterfeiting devices with different security strategies. Finally, the perspective of future developments and challenges is presented. © 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  20. Genetic modification and genetic determinism

    Directory of Open Access Journals (Sweden)

    Vorhaus Daniel B

    2006-06-01

    Full Text Available Abstract In this article we examine four objections to the genetic modification of human beings: the freedom argument, the giftedness argument, the authenticity argument, and the uniqueness argument. We then demonstrate that each of these arguments against genetic modification assumes a strong version of genetic determinism. Since these strong deterministic assumptions are false, the arguments against genetic modification, which assume and depend upon these assumptions, are therefore unsound. Serious discussion of the morality of genetic modification, and the development of sound science policy, should be driven by arguments that address the actual consequences of genetic modification for individuals and society, not by ones propped up by false or misleading biological assumptions.

  1. Regulation of the tumor suppressor PML by sequential posttranslational modifications

    Directory of Open Access Journals (Sweden)

    Lienhard eSchmitz

    2012-12-01

    Full Text Available Posttranslational modifications (PTMs regulate multiple biological functions of the PML (promyelocytic leukemia protein and also the fission, disassembly and rebuilding of PML nuclear bodies (PML-NBs during the cell cycle. Pathway-specific PML modification patterns ensure proper signal output from PML-NBs that suit the specific functional requirements. Here we comprehensively review the signaling pathways and enzymes that modify PML and also the oncogenic PML-RARα fusion protein. Many PTMs occur in a hierarchical and timely organized fashion. Phosphorylation or acetylation constitute typical starting points for many PML modifying events, while degradative ubiquitination is an irreversible end point of the modification cascade. As this hierarchical organization of PTMs frequently turns phosphorylation events as primordial events, kinases or phosphatases regulating PML phosphorylation may be interesting drug targets to manipulate the downstream modifications and thus the stability and function of PML or PML-RARα.

  2. Glycobiology Modifications in Intratumoral Hypoxia: The Breathless Side of Glycans Interaction

    Directory of Open Access Journals (Sweden)

    Antônio F. Silva-filho

    2017-04-01

    Full Text Available Post-translational and co-translational enzymatic addition of glycans (glycosylation to proteins, lipids, and other carbohydrates, is a powerful regulator of the molecular machinery involved in cell cycle, adhesion, invasion, and signal transduction, and is usually seen in both in vivo and in vitro cancer models. Glycosyltransferases can alter the glycosylation pattern of normal cells, subsequently leading to the establishment and progression of several diseases, including cancer. Furthermore, a growing amount of research has shown that different oxygen tensions, mainly hypoxia, leads to a markedly altered glycosylation, resulting in altered glycan-receptor interactions. Alteration of intracellular glucose metabolism, from aerobic cellular respiration to anaerobic glycolysis, inhibition of integrin 3α1β translocation to the plasma membrane, decreased 1,2-fucosylation of cell-surface glycans, and galectin overexpression are some consequences of the hypoxic tumor microenvironment. Additionally, increased expression of gangliosides carrying N-glycolyl sialic acid can also be significantly affected by hypoxia. For all these reasons, it is possible to realize that hypoxia strongly alters glycobiologic events within tumors, leading to changes in their behavior. This review aims to analyze the complexity and importance of glycoconjugates and their molecular interaction network in the hypoxic context of many solid tumors.

  3. Differential effects of garcinol and curcumin on histone and p53 modifications in tumour cells

    Directory of Open Access Journals (Sweden)

    Collins Hilary M

    2013-01-01

    Full Text Available Abstract Background Post-translational modifications (PTMs of histones and other proteins are perturbed in tumours. For example, reduced levels of acetylated H4K16 and trimethylated H4K20 are associated with high tumour grade and poor survival in breast cancer. Drug-like molecules that can reprogram selected histone PTMs in tumour cells are therefore of interest as potential cancer chemopreventive agents. In this study we assessed the effects of the phytocompounds garcinol and curcumin on histone and p53 modification in cancer cells, focussing on the breast tumour cell line MCF7. Methods Cell viability/proliferation assays, cell cycle analysis by flow cytometry, immunodetection of specific histone and p53 acetylation marks, western blotting, siRNA and RT-qPCR. Results Although treatment with curcumin, garcinol or the garcinol derivative LTK-14 hampered MCF7 cell proliferation, differential effects of these compounds on histone modifications were observed. Garcinol treatment resulted in a strong reduction in H3K18 acetylation, which is required for S phase progression. Similar effects of garcinol on H3K18 acetylation were observed in the osteosarcoma cells lines U2OS and SaOS2. In contrast, global levels of acetylated H4K16 and trimethylated H4K20 in MCF7 cells were elevated after garcinol treatment. This was accompanied by upregulation of DNA damage signalling markers such as γH2A.X, H3K56Ac, p53 and TIP60. In contrast, exposure of MCF7 cells to curcumin resulted in increased global levels of acetylated H3K18 and H4K16, and was less effective in inducing DNA damage markers. In addition to its effects on histone modifications, garcinol was found to block CBP/p300-mediated acetylation of the C-terminal activation domain of p53, but resulted in enhanced acetylation of p53K120, and accumulation of p53 in the cytoplasmic compartment. Finally, we show that the elevation of H4K20Me3 levels by garcinol correlated with increased expression of SUV420H2

  4. Differential effects of garcinol and curcumin on histone and p53 modifications in tumour cells

    International Nuclear Information System (INIS)

    Collins, Hilary M; Kundu, Tapas K; Heery, David M; Abdelghany, Magdy K; Messmer, Marie; Yue, Baigong; Deeves, Sian E; Kindle, Karin B; Mantelingu, Kempegowda; Aslam, Akhmed; Winkler, G Sebastiaan

    2013-01-01

    Post-translational modifications (PTMs) of histones and other proteins are perturbed in tumours. For example, reduced levels of acetylated H4K16 and trimethylated H4K20 are associated with high tumour grade and poor survival in breast cancer. Drug-like molecules that can reprogram selected histone PTMs in tumour cells are therefore of interest as potential cancer chemopreventive agents. In this study we assessed the effects of the phytocompounds garcinol and curcumin on histone and p53 modification in cancer cells, focussing on the breast tumour cell line MCF7. Cell viability/proliferation assays, cell cycle analysis by flow cytometry, immunodetection of specific histone and p53 acetylation marks, western blotting, siRNA and RT-qPCR. Although treatment with curcumin, garcinol or the garcinol derivative LTK-14 hampered MCF7 cell proliferation, differential effects of these compounds on histone modifications were observed. Garcinol treatment resulted in a strong reduction in H3K18 acetylation, which is required for S phase progression. Similar effects of garcinol on H3K18 acetylation were observed in the osteosarcoma cells lines U2OS and SaOS2. In contrast, global levels of acetylated H4K16 and trimethylated H4K20 in MCF7 cells were elevated after garcinol treatment. This was accompanied by upregulation of DNA damage signalling markers such as γH2A.X, H3K56Ac, p53 and TIP60. In contrast, exposure of MCF7 cells to curcumin resulted in increased global levels of acetylated H3K18 and H4K16, and was less effective in inducing DNA damage markers. In addition to its effects on histone modifications, garcinol was found to block CBP/p300-mediated acetylation of the C-terminal activation domain of p53, but resulted in enhanced acetylation of p53K120, and accumulation of p53 in the cytoplasmic compartment. Finally, we show that the elevation of H4K20Me3 levels by garcinol correlated with increased expression of SUV420H2, and was prevented by siRNA targeting of SUV420H2. In

  5. Secretory signal peptide modification for optimized antibody-fragment expression-secretion in Leishmania tarentolae

    Directory of Open Access Journals (Sweden)

    Klatt Stephan

    2012-07-01

    Full Text Available Abstract Background Secretory signal peptides (SPs are well-known sequence motifs targeting proteins for translocation across the endoplasmic reticulum membrane. After passing through the secretory pathway, most proteins are secreted to the environment. Here, we describe the modification of an expression vector containing the SP from secreted acid phosphatase 1 (SAP1 of Leishmania mexicana for optimized protein expression-secretion in the eukaryotic parasite Leishmania tarentolae with regard to recombinant antibody fragments. For experimental design the online tool SignalP was used, which predicts the presence and location of SPs and their cleavage sites in polypeptides. To evaluate the signal peptide cleavage site as well as changes of expression, SPs were N-terminally linked to single-chain Fragment variables (scFv’s. The ability of L. tarentolae to express complex eukaryotic proteins with highly diverse post-translational modifications and its easy bacteria-like handling, makes the parasite a promising expression system for secretory proteins. Results We generated four vectors with different SP-sequence modifications based on in-silico analyses with SignalP in respect to cleavage probability and location, named pLTEX-2 to pLTEX-5. To evaluate their functionality, we cloned four individual scFv-fragments into the vectors and transfected all 16 constructs into L. tarentolae. Independently from the expressed scFv, pLTEX-5 derived constructs showed the highest expression rate, followed by pLTEX-4 and pLTEX-2, whereas only low amounts of protein could be obtained from pLTEX-3 clones, indicating dysfunction of the SP. Next, we analysed the SP cleavage sites by Edman degradation. For pLTEX-2, -4, and -5 derived scFv’s, the results corresponded to in-silico predictions, whereas pLTEX-3 derived scFv’s contained one additional amino-acid (AA. Conclusions The obtained results demonstrate the importance of SP-sequence optimization for efficient

  6. Stability and Application of Reactive Nitrogen and Oxygen Species-Induced Hemoglobin Modifications in Dry Blood Spots As Analyzed by Liquid Chromatography Tandem Mass Spectrometry.

    Science.gov (United States)

    Chen, Hauh-Jyun Candy; Fan, Chih-Huang; Yang, Ya-Fen

    2016-12-19

    Dried blood spot (DBS) is an emerging microsampling technique for the bioanalysis of small molecules, including fatty acids, metabolites, drugs, and toxicants. DBS offers many advantages as a sample format including easy sample collection and cheap sample shipment. Hemoglobin adducts have been recognized as a suitable biomarker for monitoring chemical exposure. We previously reported that certain modified peptides in hemoglobin derived from reactive chlorine, nitrogen, and oxygen species are associated with factors including smoking, diabetes mellitus, and aging. However, the stability of these oxidation-induced modifications of hemoglobin remains unknown and whether they can be formed artifactually during storage of DBS. To answer these questions, globin extracted from the DBS cards was analyzed, and the stability of the modifications was evaluated. After storage of the DBS cards at 4 °C or room temperature up to 7 weeks, we isolated globin from a quarter of the spot every week. The extents of 11 sites and types of post-translational modifications (PTMs), including nitration and nitrosylation of tyrosine and oxidation of cysteine and methionine residues, in human hemoglobin were measured in the trypsin digest by nanoflow liquid chromatography-nanospray ionization tandem mass spectrometry (nanoLC-NSI/MS/MS) using selected reaction monitoring. The extents of all these PTMs are stable within 14 days when stored on DBS at room temperature and at 4 °C, while those from direct extraction of fresh blood are stable for at least 8 weeks when stored as an aqueous solution at -20 °C. Extraction of globin from a DBS card is of particular importance for hemolytic blood samples. To our knowledge, this is the first report on the stability of oxidative modifications of hemoglobin on DBSs, which are stable for 14 days under ambient conditions (room temperature, in air). Therefore, it is feasible and convenient to analyze these hemoglobin modifications from DBSs in studies

  7. SUMO modification of Stra13 is required for repression of cyclin D1 expression and cellular growth arrest.

    Directory of Open Access Journals (Sweden)

    Yaju Wang

    Full Text Available Stra13, a basic helix-loop-helix (bHLH transcription factor is involved in myriad biological functions including cellular growth arrest, differentiation and senescence. However, the mechanisms by which its transcriptional activity and function are regulated remain unclear. In this study, we provide evidence that post-translational modification of Stra13 by Small Ubiquitin-like Modifier (SUMO dramatically potentiates its ability to transcriptionally repress cyclin D1 and mediate G(1 cell cycle arrest in fibroblast cells. Mutation of SUMO acceptor lysines 159 and 279 located in the C-terminal repression domain has no impact on nuclear localization; however, it abrogates association with the co-repressor histone deacetylase 1 (HDAC1, attenuates repression of cyclin D1, and prevents Stra13-mediated growth suppression. HDAC1, which promotes cellular proliferation and cell cycle progression, antagonizes Stra13 sumoylation-dependent growth arrest. Our results uncover an unidentified regulatory axis between Stra13 and HDAC1 in progression through the G(1/S phase of the cell cycle, and provide new mechanistic insights into regulation of Stra13-mediated transcriptional repression by sumoylation.

  8. Differential patterns of histone acetylation in inflammatory bowel diseases

    Directory of Open Access Journals (Sweden)

    Adcock Ian M

    2011-01-01

    Full Text Available Abstract Post-translational modifications of histones, particularly acetylation, are associated with the regulation of inflammatory gene expression. We used two animal models of inflammation of the bowel and biopsy samples from patients with Crohn's disease (CD to study the expression of acetylated histones (H 3 and 4 in inflamed mucosa. Acetylation of histone H4 was significantly elevated in the inflamed mucosa in the trinitrobenzene sulfonic acid model of colitis particularly on lysine residues (K 8 and 12 in contrast to non-inflamed tissue. In addition, acetylated H4 was localised to inflamed tissue and to Peyer's patches (PP in dextran sulfate sodium (DSS-treated rat models. Within the PP, H3 acetylation was detected in the mantle zone whereas H4 acetylation was seen in both the periphery and the germinal centre. Finally, acetylation of H4 was significantly upregulated in inflamed biopsies and PP from patients with CD. Enhanced acetylation of H4K5 and K16 was seen in the PP. These results demonstrate that histone acetylation is associated with inflammation and may provide a novel therapeutic target for mucosal inflammation.

  9. Polyhedral patterns

    KAUST Repository

    Jiang, Caigui; Tang, Chengcheng; Vaxman, Amir; Wonka, Peter; Pottmann, Helmut

    2015-01-01

    We study the design and optimization of polyhedral patterns, which are patterns of planar polygonal faces on freeform surfaces. Working with polyhedral patterns is desirable in architectural geometry and industrial design. However, the classical

  10. Structural dynamic modifications via models

    Indian Academy of Sciences (India)

    The study shows that as many as half of the matrix ... the dynamicist's analytical modelling skill which would appear both in the numerator as. Figure 2. ..... Brandon J A 1990 Strategies for structural dynamic modification (New York: John Wiley).

  11. Modifications to Replacement Costs System

    International Nuclear Information System (INIS)

    Godec, M.

    1989-01-01

    The purpose of this memorandum is to document the improvements and modifications made to the Replacement Costs of Crude Oil (REPCO) Supply Analysis System. While some of this work was performed under our previous support contract to DOE/ASFE, we are presenting all modifications and improvements are presented here for completeness. The memo primarily documents revisions made to the Lower-48 Onshore Model. Revisions and modifications made to other components and models in the REPCO system which are documented elsewhere are only highlighted in this memo. Generally, the modifications made to the Lower-48 Onshore Model reflect changes that have occurred in domestic drilling, oil field costs, and reserves since 1982, the date of the most recent available data used for the original Replacement Costs report, published in 1985

  12. Standard approach to plant modifications

    International Nuclear Information System (INIS)

    Mecredy, R.C.

    1988-01-01

    Organizational and management approaches to the design, installation, and turnover of nuclear plant modifications have changed dramatically in the last 10 to 15 yr. In response to these changes, organizational and individual responsibilities have been defined and management systems have been established at Rochester Gas and Electric (RG and E) Corporation to ensure that high-quality plant modifications are installed in a timely manner that satisfies user needs at minimal cost

  13. Protein-RNA linkage and posttranslational modifications of feline calicivirus and murine norovirus VPg proteins

    Directory of Open Access Journals (Sweden)

    Allan Olspert

    2016-06-01

    Full Text Available Members of the Caliciviridae family of positive sense RNA viruses cause a wide range of diseases in both humans and animals. The detailed characterization of the calicivirus life cycle had been hampered due to the lack of robust cell culture systems and experimental tools for many of the members of the family. However, a number of caliciviruses replicate efficiently in cell culture and have robust reverse genetics systems available, most notably feline calicivirus (FCV and murine norovirus (MNV. These are therefore widely used as representative members with which to examine the mechanistic details of calicivirus genome translation and replication. The replication of the calicivirus RNA genome occurs via a double-stranded RNA intermediate that is then used as a template for the production of new positive sense viral RNA, which is covalently linked to the virus-encoded protein VPg. The covalent linkage to VPg occurs during genome replication via the nucleotidylylation activity of the viral RNA-dependent RNA polymerase. Using FCV and MNV, we used mass spectrometry-based approach to identify the specific amino acid linked to the 5′ end of the viral nucleic acid. We observed that both VPg proteins are covalently linked to guanosine diphosphate (GDP moieties via tyrosine positions 24 and 26 for FCV and MNV respectively. These data fit with previous observations indicating that mutations introduced into these specific amino acids are deleterious for viral replication and fail to produce infectious virus. In addition, we also detected serine phosphorylation sites within the FCV VPg protein with positions 80 and 107 found consistently phosphorylated on VPg-linked viral RNA isolated from infected cells. This work provides the first direct experimental characterization of the linkage of infectious calicivirus viral RNA to the VPg protein and highlights that post-translational modifications of VPg may also occur during the viral life cycle.

  14. Modification of dendritic development.

    Science.gov (United States)

    Feria-Velasco, Alfredo; del Angel, Alma Rosa; Gonzalez-Burgos, Ignacio

    2002-01-01

    Since 1890 Ramón y Cajal strongly defended the theory that dendrites and their processes and spines had a function of not just nutrient transport to the cell body, but they had an important conductive role in neural impulse transmission. He extensively discussed and supported this theory in the Volume 1 of his extraordinary book Textura del Sistema Nervioso del Hombre y de los Vertebrados. Also, Don Santiago significantly contributed to a detailed description of the various neural components of the hippocampus and cerebral cortex during development. Extensive investigation has been done in the last Century related to the functional role of these complex brain regions, and their association with learning, memory and some limbic functions. Likewise, the organization and expression of neuropsychological qualities such as memory, exploratory behavior and spatial orientation, among others, depend on the integrity and adequate functional activity of the cerebral cortex and hippocampus. It is known that brain serotonin synthesis and release depend directly and proportionally on the availability of its precursor, tryptophan (TRY). By using a chronic TRY restriction model in rats, we studied their place learning ability in correlation with the dendritic spine density of pyramidal neurons in field CA1 of the hippocampus during postnatal development. We have also reported alterations in the maturation pattern of the ability for spontaneous alternation and task performance evaluating short-term memory, as well as adverse effects on the density of dendritic spines of hippocampal CA1 field pyramidal neurons and on the dendritic arborization and the number of dendritic spines of pyramidal neurons from the third layer of the prefrontal cortex using the same model of TRY restriction. The findings obtained in these studies employing a modified Golgi method, can be interpreted as a trans-synaptic plastic response due to understimulation of serotoninergic receptors located in the

  15. Active hippocampal networks undergo spontaneous synaptic modification.

    Directory of Open Access Journals (Sweden)

    Masako Tsukamoto-Yasui

    Full Text Available The brain is self-writable; as the brain voluntarily adapts itself to a changing environment, the neural circuitry rearranges its functional connectivity by referring to its own activity. How the internal activity modifies synaptic weights is largely unknown, however. Here we report that spontaneous activity causes complex reorganization of synaptic connectivity without any external (or artificial stimuli. Under physiologically relevant ionic conditions, CA3 pyramidal cells in hippocampal slices displayed spontaneous spikes with bistable slow oscillations of membrane potential, alternating between the so-called UP and DOWN states. The generation of slow oscillations did not require fast synaptic transmission, but their patterns were coordinated by local circuit activity. In the course of generating spontaneous activity, individual neurons acquired bidirectional long-lasting synaptic modification. The spontaneous synaptic plasticity depended on a rise in intracellular calcium concentrations of postsynaptic cells, but not on NMDA receptor activity. The direction and amount of the plasticity varied depending on slow oscillation patterns and synapse locations, and thus, they were diverse in a network. Once this global synaptic refinement occurred, the same neurons now displayed different patterns of spontaneous activity, which in turn exhibited different levels of synaptic plasticity. Thus, active networks continuously update their internal states through ongoing synaptic plasticity. With computational simulations, we suggest that with this slow oscillation-induced plasticity, a recurrent network converges on a more specific state, compared to that with spike timing-dependent plasticity alone.

  16. Epigenetic Modifications and Diabetic Retinopathy

    Directory of Open Access Journals (Sweden)

    Renu A. Kowluru

    2013-01-01

    Full Text Available Diabetic retinopathy remains one of the most debilitating chronic complications, but despite extensive research in the field, the exact mechanism(s responsible for how retina is damaged in diabetes remains ambiguous. Many metabolic pathways have been implicated in its development, and genes associated with these pathways are altered. Diabetic environment also facilitates epigenetics modifications, which can alter the gene expression without permanent changes in DNA sequence. The role of epigenetics in diabetic retinopathy is now an emerging area, and recent work has shown that genes encoding mitochondrial superoxide dismutase (Sod2 and matrix metalloproteinase-9 (MMP-9 are epigenetically modified, activates of epigenetic modification enzymes, histone lysine demethylase 1 (LSD1, and DNA methyltransferase are increased, and the micro RNAs responsible for regulating nuclear transcriptional factor and VEGF are upregulated. With the growing evidence of epigenetic modifications in diabetic retinopathy, better understanding of these modifications has potential to identify novel targets to inhibit this devastating disease. Fortunately, the inhibitors and mimics targeted towards histone modification, DNA methylation, and miRNAs are now being tried for cancer and other chronic diseases, and better understanding of the role of epigenetics in diabetic retinopathy will open the door for their possible use in combating this blinding disease.

  17. The surface modification of polystyrene

    International Nuclear Information System (INIS)

    Tremlett, C.

    2000-03-01

    Polymers have ideal bulk properties for many applications. However, adhesion to many polymers is poor without surface pretreatment. This can result, for example, in peeling paint and printing, adhesive joint failure and bio-incompatibility. In applications such as painting, printing, adhesive bonding and biocompatibility, various cleaning or surface chemical modifications may be employed. A commodity polymer where pretreatment is sometimes needed is polystyrene. This project investigated, in detail, the effects of a novel method of modification namely mediated electrochemical oxidation (MEO), as a mode of surface modification on polystyrene and a comparison was made with other polymers. The resulting modification was investigated using a range of surface analysis techniques to obtain complementary information. These included, X-ray photoelectron spectroscopy, contact angles, static secondary ion mass spectrometry, atomic force microscopy, chemical derivatization, scanning electron microscopy, attenuated total reflection Fourier Transform infrared spectroscopy and composite lap shear joint testing. It has been shown that MEO modifies the surface of polystyrene introduced oxygen mainly as hydroxyl groups, and a small number of carbonyl groups, that are positioned only on the backbone hydrocarbon chain. This modification improved adhesion, was stable and samples could be stored in aqueous media. The resulting hydroxylation was further derivatized using an amino acid to provide a specialised surface. This was very different from the multiple oxygen functionalities introduced in the comparison studies by UV/ozone and plasma treatments. (author)

  18. Hydroxyl Radical-Mediated Novel Modification of Peptides: N-Terminal Cyclization through the Formation of α-Ketoamide.

    Science.gov (United States)

    Lee, Seon Hwa; Kyung, Hyunsook; Yokota, Ryo; Goto, Takaaki; Oe, Tomoyuki

    2015-01-20

    The hydroxyl radical-mediated oxidation of peptides and proteins constitutes a large group of post-translational modifications that can result in structural and functional changes. These oxidations can lead to hydroxylation, sulfoxidation, or carbonylation of certain amino acid residues and cleavage of peptide bonds. In addition, hydroxyl radicals can convert the N-terminus of peptides to an α-ketoamide via abstraction of the N-terminal α-hydrogen and hydrolysis of the ketimine intermediate. In the present study, we identified N-terminal cyclization as a novel modification mediated by a hydroxyl radical. The reaction of angiotensin (Ang) II (DRVYIHPF) and the hydroxyl radical generated by the Cu(II)/ascorbic acid (AA) system or UV/hydrogen peroxide system produced N-terminal cyclized-Ang II (Ang C) and pyruvamide-Ang II (Ang P, CH3COCONH-RVYIHPF). The structure of Ang C was confirmed by mass spectrometry and comparison to an authentic standard. The subsequent incubation of isolated Ang P in the presence of Cu(II)/AA revealed that Ang P was the direct precursor of Ang C. The proposed mechanism involves the formation of a nitrogen-centered (aminyl) radical, which cyclizes to form a five-membered ring containing the alkoxy radical. The subsequent β-scission reaction of the alkoxyl radical results in the cleavage of the terminal CH3CO group. The initial aminyl radical can be stabilized by chelation to the Cu(II) ions. The affinity of Ang C toward the Ang II type 1 receptor was significantly lower than that of Ang II or Ang P. Ang C was not further metabolized by aminopeptidase A, which converts Ang II to Ang III. Hydroxyl radical-mediated N-terminal cyclization was also observed in other Ang peptides containing N-terminal alanine, arginine, valine, and amyloid β 1-11 (DAEFRHDSGYE).

  19. Electron beam modification of vanadium dioxide oscillators

    Energy Technology Data Exchange (ETDEWEB)

    Belyaev, Maksim; Velichko, Andrey; Putrolaynen, Vadim; Perminov, Valentin; Pergament, Alexander [Petrozavodsk State University, Petrozavodsk (Russian Federation)

    2017-03-15

    The paper presents the results of a study of electron-beam modification (EBM) of VO{sub 2}-switch I-V curve threshold parameters and the self-oscillation frequency of a circuit containing such a switching device. EBM in vacuum is reversible and the parameters are restored when exposed to air at pressure of 150 Pa. At EBM with a dose of 3 C cm{sup -2}, the voltages of switching-on (V{sub th}) and off (V{sub h}), as well as the OFF-state resistance R{sub off}, decrease down to 50% of the initial values, and the oscillation frequency increases by 30% at a dose of 0.7 C cm{sup -2}. Features of physics of EBM of an oscillator are outlined considering the contribution of the metal and semiconductor phases of the switching channel. Controlled modification allows EBM forming of switches with preset parameters. Also, it might be used in artificial oscillatory neural networks for pattern recognition based on frequency shift keying. (copyright 2017 WILEY-VCH Verlag GmbH and Co. KGaA, Weinheim)

  20. Minimal modification to tribimaximal mixing

    International Nuclear Information System (INIS)

    He Xiaogang; Zee, A.

    2011-01-01

    We explore some ways of minimally modifying the neutrino mixing matrix from tribimaximal, characterized by introducing at most one mixing angle and a CP violating phase thus extending our earlier work. One minimal modification, motivated to some extent by group theoretic considerations, is a simple case with the elements V α2 of the second column in the mixing matrix equal to 1/√(3). Modifications by keeping one of the columns or one of the rows unchanged from tribimaximal mixing all belong to the class of minimal modification. Some of the cases have interesting experimentally testable consequences. In particular, the T2K and MINOS collaborations have recently reported indications of a nonzero θ 13 . For the cases we consider, the new data sharply constrain the CP violating phase angle δ, with δ close to 0 (in some cases) and π disfavored.

  1. DNA modification by alkylating compounds

    Energy Technology Data Exchange (ETDEWEB)

    Kruglyakova, E.E.

    1985-09-01

    Results are given for research on the physico-chemical properties of alkylating compounds - nitroso alkyl ureas (NAU) which possess a broad spectrum of biological activity, such as mutagenic, carcinogenic, and anti-tumor action that is due to the alkylation and carbamoylation of DNA as well as other cellular components. Identified chemical products of NAU interaction with DNA and its components are cited. Structural conversions of a DNA macromolecule resulting from its chemical modification are examined. NAU are used to discuss possible biological consequences of DNA modification. 148 references.

  2. Corrosion principles and surface modification

    International Nuclear Information System (INIS)

    Kruger, J.

    1982-01-01

    This chapter examines the important strategies provided by the newer ideas of corrosion science and engineering that surface modification techniques must utilize to help prevent corrosion, especially the most damaging kind of aqueous corrosion, localized corrosion. Provides a brief introduction to the principles underlying the phenomenon of corrosion in order to use them to discuss surface modification strategies to combat corrosion. Discusses the electrochemistry of corrosion; the thermodynamics of corrosion; the kinetics of corrosion; thermodynamic strategies; and kinetic strategies (formation of more protective passive films; resistance to breakdown; ductility; repassivation)

  3. Polyhedral patterns

    KAUST Repository

    Jiang, Caigui

    2015-10-27

    We study the design and optimization of polyhedral patterns, which are patterns of planar polygonal faces on freeform surfaces. Working with polyhedral patterns is desirable in architectural geometry and industrial design. However, the classical tiling patterns on the plane must take on various shapes in order to faithfully and feasibly approximate curved surfaces. We define and analyze the deformations these tiles must undertake to account for curvature, and discover the symmetries that remain invariant under such deformations. We propose a novel method to regularize polyhedral patterns while maintaining these symmetries into a plethora of aesthetic and feasible patterns.

  4. Genetic analysis of the heparan modification network in Caenorhabditis elegans.

    Science.gov (United States)

    Townley, Robert A; Bülow, Hannes E

    2011-05-13

    Heparan sulfates (HS) are highly modified sugar polymers in multicellular organisms that function in cell adhesion and cellular responses to protein signaling. Functionally distinct, cell type-dependent HS modification patterns arise as the result of a conserved network of enzymes that catalyze deacetylations, sulfations, and epimerizations in specific positions of the sugar residues. To understand the genetic interactions of the enzymes during the HS modification process, we have measured the composition of HS purified from mutant strains of Caenorhabditis elegans. From these measurements we have developed a genetic network model of HS modification. We find the interactions to be highly recursive positive feed-forward and negative feedback loops. Our genetic analyses show that the HS C-5 epimerase hse-5, the HS 2-O-sulfotransferase hst-2, or the HS 6-O-sulfotransferase hst-6 inhibit N-sulfation. In contrast, hse-5 stimulates both 2-O- and 6-O-sulfation and, hst-2 and hst-6 inhibit 6-O- and 2-O-sulfation, respectively. The effects of hst-2 and hst-6 on N-sulfation, 6-O-sulfation, and 2-O-sulfation appear largely dependent on hse-5 function. This core of regulatory interactions is further modulated by 6-O-endosulfatase activity (sul-1). 47% of all 6-O-sulfates get removed from HS and this editing process is dependent on hst-2, thereby providing additional negative feedback between 2-O- and 6-O-sulfation. These findings suggest that the modification patterns are highly sensitive to the relative composition of the HS modification enzymes. Our comprehensive genetic analysis forms the basis of understanding the HS modification network in metazoans.

  5. Genetic Analysis of the Heparan Modification Network in Caenorhabditis elegans*

    Science.gov (United States)

    Townley, Robert A.; Bülow, Hannes E.

    2011-01-01

    Heparan sulfates (HS) are highly modified sugar polymers in multicellular organisms that function in cell adhesion and cellular responses to protein signaling. Functionally distinct, cell type-dependent HS modification patterns arise as the result of a conserved network of enzymes that catalyze deacetylations, sulfations, and epimerizations in specific positions of the sugar residues. To understand the genetic interactions of the enzymes during the HS modification process, we have measured the composition of HS purified from mutant strains of Caenorhabditis elegans. From these measurements we have developed a genetic network model of HS modification. We find the interactions to be highly recursive positive feed-forward and negative feedback loops. Our genetic analyses show that the HS C-5 epimerase hse-5, the HS 2-O-sulfotransferase hst-2, or the HS 6-O-sulfotransferase hst-6 inhibit N-sulfation. In contrast, hse-5 stimulates both 2-O- and 6-O-sulfation and, hst-2 and hst-6 inhibit 6-O- and 2-O-sulfation, respectively. The effects of hst-2 and hst-6 on N-sulfation, 6-O-sulfation, and 2-O-sulfation appear largely dependent on hse-5 function. This core of regulatory interactions is further modulated by 6-O-endosulfatase activity (sul-1). 47% of all 6-O-sulfates get removed from HS and this editing process is dependent on hst-2, thereby providing additional negative feedback between 2-O- and 6-O-sulfation. These findings suggest that the modification patterns are highly sensitive to the relative composition of the HS modification enzymes. Our comprehensive genetic analysis forms the basis of understanding the HS modification network in metazoans. PMID:21454666

  6. Specialization Patterns

    DEFF Research Database (Denmark)

    Schultz, Ulrik Pagh; Lawall, Julia Laetitia; Consel, Charles

    2000-01-01

    Design patterns offer many advantages for software development, but can introduce inefficiency into the final program. Program specialization can eliminate such overheads, but is most effective when targeted by the user to specific bottlenecks. Consequently, we propose that these concepts...... are complementary. Program specialization can optimize programs written using design patterns, and design patterns provide information about the program structure that can guide specialization. Concretely, we propose specialization patterns, which describe how to apply program specialization to optimize uses...... of design patterns. In this paper, we analyze the specialization opportunities provided by specific uses of design patterns. Based on the analysis of each design pattern, we define the associated specialization pattern. These specialization opportunities can be declared using the specialization classes...

  7. Superhydrophobic cotton by fluorosilane modification

    CSIR Research Space (South Africa)

    Erasmus, E

    2009-12-01

    Full Text Available the treatment with fluorinated or silicon compounds)1-4 and by enhancing the surface roughness with a fractal structure5-8. Cotton, a cellulose-based material, that is greatly hydrophilic, is more benefited when made hydrophobic. Modification of cotton...

  8. Complex exon-intron marking by histone modifications is not determined solely by nucleosome distribution.

    Directory of Open Access Journals (Sweden)

    Pawandeep Dhami

    2010-08-01

    Full Text Available It has recently been shown that nucleosome distribution, histone modifications and RNA polymerase II (Pol II occupancy show preferential association with exons ("exon-intron marking", linking chromatin structure and function to co-transcriptional splicing in a variety of eukaryotes. Previous ChIP-sequencing studies suggested that these marking patterns reflect the nucleosomal landscape. By analyzing ChIP-chip datasets across the human genome in three cell types, we have found that this marking system is far more complex than previously observed. We show here that a range of histone modifications and Pol II are preferentially associated with exons. However, there is noticeable cell-type specificity in the degree of exon marking by histone modifications and, surprisingly, this is also reflected in some histone modifications patterns showing biases towards introns. Exon-intron marking is laid down in the absence of transcription on silent genes, with some marking biases changing or becoming reversed for genes expressed at different levels. Furthermore, the relationship of this marking system with splicing is not simple, with only some histone modifications reflecting exon usage/inclusion, while others mirror patterns of exon exclusion. By examining nucleosomal distributions in all three cell types, we demonstrate that these histone modification patterns cannot solely be accounted for by differences in nucleosome levels between exons and introns. In addition, because of inherent differences between ChIP-chip array and ChIP-sequencing approaches, these platforms report different nucleosome distribution patterns across the human genome. Our findings confound existing views and point to active cellular mechanisms which dynamically regulate histone modification levels and account for exon-intron marking. We believe that these histone modification patterns provide links between chromatin accessibility, Pol II movement and co-transcriptional splicing.

  9. Early cytoskeletal protein modifications precede overt structural degeneration in the DBA/2J mouse model of glaucoma

    Directory of Open Access Journals (Sweden)

    Gina Nicole Wilson

    2016-11-01

    Full Text Available Axonal transport deficits precede structural loss in glaucoma and other neurodegenerations. Impairments in structural support, including modified cytoskeletal proteins and microtubule-destabilizing elements, could be initiating factors in glaucoma pathogenesis. We investigated the time course of changes in protein levels and post-translational modifications in the DBA/2J mouse model of glaucoma. Using anterograde tract tracing of the retinal projection, we assessed major cytoskeletal and transported elements as a function of transport integrity in different stages of pathological progression. Using capillary-based electrophoresis, single- and multiplex immunosorbent assays, and immunofluorescence, we quantified hyperphosphorylated neurofilament-heavy chain, phosphorylated tau (ptau, calpain-mediated spectrin breakdown product (145/150kDa, β –tubulin, and amyloid-β42 proteins based on age and transport outcome to the superior colliculus (SC, the main retinal target in mice. Phosphorylated neurofilament-heavy chain (pNF-H was elevated within the optic nerve (ON and SC of 8-10 month-old DBA/2J mice, but was not evident in the retina until 12-15 months, suggesting that cytoskeletal modifications first appear in the distal retinal projection. As expected, higher pNF-H levels in the SC and retina were correlated with axonal transport deficits. Elevations in hyperphosphorylated tau (ptau occurred in ON and SC between 3-8 month of age while retinal ptau accumulations occurred at 12-15 months in DBA/2J mice. In vitro co-immunoprecipitation experiments suggested increased affinity of ptau for the retrograde motor complex protein, dynactin. We observed a transport-related decrease of β-tubulin in ON of 10-12 month-old DBA/2J mice, suggesting destabilized microtubule array. Elevations in calpain-mediated spectrin breakdown product were seen in ON and SC at the earliest age examined, well before axonal transport loss is evident. Finally, transport

  10. In vivo modification of tyrosine residues in recombinant mussel adhesive protein by tyrosinase co-expression in Escherichia coli

    Directory of Open Access Journals (Sweden)

    Choi Yoo

    2012-10-01

    Full Text Available Abstract Background In nature, mussel adhesive proteins (MAPs show remarkable adhesive properties, biocompatibility, and biodegradability. Thus, they have been considered promising adhesive biomaterials for various biomedical and industrial applications. However, limited production of natural MAPs has hampered their practical applications. Recombinant production in bacterial cells could be one alternative to obtain useable amounts of MAPs, although additional post-translational modification of tyrosine residues into 3,4-dihydroxyphenyl-alanine (Dopa and Dopaquinone is required. The superior properties of MAPs are mainly attributed to the introduction of quinone-derived intermolecular cross-links. To solve this problem, we utilized a co-expression strategy of recombinant MAP and tyrosinase in Escherichia coli to successfully modify tyrosine residues in vivo. Results A recombinant hybrid MAP, fp-151, was used as a target for in vivo modification, and a dual vector system of pET and pACYC-Duet provided co-expression of fp-151 and tyrosinase. As a result, fp-151 was over-expressed and mainly obtained from the soluble fraction in the co-expression system. Without tyrosinase co-expression, fp-151 was over-expressed in an insoluble form in inclusion bodies. The modification of tyrosine residues in the soluble-expressed fp-151 was clearly observed from nitroblue tetrazolium staining and liquid-chromatography-mass/mass spectrometry analyses. The purified, in vivo modified, fp-151 from the co-expression system showed approximately 4-fold higher bulk-scale adhesive strength compared to in vitro tyrosinase-treated fp-151. Conclusion Here, we reported a co-expression system to obtain in vivo modified MAP; additional in vitro tyrosinase modification was not needed to obtain adhesive properties and the in vivo modified MAP showed superior adhesive strength compared to in vitro modified protein. It is expected that this co-expression strategy will accelerate

  11. Robust dynamical pattern formation from a multifunctional minimal genetic circuit

    Directory of Open Access Journals (Sweden)

    Carrera Javier

    2010-04-01

    Full Text Available Abstract Background A practical problem during the analysis of natural networks is their complexity, thus the use of synthetic circuits would allow to unveil the natural mechanisms of operation. Autocatalytic gene regulatory networks play an important role in shaping the development of multicellular organisms, whereas oscillatory circuits are used to control gene expression under variable environments such as the light-dark cycle. Results We propose a new mechanism to generate developmental patterns and oscillations using a minimal number of genes. For this, we design a synthetic gene circuit with an antagonistic self-regulation to study the spatio-temporal control of protein expression. Here, we show that our minimal system can behave as a biological clock or memory, and it exhibites an inherent robustness due to a quorum sensing mechanism. We analyze this property by accounting for molecular noise in an heterogeneous population. We also show how the period of the oscillations is tunable by environmental signals, and we study the bifurcations of the system by constructing different phase diagrams. Conclusions As this minimal circuit is based on a single transcriptional unit, it provides a new mechanism based on post-translational interactions to generate targeted spatio-temporal behavior.

  12. Ion bombardment modification of surfaces

    International Nuclear Information System (INIS)

    Auciello, O.

    1984-01-01

    An historical overview of the main advances in the understanding of bombardment-induced surface topography is presented. The implantation and sputtering mechanisms which are relevant to ion bombardment modification of surfaces and consequent structural, electronic and compositional changes are described. Descriptions of plasma and ion-beam sputtering-induced film formation, primary ion-beam deposition, dual beam techniques, cluster of molecule ion-beam deposition, and modification of thin film properties by ion bombardment during deposition are presented. A detailed account is given of the analytical and computational modelling of topography from the viewpoint of first erosion theory. Finally, an account of the possible application and/or importance of textured surfaces in technologies and/or experimental techniques not considered in previous chapters is presented. refs.; figs.; tabs

  13. Epigenetic modifications in prostate cancer.

    Science.gov (United States)

    Ngollo, Marjolaine; Dagdemir, Aslihan; Karsli-Ceppioglu, Seher; Judes, Gaelle; Pajon, Amaury; Penault-Llorca, Frederique; Boiteux, Jean-Paul; Bignon, Yves-Jean; Guy, Laurent; Bernard-Gallon, Dominique J

    2014-01-01

    Prostate cancer is the most common cancer in men and the second leading cause of cancer deaths in men in France. Apart from the genetic alterations in prostate cancer, epigenetics modifications are involved in the development and progression of this disease. Epigenetic events are the main cause in gene regulation and the three most epigenetic mechanisms studied include DNA methylation, histone modifications and microRNA expression. In this review, we summarized epigenetic mechanisms in prostate cancer. Epigenetic drugs that inhibit DNA methylation, histone methylation and histone acetylation might be able to reactivate silenced gene expression in prostate cancer. However, further understanding of interactions of these enzymes and their effects on transcription regulation in prostate cancer is needed and has become a priority in biomedical research. In this study, we summed up epigenetic changes with emphasis on pharmacologic epigenetic target agents.

  14. Cradle modification for hydraulic ram

    International Nuclear Information System (INIS)

    Koons, B.M.

    1995-01-01

    The analysis of the cradle hydraulic system considers stress, weld strength, and hydraulic forces required to lift and support the cradle/pump assembly. The stress and weld strength of the cradle modifications is evaluated to ensure that they meet the requirements of the American Institute for Steel Construction (AISC 1989). The hydraulic forces are evaluated to ensure that the hydraulic system is capable of rotating the cradle and pump assembly to the vertical position (between 70 degrees and 90 degrees)

  15. MARCC (Matrix-Assisted Reader Chromatin Capture): an antibody-free method to enrich and analyze combinatorial nucleosome modifications

    Science.gov (United States)

    Su, Zhangli

    2016-01-01

    Combinatorial patterns of histone modifications are key indicators of different chromatin states. Most of the current approaches rely on the usage of antibodies to analyze combinatorial histone modifications. Here we detail an antibody-free method named MARCC (Matrix-Assisted Reader Chromatin Capture) to enrich combinatorial histone modifications. The combinatorial patterns are enriched on native nucleosomes extracted from cultured mammalian cells and prepared by micrococcal nuclease digestion. Such enrichment is achieved by recombinant chromatin-interacting protein modules, or so-called reader domains, which can bind in a combinatorial modification-dependent manner. The enriched chromatin can be quantified by western blotting or mass spectrometry for the co-existence of histone modifications, while the associated DNA content can be analyzed by qPCR or next-generation sequencing. Altogether, MARCC provides a reproducible, efficient and customizable solution to enrich and analyze combinatorial histone modifications. PMID:26131849

  16. Ion bombardment modification of surfaces

    International Nuclear Information System (INIS)

    Auciello, O.

    1984-01-01

    Ion bombardment-induced modification of surfaces may be considered one of the significant scientific and technological developments of the last two decades. The understanding acquired concerning the underlying mechanisms of several phenomena occurring during ion-surface interactions has led to applications within different modern technologies. These include microelectronics, surface acoustical and optical technologies, solar energy conversion, thin film technology, ion implantation metallurgy, nuclear track technology, thermonuclear fusion, vacuum technology, cold welding technology, biomedicine (implantology). It has become clear that information on many relevant advances, regarding ion bombardment modification of surfaces is dispersed among journals involving fields sometimes not clearly related. This may result, in some cases, in a loss of the type of interdisciplinary exchange of ideas, which has proved to be so fruitful for the advancement of science and technology. This book has been planned in an attempt to collect at least some of today's relevant information about the experimental and theoretical knowledge related to surface modification and its application to technology. (Auth.)

  17. Specialization Patterns

    OpenAIRE

    Schultz , Ulrik Pagh; Lawall , Julia ,; Consel , Charles

    1999-01-01

    Design patterns offer numerous advantages for software development, but can introduce inefficiency into the finished program. Program specialization can eliminate such overheads, but is most effective when targeted by the user to specific bottlenecks. Consequently, we propose to consider program specialization and design patterns as complementary concepts. On the one hand, program specialization can optimize object-oriented programs written using design patterns. On the other hand, design pat...

  18. Pattern recognition

    CERN Document Server

    Theodoridis, Sergios

    2003-01-01

    Pattern recognition is a scientific discipline that is becoming increasingly important in the age of automation and information handling and retrieval. Patter Recognition, 2e covers the entire spectrum of pattern recognition applications, from image analysis to speech recognition and communications. This book presents cutting-edge material on neural networks, - a set of linked microprocessors that can form associations and uses pattern recognition to ""learn"" -and enhances student motivation by approaching pattern recognition from the designer's point of view. A direct result of more than 10

  19. Modification of P-selectin glycoprotein ligand-1 with a natural killer cell-restricted sulfated lactosamine creates an alternate ligand for L-selectin

    Science.gov (United States)

    André, Pascale; Spertini, Olivier; Guia, Sophie; Rihet, Pascal; Dignat-George, Françoise; Brailly, Hervé; Sampol, José; Anderson, Paul J.; Vivier, Eric

    2000-01-01

    Natural killer (NK) cells are components of the innate immune system that can recognize and kill virally infected cells, tumor cells, and allogeneic cells without prior sensitization. NK cells also elaborate cytokines (e.g., interferon-γ and tumor necrosis factor-α) and chemokines (e.g., macrophage inflammatory protein-1α) that promote the acquisition of antigen-specific immunity. NK cell differentiation is accompanied by the cell surface expression of a mucin-like glycoprotein bearing an NK cell-restricted keratan sulfate-related lactosamine carbohydrate, the PEN5 epitope. Here, we report that PEN5 is a post-translational modification of P-selectin glycoprotein ligand-1 (PSGL-1). The PEN5 epitope creates on PSGL-1 a unique binding site for L-selectin, which is independent of PSGL-1 tyrosine sulfation. On the surface of NK cells, the expression of PEN5 is coordinated with the disappearance of L-selectin and the up-regulation of Killer cell Ig-like Receptors (KIR). These results indicate that NK cell differentiation is accompanied by the acquisition of a unique carbohydrate, PEN5, that can serve as part of a combination code to deliver KIR+ NK cells to specific tissues. PMID:10725346

  20. Sleep modifications in acute transient global amnesia.

    Science.gov (United States)

    Della Marca, Giacomo; Mazza, Marianna; Losurdo, Anna; Testani, Elisa; Broccolini, Aldobrando; Frisullo, Giovanni; Marano, Giuseppe; Morosetti, Roberta; Pilato, Fabio; Profice, Paolo; Vollono, Catello; Di Lazzaro, Vincenzo

    2013-09-15

    Transient global amnesia (TGA) is a temporary memory loss characterized by an abrupt onset of antero-grade and retrograde amnesia, totally reversible. Since sleep plays a major role in memory consolidation, and in the storage of memory-related traces into the brain cortex, the aims of the present study were: (1) to evaluate changes in sleep macro-structure in TGA; (2) to assess modifications in sleep micro-structure in TGA, with particular reference to the arousal EEG and to cyclic alternating pattern (CAP); (3) to compare sleep parameters in TGA patients with a control group of patients with acute ischemic events ("minor stroke" or transient ischemic attack [TIA]) clinically and neuroradiologically "similar" to the TGA. TGA GROUP: 17 patients, (8 men and 9 women, 60.2 ± 12.5 years). Stroke or TIA (SoT) group: 17 patients hospitalized in the Stroke Unit for recent onset of minor stroke or TIA with hemispheric localization; healthy controls (HC) group: 17 healthy volunteers, matched for age and sex. Patients and controls underwent full-night polysomnography. In the multivariate analysis (conditions TGA, SoT, and HC) a significant effect of the condition was observed for sleep efficiency index, number of awakenings longer 1 min, REM latency, CAP time, and CAP rate. TGA and SoT differed only for CAP time and CAP rate, which were lower in the TGA group. Microstructural modification associated with tga could be consequent to: (1) hippocampal dysfunction and memory impairment; (2) impairment of arousal-related structures (in particular, cholinergic pathways); (3) emotional distress.

  1. Histone modifications: Cycling with chromosomal replication

    DEFF Research Database (Denmark)

    Thon, Genevieve

    2008-01-01

    Histone modifications tend to be lost during chromosome duplication. Several recent studies suggest that the RNA interference pathway becomes active during the weakened transcriptional repression occurring at centromeres in S phase, resulting in the re-establishment of histone modifications...

  2. BIOCHAR MODIFICATION, THERMAL STABILITY AND TOXICITY OF PRODUCTS MODIFICATION

    Directory of Open Access Journals (Sweden)

    Romana FRIEDRICHOVÁ

    2017-12-01

    Full Text Available Biochar is a product obtained from processing of waste biomass. The main application of biochar is in soil and environment remediation. Some new applications of this carbonaceous material take advantage of its adsorption capacity use it as a heterogeneous catalyst for energy storage and conversion etc. This contribution describes thermal stability of the original biochar. It discusses biochar modified by chemical and physical methods including a new compound of biochar-graphene oxide. The purpose of the modifications is to increase its active surface to introduce active functional groups into the carbon structure of biochar in relation to fire safety and toxicity of those products.

  3. RNomics and Modomics in the halophilic archaea Haloferax volcanii: identification of RNA modification genes

    Directory of Open Access Journals (Sweden)

    Decatur Wayne A

    2008-10-01

    Full Text Available Abstract Background Naturally occurring RNAs contain numerous enzymatically altered nucleosides. Differences in RNA populations (RNomics and pattern of RNA modifications (Modomics depends on the organism analyzed and are two of the criteria that distinguish the three kingdoms of life. If the genomic sequences of the RNA molecules can be derived from whole genome sequence information, the modification profile cannot and requires or direct sequencing of the RNAs or predictive methods base on the presence or absence of the modifications genes. Results By employing a comparative genomics approach, we predicted almost all of the genes coding for the t+rRNA modification enzymes in the mesophilic moderate halophile Haloferax volcanii. These encode both guide RNAs and enzymes. Some are orthologous to previously identified genes in Archaea, Bacteria or in Saccharomyces cerevisiae, but several are original predictions. Conclusion The number of modifications in t+rRNAs in the halophilic archaeon is surprisingly low when compared with other Archaea or Bacteria, particularly the hyperthermophilic organisms. This may result from the specific lifestyle of halophiles that require high intracellular salt concentration for survival. This salt content could allow RNA to maintain its functional structural integrity with fewer modifications. We predict that the few modifications present must be particularly important for decoding, accuracy of translation or are modifications that cannot be functionally replaced by the electrostatic interactions provided by the surrounding salt-ions. This analysis also guides future experimental validation work aiming to complete the understanding of the function of RNA modifications in Archaeal translation.

  4. Fabrication and Modification of Nanoporous Silicon Particles

    Science.gov (United States)

    Ferrari, Mauro; Liu, Xuewu

    2010-01-01

    Silicon-based nanoporous particles as biodegradable drug carriers are advantageous in permeation, controlled release, and targeting. The use of biodegradable nanoporous silicon and silicon dioxide, with proper surface treatments, allows sustained drug release within the target site over a period of days, or even weeks, due to selective surface coating. A variety of surface treatment protocols are available for silicon-based particles to be stabilized, functionalized, or modified as required. Coated polyethylene glycol (PEG) chains showed the effective depression of both plasma protein adsorption and cell attachment to the modified surfaces, as well as the advantage of long circulating. Porous silicon particles are micromachined by lithography. Compared to the synthesis route of the nanomaterials, the advantages include: (1) the capability to make different shapes, not only spherical particles but also square, rectangular, or ellipse cross sections, etc.; (2) the capability for very precise dimension control; (3) the capacity for porosity and pore profile control; and (4) allowance of complex surface modification. The particle patterns as small as 60 nm can be fabricated using the state-of-the-art photolithography. The pores in silicon can be fabricated by exposing the silicon in an HF/ethanol solution and then subjecting the pores to an electrical current. The size and shape of the pores inside silicon can be adjusted by the doping of the silicon, electrical current application, the composition of the electrolyte solution, and etching time. The surface of the silicon particles can be modified by many means to provide targeted delivery and on-site permanence for extended release. Multiple active agents can be co-loaded into the particles. Because the surface modification of particles can be done on wafers before the mechanical release, asymmetrical surface modification is feasible. Starting from silicon wafers, a treatment, such as KOH dipping or reactive ion

  5. Energy conservation potential of surface modification technologies

    Energy Technology Data Exchange (ETDEWEB)

    Le, H.K.; Horne, D.M.; Silberglitt, R.S.

    1985-09-01

    This report assesses the energy conservation impact of surface modification technologies on the metalworking industries. The energy conservation impact of surface modification technologies on the metalworking industries is assessed by estimating their friction and wear tribological sinks and the subsequent reduction in these sinks when surface modified tools are used. Ion implantation, coatings, and laser and electron beam surface modifications are considered.

  6. Geodesic patterns

    KAUST Repository

    Pottmann, Helmut; Huang, Qixing; Deng, Bailin; Schiftner, Alexander; Kilian, Martin; Guibas, Leonidas J.; Wallner, Johannes

    2010-01-01

    Geodesic curves in surfaces are not only minimizers of distance, but they are also the curves of zero geodesic (sideways) curvature. It turns out that this property makes patterns of geodesics the basic geometric entity when dealing with the cladding of a freeform surface with wooden panels which do not bend sideways. Likewise a geodesic is the favored shape of timber support elements in freeform architecture, for reasons of manufacturing and statics. Both problem areas are fundamental in freeform architecture, but so far only experimental solutions have been available. This paper provides a systematic treatment and shows how to design geodesic patterns in different ways: The evolution of geodesic curves is good for local studies and simple patterns; the level set formulation can deal with the global layout of multiple patterns of geodesics; finally geodesic vector fields allow us to interactively model geodesic patterns and perform surface segmentation into panelizable parts. © 2010 ACM.

  7. Foam patterns

    Science.gov (United States)

    Chaudhry, Anil R; Dzugan, Robert; Harrington, Richard M; Neece, Faurice D; Singh, Nipendra P; Westendorf, Travis

    2013-11-26

    A method of creating a foam pattern comprises mixing a polyol component and an isocyanate component to form a liquid mixture. The method further comprises placing a temporary core having a shape corresponding to a desired internal feature in a cavity of a mold and inserting the mixture into the cavity of the mold so that the mixture surrounds a portion of the temporary core. The method optionally further comprises using supporting pins made of foam to support the core in the mold cavity, with such pins becoming integral part of the pattern material simplifying subsequent processing. The method further comprises waiting for a predetermined time sufficient for a reaction from the mixture to form a foam pattern structure corresponding to the cavity of the mold, wherein the foam pattern structure encloses a portion of the temporary core and removing the temporary core from the pattern independent of chemical leaching.

  8. Geodesic patterns

    KAUST Repository

    Pottmann, Helmut

    2010-07-26

    Geodesic curves in surfaces are not only minimizers of distance, but they are also the curves of zero geodesic (sideways) curvature. It turns out that this property makes patterns of geodesics the basic geometric entity when dealing with the cladding of a freeform surface with wooden panels which do not bend sideways. Likewise a geodesic is the favored shape of timber support elements in freeform architecture, for reasons of manufacturing and statics. Both problem areas are fundamental in freeform architecture, but so far only experimental solutions have been available. This paper provides a systematic treatment and shows how to design geodesic patterns in different ways: The evolution of geodesic curves is good for local studies and simple patterns; the level set formulation can deal with the global layout of multiple patterns of geodesics; finally geodesic vector fields allow us to interactively model geodesic patterns and perform surface segmentation into panelizable parts. © 2010 ACM.

  9. Epigenetic modifications and diabetic nephropathy

    Directory of Open Access Journals (Sweden)

    Marpadga A. Reddy

    2012-09-01

    Full Text Available Diabetic nephropathy (DN is a major complication associated with both type 1 and type 2 diabetes, and a leading cause of end-stage renal disease. Conventional therapeutic strategies are not fully efficacious in the treatment of DN, suggesting an incomplete understanding of the gene regulation mechanisms involved in its pathogenesis. Furthermore, evidence from clinical trials has demonstrated a “metabolic memory” of prior exposure to hyperglycemia that continues to persist despite subsequent glycemic control. This remains a major challenge in the treatment of DN and other vascular complications. Epigenetic mechanisms such as DNA methylation, nucleosomal histone modifications, and noncoding RNAs control gene expression through regulation of chromatin structure and function and post-transcriptional mechanisms without altering the underlying DNA sequence. Emerging evidence indicates that multiple factors involved in the etiology of diabetes can alter epigenetic mechanisms and regulate the susceptibility to diabetes complications. Recent studies have demonstrated the involvement of histone lysine methylation in the regulation of key fibrotic and inflammatory genes related to diabetes complications including DN. Interestingly, histone lysine methylation persisted in vascular cells even after withdrawal from the diabetic milieu, demonstrating a potential role of epigenetic modifications in metabolic memory. Rapid advances in high-throughput technologies in the fields of genomics and epigenomics can lead to the identification of genome-wide alterations in key epigenetic modifications in vascular and renal cells in diabetes. Altogether, these findings can lead to the identification of potential predictive biomarkers and development of novel epigenetic therapies for diabetes and its associated complications.

  10. Modification Propagation in Complex Networks

    Science.gov (United States)

    Mouronte, Mary Luz; Vargas, María Luisa; Moyano, Luis Gregorio; Algarra, Francisco Javier García; Del Pozo, Luis Salvador

    To keep up with rapidly changing conditions, business systems and their associated networks are growing increasingly intricate as never before. By doing this, network management and operation costs not only rise, but are difficult even to measure. This fact must be regarded as a major constraint to system optimization initiatives, as well as a setback to derived economic benefits. In this work we introduce a simple model in order to estimate the relative cost associated to modification propagation in complex architectures. Our model can be used to anticipate costs caused by network evolution, as well as for planning and evaluating future architecture development while providing benefit optimization.

  11. Postulated weather modification effects of large energy releases

    Energy Technology Data Exchange (ETDEWEB)

    Ramsdell, J.V.; Scott, B.C.; Orgill, M.M.; Renne, D.S.; Hubbard, J.E.; McGinnis, K.A.

    1977-02-01

    Postulated impacts of large energy releases were examined in the light of existing technical information. The magnitudes of direct atmospheric modifications were estimated, and the ecological and economic implications of the modifications were explored. Energy releases from energy centers (10 to 40 power plants at a single site) and individual power plant clusters (1 to 4 power plants) were considered. In the atmosphere the energy will exist initially as increased temperature (sensible heat), moisture (latent heat), and air motion (kinetic energy). Addition of energy could result in increased cloudiness and fog, and changed precipitation patterns. A framework for economic analysis of the impacts of the postulated atmospheric modifications was established on the basis of costs and benefits. Willingness-to-pay was selected as the appropriate measure for valuing each impact. The primary and secondary atmospheric modifications may affect recreation, transportation, and aesthetics as well as agriculture and forestry. Economic values can be placed on some of the effects. However, the willingness of people to pay to gain benefits and avoid damages in many cases can only be determined through extensive surveys. The economic consequences of a given energy release would be highly site specific.

  12. Postulated weather modification effects of large energy releases

    International Nuclear Information System (INIS)

    Ramsdell, J.V.; Scott, B.C.; Orgill, M.M.; Renne, D.S.; Hubbard, J.E.; McGinnis, K.A.

    1977-02-01

    Postulated impacts of large energy releases were examined in the light of existing technical information. The magnitudes of direct atmospheric modifications were estimated, and the ecological and economic implications of the modifications were explored. Energy releases from energy centers (10 to 40 power plants at a single site) and individual power plant clusters (1 to 4 power plants) were considered. In the atmosphere the energy will exist initially as increased temperature (sensible heat), moisture (latent heat), and air motion (kinetic energy). Addition of energy could result in increased cloudiness and fog, and changed precipitation patterns. A framework for economic analysis of the impacts of the postulated atmospheric modifications was established on the basis of costs and benefits. Willingness-to-pay was selected as the appropriate measure for valuing each impact. The primary and secondary atmospheric modifications may affect recreation, transportation, and aesthetics as well as agriculture and forestry. Economic values can be placed on some of the effects. However, the willingness of people to pay to gain benefits and avoid damages in many cases can only be determined through extensive surveys. The economic consequences of a given energy release would be highly site specific

  13. Structure-Function, Stability, and Chemical Modification of the Cyanobacterial Cytochrome b6f Complex from Nostoc sp. PCC 7120*

    Science.gov (United States)

    Baniulis, Danas; Yamashita, Eiki; Whitelegge, Julian P.; Zatsman, Anna I.; Hendrich, Michael P.; Hasan, S. Saif; Ryan, Christopher M.; Cramer, William A.

    2009-01-01

    The crystal structure of the cyanobacterial cytochrome b6f complex has previously been solved to 3.0-Å resolution using the thermophilic Mastigocladus laminosus whose genome has not been sequenced. Several unicellular cyanobacteria, whose genomes have been sequenced and are tractable for mutagenesis, do not yield b6f complex in an intact dimeric state with significant electron transport activity. The genome of Nostoc sp. PCC 7120 has been sequenced and is closer phylogenetically to M. laminosus than are unicellular cyanobacteria. The amino acid sequences of the large core subunits and four small peripheral subunits of Nostoc are 88 and 80% identical to those in the M. laminosus b6f complex. Purified b6f complex from Nostoc has a stable dimeric structure, eight subunits with masses similar to those of M. laminosus, and comparable electron transport activity. The crystal structure of the native b6f complex, determined to a resolution of 3.0Å (PDB id: 2ZT9), is almost identical to that of M. laminosus. Two unique aspects of the Nostoc complex are: (i) a dominant conformation of heme bp that is rotated 180° about the α- and γ-meso carbon axis relative to the orientation in the M. laminosus complex and (ii) acetylation of the Rieske iron-sulfur protein (PetC) at the N terminus, a post-translational modification unprecedented in cyanobacterial membrane and electron transport proteins, and in polypeptides of cytochrome bc complexes from any source. The high spin electronic character of the unique heme cn is similar to that previously found in the b6f complex from other sources. PMID:19189962

  14. Search Patterns

    CERN Document Server

    Morville, Peter

    2010-01-01

    What people are saying about Search Patterns "Search Patterns is a delight to read -- very thoughtful and thought provoking. It's the most comprehensive survey of designing effective search experiences I've seen." --Irene Au, Director of User Experience, Google "I love this book! Thanks to Peter and Jeffery, I now know that search (yes, boring old yucky who cares search) is one of the coolest ways around of looking at the world." --Dan Roam, author, The Back of the Napkin (Portfolio Hardcover) "Search Patterns is a playful guide to the practical concerns of search interface design. It cont

  15. The effectiveness of workplace dietary modification interventions: a systematic review.

    Science.gov (United States)

    Geaney, F; Kelly, C; Greiner, B A; Harrington, J M; Perry, I J; Beirne, P

    2013-11-01

    To evaluate the effectiveness of workplace dietary modification interventions alone or in combination with nutrition education on employees' dietary behaviour, health status, self-efficacy, perceived health, determinants of food choice, nutrition knowledge, co-worker support, job satisfaction, economic cost and food-purchasing patterns. Data sources included PubMed, Medline, Embase, Psych Info., Web of Knowledge and Cochrane Library (November 2011). This review was guided by the PRISMA (Preferred Reporting Items for Systematic Reviews and Meta-Analyses) statement. Studies were randomised controlled trials and controlled studies. Interventions were implemented for at least three months. Cochrane Collaboration's risk of bias tool measured potential biases. Heterogeneity precluded meta-analysis. Results were presented in a narrative summary. Six studies conducted in Brazil, the USA, Netherlands and Belgium met the inclusion criteria. Four studies reported small increases in fruit and vegetable consumption (≤half serving/day). These studies involved workplace dietary modifications and three incorporated nutrition education. Other outcomes reported included health status, co-worker support, job satisfaction, perceived health, self-efficacy and food-purchasing patterns. All studies had methodological limitations that weakened confidence in the results. Limited evidence suggests that workplace dietary modification interventions alone and in combination with nutrition education increase fruit and vegetable intakes. These interventions should be developed with recommended guidelines, workplace characteristics, long-term follow-up and objective outcomes for diet, health and cost. © 2013.

  16. Genetic Control of the Secondary Modification of Deoxyribonucleic Acid in Escherichia coli1

    Science.gov (United States)

    Mamelak, Linda; Boyer, Herbert W.

    1970-01-01

    The wild-type restriction and modification alleles of Escherichia coli K-12 and B were found to have no measurable effect on the patterns of methylated bases in the deoxyribonucleic acid (DNA) of these strains. The genetic region controlling the methylation of cytosine in E. coli K-12 was mapped close to his, and the presence or absence of this gene in E. coli B or E. coli K had no effect on the restriction and modification properties of these strains. Thus, only a few of the methylated bases in the DNA of these strains are involved in host modification, and the biological role of the remainder remains obscure. PMID:4919756

  17. Laser surface modification of PEEK

    Energy Technology Data Exchange (ETDEWEB)

    Riveiro, A., E-mail: ariveiro@uvigo.es [Applied Physics Department, University of Vigo ETSII, Lagoas-Marcosende, 9, Vigo 36310 (Spain); Centro Universitario de la Defensa, Escuela Naval Militar, Plaza de Espana 2, 36920 Marin (Spain); Soto, R.; Comesana, R.; Boutinguiza, M.; Val, J. del; Quintero, F.; Lusquinos, F.; Pou, J. [Applied Physics Department, University of Vigo ETSII, Lagoas-Marcosende, 9, Vigo 36310 (Spain)

    2012-09-15

    Highlights: Black-Right-Pointing-Pointer Role of laser irradiation wavelength on the surface modification of PEEK (polyether-ether-ketone) was investigated. Black-Right-Pointing-Pointer Adequate processing conditions to improve wettability, roughness, and cell adhesion characteristics are determined. Black-Right-Pointing-Pointer A design of experiments (DOE) methodology was performed. Black-Right-Pointing-Pointer UV (355 nm) radiation is the most promising laser radiation for improving the adhesive surface properties of PEEK. - Abstract: Polyether-ether-ketone (PEEK) is a synthetic thermoplastic polymer with excellent mechanical and chemical properties, which make it attractive for the field of reconstructive surgery. Nevertheless, this material has a poor interfacial biocompatibility due to its large chemical stability which induces poor adhesive bonding properties. The possibilities of enhancing the PEEK adhesive properties by laser treatments have been explored in the past. This paper presents a systematic approach to discern the role of laser irradiation wavelength on the surface modification of PEEK under three laser wavelengths ({lambda} = 1064, 532, and 355 nm) with the aim to determine the most adequate processing conditions to increase the roughness and wettability, the main parameters affecting cell adhesion characteristics of implants. Overall results show that the ultraviolet ({lambda} = 355 nm) laser radiation is the most suitable one to enhance surface wettability of PEEK.

  18. Structural modifications of spinels under radiation

    International Nuclear Information System (INIS)

    Quentin, A.

    2010-12-01

    This work is devoted to the study of spinel structure materials under radiation. For that purpose, samples of polycrystalline ZnAl 2 O 4 and monocrystalline MgAl 2 O 4 were irradiated by different heavy ions with different energies. Samples of ZnAl 2 O 4 were studied par electron transmission microscopy, and by grazing incidence X-Ray diffraction and Rietveld analysis. Samples of MgAl 2 O 4 were studied by optical spectroscopy. Most of the results concern amorphization and crystalline structure modification of ZnAl 2 O 4 especially the inversion. We were able to determine a stopping power threshold for amorphization, between 11 keV/nm and 12 keV/nm, and also the amorphization process, which is a multiple impacts process. We studied the evolution of the amorphous phase by TEM and showed a nano-patterning phenomenon. Concerning the inversion, we determined that it did happen by a single impact process, and the saturation value did not reach the random cation distribution value. Inversion and amorphization have different, but close, stopping power threshold. However, amorphization seems to be conditioned by a pre-damage of the material which consists in inversion. (author)

  19. Task Modification and Knowledge Utilization by Korean Prospective Mathematics Teachers.

    Directory of Open Access Journals (Sweden)

    Kyeong-Hwa Lee

    2016-11-01

    Full Text Available It has been asserted that mathematical tasks play a critical role in the teaching and learning of mathematics. Modification of tasks included in intended curriculum materials, such as textbooks, can be an effective activity for prospective teachers to understand the role of mathematical tasks in the teaching and learning of mathematics; designing of new tasks requires more knowledge and experience. This study aims to identify the patterns that Korean prospective mathematics teachers seem to follow when they modify the mathematical tasks in textbooks. Knowledge utilized by prospective teachers while they modify textbook tasks is identified and characterized in order to understand the possible factors that have an impact on Korean prospective mathematics teachers' modification of tasks.

  20. Surface Modification for Microreactor Fabrication

    Directory of Open Access Journals (Sweden)

    Wladyslaw Torbicz

    2006-04-01

    Full Text Available In this paper, methods of surface modification of different supports, i.e. glass andpolymeric beads for enzyme immobilisation are described. The developed method ofenzyme immobilisation is based on Schiff’s base formation between the amino groups onthe enzyme surface and the aldehyde groups on the chemically modified surface of thesupports. The surface of silicon modified by APTS and GOPS with immobilised enzymewas characterised by atomic force microscopy (AFM, time-of-flight secondary ion massspectroscopy (ToF-SIMS and infrared spectroscopy (FTIR. The supports withimmobilised enzyme (urease were also tested in combination with microreactors fabricatedin silicon and Perspex, operating in a flow-through system. For microreactors filled withurease immobilised on glass beads (Sigma and on polymeric beads (PAN, a very high andstable signal (pH change was obtained. The developed method of urease immobilisationcan be stated to be very effective.

  1. Ion beam modification of polymers

    International Nuclear Information System (INIS)

    Sofield, C.J.; Sugden, S.; Ing, J.; Bridwell, L.B.; Wang, Y.Q.

    1993-01-01

    The implantation of polymers has received considerable attention in recent years, primarily to examine doping of conducting polymers and to increase the surface conductivity (by many orders of magnitude) of highly insulating polymers. The interest in these studies was partly motivated by possible applications to microelectronic device fabrication. More recently it has been observed that ion implantation can under some conditions lead to the formation of a hard (e.g. as hard as steel, ca. 3 MPa) and conducting surface layer. This paper will review the ion beam modification of polymers resulting from ion implantation with reference to fundamental ion-solid interactions. This leads us to examine whether or not implantation of polymers is a contradiction in terms. (Author)

  2. Soy protein modification: A review

    Directory of Open Access Journals (Sweden)

    Barać Miroljub B.

    2004-01-01

    Full Text Available Soy protein products such as flour, concentrates and isolates are used in food formulation because of their functionality, nutritional value and low cost. To obtain their optimal nutritive and functional properties as well as desirable flavor different treatments are used. Soybean proteins can be modified by physical, chemical and enzymatic treatments. Different thermal treatments are most commonly used, while the most appropriate way of modifying soy proteins from the standpoint of safety is their limited proteolysis. These treatments cause physical and chemical changes that affect their functional properties. This review discusses three principal methods used for modification of soy protein products, their effects on dominant soy protein properties and some biologically active compounds.

  3. Modification of carotenoid levels by abscission agents and expression of carotenoid biosynthetic genes in 'valencia' sweet orange.

    Science.gov (United States)

    Alferez, Fernando; Pozo, Luis V; Rouseff, Russell R; Burns, Jacqueline K

    2013-03-27

    The effect of 5-chloro-3-methyl-4-nitro-1H-pyrazole (CMNP) and ethephon on peel color, flavedo carotenoid gene expression, and carotenoid accumulation was investigated in mature 'Valencia' orange ( Citrus sinensis L. Osbeck) fruit flavedo at three maturation stages. Abscission agent application altered peel color. CMNP was more effective than ethephon in promoting green-to-red (a) and blue-to-yellow (b) color at the middle and late maturation stages and total carotenoid changes at all maturation stages. Altered flow of carotenoid precursors during maturation due to abscission agents was suggested by changes in phytoene desaturase (Pds) and ζ-carotene desaturase (Zds) gene expression. However, each abscission agent affected downstream expression differentially. Ethephon application increased β-carotene hydroxilase (β-Chx) transcript accumulation 12-fold as maturation advanced from the early to middle and late stages. CMNP markedly increased β- and ε-lycopene cyclase (Lcy) transcript accumulation 45- and 15-fold, respectively, at midmaturation. Patterns of carotenoid accumulation in flavedo were supported in part by gene expression changes. CMNP caused greater accumulation of total flavedo carotenoids at all maturation stages when compared with ethephon or controls. In general, CMNP treatment increased total red carotenoids more than ethephon or the control but decreased total yellow carotenoids at each maturation stage. In control fruit flavedo, total red carotenoids increased and yellow carotenoids decreased as maturation progressed. Trends in total red carotenoids during maturation were consistent with measured a values. Changes in carotenoid accumulation and expression patterns in flavedo suggest that regulation of carotenoid accumulation is under transcriptional, translational, and post-translational control.

  4. TWO MODIFICATIONS OF SCALERS FOR RATE STUDIES

    Energy Technology Data Exchange (ETDEWEB)

    Hodgson, T. S.; Gordon, B. E.

    1964-04-15

    Two simple modifications to nuclear scalers have been developed. The first involves an automatic stepping switch which permits a single scaler to be actuated by up to four Geiger counters in sequence and to record the time to reach a preset count for each. The second modification is designed to pick a pulse off a conventional scaler when a preset count has been reached and to use the pulse to actuate a recorder. Both modifications considerably extend the utility of conventional scalers.

  5. Modification of metallic corrosion by ion implantation

    International Nuclear Information System (INIS)

    Clayton, C.R.

    1981-01-01

    This review will consider some of the properties of surface alloys, formed by ion implantation, which are effective in modifying corrosion behaviour. Examples will be given of the modification of the corrosion behaviour of pure metals, steels and other engineering alloys, resulting from implantation with metals and metalloids. Emphasis will be given to the modification of anodic processes produced by ion implantation since a review will be given elsewhere in the proceedings concerning the modification of cathodic processes. (orig.)

  6. Chemical Modifications of Starch: Microwave Effect

    OpenAIRE

    Lewicka, Kamila; Siemion, Przemysław; Kurcok, Piotr

    2015-01-01

    This paper presents basic methods of starch chemical modification, the effect of microwave radiation on the modification process, and the physicochemical properties of starch. It has been shown that the modifications contribute to improvement of the material performance and likewise to significant improvement of its mechanical properties. As a result, more and more extensive use of starch is possible in various industries. In addition, methods of oxidized starch and starch esters preparation ...

  7. Chemical Modifications of Starch: Microwave Effect

    Directory of Open Access Journals (Sweden)

    Kamila Lewicka

    2015-01-01

    Full Text Available This paper presents basic methods of starch chemical modification, the effect of microwave radiation on the modification process, and the physicochemical properties of starch. It has been shown that the modifications contribute to improvement of the material performance and likewise to significant improvement of its mechanical properties. As a result, more and more extensive use of starch is possible in various industries. In addition, methods of oxidized starch and starch esters preparation are discussed. Properties of microwave radiation and its impact on starch (with particular regard to modifications described in literature are characterized.

  8. Root patterning

    NARCIS (Netherlands)

    Scheres, Ben; Laskowski, Marta

    2016-01-01

    The mechanisms that pattern lateral root primordial are essential for the elaboration of root system architecture, a trait of key importance for future crop breeding. But which are most important: periodic or local cues? In this issue of Journal of Experimental Botany (pages 1411-1420), Kircher

  9. Granular patterns

    CERN Document Server

    Aranson, Igor S

    2009-01-01

    This title presents a review of experiments and novel theoretical concepts needed to understand the mechanisms of pattern formation in granular materials. An effort is made to connect concepts and ideas developed in granular physics with new emergent fields, especially in biology, such as cytoskeleton dynamics.

  10. Identification of histone modifications in biomedical text for supporting epigenomic research.

    Science.gov (United States)

    Kolárik, Corinna; Klinger, Roman; Hofmann-Apitius, Martin

    2009-01-30

    Posttranslational modifications of histones influence the structure of chromatine and in such a way take part in the regulation of gene expression. Certain histone modification patterns, distributed over the genome, are connected to cell as well as tissue differentiation and to the adaption of organisms to their environment. Abnormal changes instead influence the development of disease states like cancer. The regulation mechanisms for modifying histones and its functionalities are the subject of epigenomics investigation and are still not completely understood. Text provides a rich resource of knowledge on epigenomics and modifications of histones in particular. It contains information about experimental studies, the conditions used, and results. To our knowledge, no approach has been published so far for identifying histone modifications in text. We have developed an approach for identifying histone modifications in biomedical literature with Conditional Random Fields (CRF) and for resolving the recognized histone modification term variants by term standardization. For the term identification F1 measures of 0.84 by 10-fold cross-validation on the training corpus and 0.81 on an independent test corpus have been obtained. The standardization enabled the correct transformation of 96% of the terms from training and 98% from test the corpus. Due to the lack of terminologies exhaustively covering specific histone modification types, we developed a histone modification term hierarchy for use in a semantic text retrieval system. The developed approach highly improves the retrieval of articles describing histone modifications. Since text contains context information about performed studies and experiments, the identification of histone modifications is the basis for supporting literature-based knowledge discovery and hypothesis generation to accelerate epigenomic research.

  11. Infections associated with body modification

    Directory of Open Access Journals (Sweden)

    Samson Sai-Yin Wong

    2012-12-01

    Full Text Available Although exact statistics are lacking, body modifications for cosmetic purposes are performed in many countries. The commonest forms include tattooing, body piercing, and breast and facial augmentation using implants or injectable fillers. Liposuction and, to a lesser extent, mesotherapy are also practiced in many countries. Infective complications of these procedures include local infections, transmission of bloodborne pathogens (viral hepatitis and human immunodeficiency virus, and distant infections such as infective endocarditis. Presence of foreign bodies, long healing time of piercing wounds, and poor compliance with infection control practices of some practitioners all predispose the recipients to infections. Apart from the endogenous microbial flora of the skin and mucosae, atypical mycobacteria, especially the rapid growers, have emerged as some of the most important pathogens in such settings. Outbreaks of infection are commonly reported. We hereby review the current knowledge of the topic with specific focus on infections associated with tattooing, body piercing, breast augmentation, mesotherapy, liposuction, and tissue filler injections. Greater awareness among consumers and health-care professionals, as well as more stringent regulations by the health authorities, is essential to minimize the health risks arising from these procedures.

  12. Quality planning for major plant design modifications

    International Nuclear Information System (INIS)

    Dulee, R.J.

    1988-01-01

    This paper reviews the approach and activities undertaken by Public Service Electric and Gas Company's (PSE and G's) nuclear quality assurance (QA) department to support major plant design modifications conducted during refueling outages at Salem Generating Station. It includes the planning and implementation of quality plans developed to provide both QA and quality control (QC) coverage of modification performed by contracted service organizations

  13. Modification of the perturbative QCD towards confinement

    International Nuclear Information System (INIS)

    Arodz, H.

    1981-01-01

    Modification of the low momentum behaviour of the perturbative SU(2) gauge theory is proposed. The modification is closely related (although not equivalent) to a nonstandard choice of boundary condition for the Euclidean 2-point gluonic Green function. In the resulting theory already single graphs lead to the confining potential between heavy, static quarks, V(r) = ar 2 for r → infinity. (author)

  14. 75 FR 41529 - Petitions for Modification

    Science.gov (United States)

    2010-07-16

    ... application of existing mandatory safety standards published in Title 30 of the Code of Federal Regulations... DEPARTMENT OF LABOR Mine Safety and Health Administration Petitions for Modification AGENCY: Mine Safety and Health Administration (MSHA), Labor. ACTION: Notice of petitions for modification of existing...

  15. 75 FR 29784 - Petitions for Modification

    Science.gov (United States)

    2010-05-27

    ... application of existing mandatory safety standards published in Title 30 of the Code of Federal Regulations... DEPARTMENT OF LABOR Mine Safety and Health Administration Petitions for Modification AGENCY: Mine Safety and Health Administration (MSHA), Labor. ACTION: Notice of petitions for modification of existing...

  16. 75 FR 34486 - Petitions for Modification

    Science.gov (United States)

    2010-06-17

    ... application of existing mandatory safety standards published in Title 30 of the Code of Federal Regulations... DEPARTMENT OF LABOR Mine Safety and Health Administration Petitions for Modification AGENCY: Mine Safety and Health Administration (MSHA), Labor. ACTION: Notice of petitions for modification of existing...

  17. Proof patterns

    CERN Document Server

    Joshi, Mark

    2015-01-01

    This innovative textbook introduces a new pattern-based approach to learning proof methods in the mathematical sciences. Readers will discover techniques that will enable them to learn new proofs across different areas of pure mathematics with ease. The patterns in proofs from diverse fields such as algebra, analysis, topology and number theory are explored. Specific topics examined include game theory, combinatorics, and Euclidean geometry, enabling a broad familiarity. The author, an experienced lecturer and researcher renowned for his innovative view and intuitive style, illuminates a wide range of techniques and examples from duplicating the cube to triangulating polygons to the infinitude of primes to the fundamental theorem of algebra. Intended as a companion for undergraduate students, this text is an essential addition to every aspiring mathematician’s toolkit.

  18. Pattern classification

    CERN Document Server

    Duda, Richard O; Stork, David G

    2001-01-01

    The first edition, published in 1973, has become a classic reference in the field. Now with the second edition, readers will find information on key new topics such as neural networks and statistical pattern recognition, the theory of machine learning, and the theory of invariances. Also included are worked examples, comparisons between different methods, extensive graphics, expanded exercises and computer project topics. An Instructor's Manual presenting detailed solutions to all the problems in the book is available from the Wiley editorial department.

  19. Modification Semantics in Now-Relative Databases

    DEFF Research Database (Denmark)

    Torp, Kristian; Jensen, Christian Søndergaard; Snodgrass, R. T.

    2004-01-01

    Most real-world databases record time-varying information. In such databases, the notion of ??the current time,?? or NOW, occurs naturally and prominently. For example, when capturing the past states of a relation using begin and end time columns, tuples that are part of the current state have some...... past time as their begin time and NOW as their end time. While the semantics of such variable databases has been described in detail and is well understood, the modification of variable databases remains unexplored. This paper defines the semantics of modifications involving the variable NOW. More...... specifically,  the problems with modifications in the presence of NOW are explored, illustrating that the main problems are with modifications of tuples that reach into the future. The paper defines the semantics of modifications?including insertions, deletions, and updates?of databases without NOW, with NOW...

  20. Methodological guidelines for developing accident modification functions

    DEFF Research Database (Denmark)

    Elvik, Rune

    2015-01-01

    This paper proposes methodological guidelines for developing accident modification functions. An accident modification function is a mathematical function describing systematic variation in the effects of road safety measures. The paper describes ten guidelines. An example is given of how to use...... limitations in developing accident modification functions are the small number of good evaluation studies and the often huge variation in estimates of effect. It is therefore still not possible to develop accident modification functions for very many road safety measures. © 2015 Elsevier Ltd. All rights...... the guidelines. The importance of exploratory analysis and an iterative approach in developing accident modification functions is stressed. The example shows that strict compliance with all the guidelines may be difficult, but represents a level of stringency that should be strived for. Currently the main...

  1. Application of radiation technology in starch modification

    International Nuclear Information System (INIS)

    Chen Huiyuan; Peng Zhigang; Ding Zhongmin; Lu Jiajiu

    2007-01-01

    In order to commercialize the radiation modification of starch, corn starch was irradiated with different dose of 60 Co gamma radiations. Some basic physical and chemical properties of the resulted modified starch paste were measured with emphasis on the viscosity stability and tensile strength. The results indicate that irradiation of corn starch with a dose of 4-10 kGy can decrease its viscosity to 5-14 mPa·s, and the tensile strength can meet the standard set up for textile paste. In comparison with chemical modification for starch, radiation modification is simpler in technology, more convenient in operation, more stable in modification quality, and easier to control. The mechanism of radiation modification of starch was also discussed. (authors)

  2. Modifications to the INSPECT model

    Energy Technology Data Exchange (ETDEWEB)

    Dickinson, S [AEA Technology, Winfrith (United Kingdom); Sims, H E [AEA Technology, Harwell (United Kingdom)

    1996-12-01

    The prediction of iodine behaviour in the containment of a PWR following a loss of coolant accident requires a reliable model of the chemistry of iodine in aqueous solution. The INSPECT model, which was developed several years ago, contains a large number of the relevant chemical reactions of iodine and water radiation chemistry. Since the reactions set was first assembled, however, new data on rate constants and mechanisms have become available. In addition, the application of the model to various small-scale experiments has revealed problems in the modelling of some reactions, leading to an under-prediction of the iodine volatility at high pH, although the experiments have demonstrated that the high pH volatility remains satisfactory low. This paper describes the modifications which have been made to the INSPECT model to take account of new data and to improve the modelling where appropriate. The main changes which have been made to the reaction set are as follows: - The rate constants and activation energies for the reactions describing the radiolysis of water have been updated in with recent assessments, and the temperature dependence of the G value for the primary species have been accordance taken into account, - The mechanism and rates of I{sub 2} hydrolysis have been modified in accordance with the latest assessments of this reaction, - The mechanism for the reaction of I{sub 2} with H{sub 2}O{sub 2} has been changed to a form which produces a more realistic pH dependence under neutral and alkaline conditions, - An addition mechanism for the disproportionation of the O{sub 2} ion has been included, reflecting experimental observations that this reaction has a significant first-order component under all but the purest conditions, - Atomic I is treated as a volatile species, with a partition coefficient of 1.9 at 298 K. (Abstract Truncated)

  3. Surface modifications of dental implants.

    Science.gov (United States)

    Stanford, C M

    2008-06-01

    Dental implant surface technologies have been evolving rapidly to enhance a more rapid bone formation on their surface and hold a potential to increase the predictability of expedited implant therapy. While implant outcomes have become highly predictable, there are sites and conditions that result in elevated implant loss. This paper reviews the impact of macro-retentive features which includes approaches to surface oxide modification, thread design, press-fit and sintered-bead technologies to increase predictability of outcomes. Implant designs that lead to controlled lateral compression of the bone can improve primary stability as long as the stress does not exceed the localized yield strength of the cortical bone. Some implant designs have reduced crestal bone loss by use of multiple cutting threads that are closely spaced, smoothed on the tip but designed to create a hoop-stress stability of the implant as it is completely seated in the osteotomy. Following the placement of the implant, there is a predictable sequence of bone turnover and replacement at the interface that allows the newly formed bone to adapt to microscopic roughness on the implant surface, and on some surfaces, a nanotopography (<10(-9) m scale) that has been shown to preferably influence the formation of bone. Newly emerging studies show that bone cells are exquisitely sensitive to these topographical features and will upregulate the expression of bone related genes for new bone formation when grown on these surfaces. We live in an exciting time of rapid changes in the modalities we can offer patients for tooth replacement therapy. Given this, it is our responsibility to be critical when claims are made, incorporate into our practice what is proven and worthwhile, and to continue to support and provide the best patient care possible.

  4. The effect of habitat modification on plant-pollinator network

    Science.gov (United States)

    Aminatun, Tien; Putra, Nugroho Susetya

    2017-08-01

    The research aimed to determine; (1) the mutualism interaction pattern of plant-pollinator on several habitat modifications; and (2) the habitat modification which showed the most stable pattern of interaction. The study was conducted in one planting season with 20 plots which each plot had 2x2 m2 width and 2 m spacing among plots, and each plot was planted with the same variety of tomato plants, i.e. "intan". Nitrogen manipulation treatment was conducted with four kinds of fertilizers, i.e. NPK (code PU), compost (code PKM), vermicompost (code PC), and manure (code PK). Each treatment had 5 plot replications. We observed the growth of tomato plants, weed and arthropod populationstwo weekly while pollinator visitation twice a week during tomato plant flowering with counting population and visitation frequence of each pollinator on each sample of tomato plants. The nectar of tomato plant flower of each treatment was tested in laboratory to see its reducing sugar and sucrose. Oganic matter and nitrogen of the soil samples of each treatment were tested in laboratory in the beginning and the end of this research. We analized the plant-pollinator network with bipartite program in R-statistics, and the abiotic and other biotic factors with descriptive analysis. The results of the research were; (1) the mutualism interaction pattern of plant-pollinator network of four treatments were varied, and (2) The pattern of plant-pollinator network of NPK fertilizer treatment showed the more stable interaction based on analysis of interaction evenness, Shannon diversity, frequency and longevity of pollinator visitation.

  5. Antibiotic Utilization and Prescribing Patterns in a Nigerian ...

    African Journals Online (AJOL)

    The study of prescribing pattern seeks to monitor, evaluate and suggest a modification in prescriber's prescribing habits so as to make medical care rational and cost effective. Information about antibiotic use pattern is necessary for a constructive approach to problems that arise from multiple antibiotics available. To identify ...

  6. Consequences for central nervous system functional state of exposure to ionizing radiation modification with antioxidants

    International Nuclear Information System (INIS)

    Tukalenko, Je.V.; Varets'kij, V.V.; Rakochyi, O.G.; Dmyitryijeva, Yi.R.

    2004-01-01

    Aim: to estimate the pattern of ionizing radiation effects modification by antioxidants using central nervous system functional state indices. The studies were carried out using 84 rats. Beta-carotene and alpha-tocopherol were found to significantly improve conditioned activity indices level of the animals exposed to ionizing radiation and emotional-pain stress

  7. Interlanguage request modification: a case in vocational college

    Science.gov (United States)

    Widanta, I. M. R. J.; Sitawati, A. A. R.; Suciani, N. K.; Handayani, L. N. C.

    2018-01-01

    There has been much attention given by scholars to the investigation of inter-language pragmatics (ILP), and some of them have been concentrating on how ILP speakers modify their speech acts (SA) of request. This study was aimed at investigating request modification produced by Indonesian English speakers. A group of 23 college students majoring in tourism was involved as research participants. The participants were given two tests using two role play cards with two hotel-context request situations, i.e. low imposing request (R-Rq) and high imposing request (R+ Rq). Pretest was given prior to and post-test was given upon treatment. The situation was chosen based on[1] exemplar generation1 model. The data of request utterances was analyzed and compared with request taxonomies proposed by some scholars. Data analysis showed that the research participants were more competent pragmatically upon the treatment, indicated with the fact where they were able to produce 13 request modification patterns being compared to 11 patterns prior to the treatment.

  8. Surface Topographical Modification of Coronary Stent: A Review

    Science.gov (United States)

    Tan, C. H.; Muhamad, N.; Abdullah, M. M. A. B.

    2017-06-01

    Driven by the urge of mediating the inflammatory response from coronary stent implant to improve patency rates of the current coronary stent, concern has been focusing on reducing the risk of in-stent restenosis and thrombosis for long-term safety. Surface modification approach has been found to carry great potential due to the surface is the vital parts that act as a buffer layer between the biomaterial and the organic material like blood and vessel tissues. Nevertheless, manipulating cell response in situ using physical patterning is very complex as the exact mechanism were yet elucidated. Thus, the aim of this review is to summarise the recent efforts on modifying the surface topography of coronary stent at the micro- and nanometer scale with the purpose of inducing rapid in situ endothelialization to regenerate a healthy endothelium layer on biomaterial surface. In particular, a discussion on the surface patterns that have been investigated on cell selective behaviour together with the methods used to generate them are presented. Furthermore, the probable future work involving the surface modification of coronary stent were indicated.

  9. Modification of Sr on 4004 Aluminum Alloy

    Science.gov (United States)

    Guo, Erjun; Cao, Guojian; Feng, Yicheng; Wang, Liping; Wang, Guojun; Lv, Xinyu

    2013-05-01

    As a brazing foil, 4004 Al alloy has good welding performance. However, the high Si content decreases the plasticity of the alloy. To improve the plasticity of 4004 Al alloy and subsequently improve the productivity of 4004 Al foil or 434 composite foil, 4004 Al alloy was modified by Al-10%Sr master alloy. Modification effects of an additional amount of Sr, modification temperature, and holding time on 4004 aluminum alloy were studied by orthogonal design. The results showed that the greatest impact parameter of 4004 aluminum alloy modification was the additional amount of Sr, followed by holding time and modification temperature. The optimum modification parameters obtained by orthogonal design were as follows: Sr addition of 0.04%, holding time of 60 min, and modification temperature of 760°C. The effect of Sr addition on modification was analyzed in detail based on orthogonal results. With increasing of Sr addition, elongation of 4004 alloy increased at first, and decreased after reaching the maximum value.

  10. Final Report for Award DE-FG02-09ER16008

    Energy Technology Data Exchange (ETDEWEB)

    Somerville, Chris

    2014-04-26

    The research objectives of this project was to characterize the factors that regulate the pattern, amount and quality of cellulose deposition in higher plants. We pursued this goal in four ways: (1) by searching for proteins that interact with the cellulose synthase complex using novel bioinformatics tools to identify genes which exhibit gene expression patterns that are highly correlated with cellulose synthase components, (2) by using cellulose synthase domains as bait in 2-hybrid screens, and then characterizing any proteins that are identified by both gene expression network analysis and 2-hybrid screens (3) by testing the function of all known post-translational modifications, (4) by identifying and characterizing kinases that carry out most of the known post-translational modification of cellulose synthase components.

  11. Chemical modification of flax reinforced polypropylene composites

    CSIR Research Space (South Africa)

    Jacob John, Maya

    2009-04-01

    Full Text Available This paper presents an experimental study on the static and dynamic mechanical properties of nonwoven based flax fibre reinforced polypropylene composites. The effect of zein modification on flax fibres is also reported. Flax nonwovens were treated...

  12. Modifications of the Weibull distribution: A review

    International Nuclear Information System (INIS)

    Almalki, Saad J.; Nadarajah, Saralees

    2014-01-01

    It is well known that the Weibull distribution is the most popular and the most widely used distribution in reliability and in analysis of lifetime data. Unfortunately, its hazard function cannot exhibit non-monotonic shapes like the bathtub shape or the unimodal shape. Since 1958, the Weibull distribution has been modified by many researchers to allow for non-monotonic hazard functions. This paper gives an extensive review of some discrete and continuous versions of the modifications of the Weibull distribution. - Highlights: • A comprehensive review of known discrete modifications and generalizations of the Weibull distribution. • A comprehensive review of known continuous modifications and generalizations of the Weibull distribution. • Over 110 references on modifications/generalizations of the Weibull distribution. • More than 55% of the cited references appeared in the last 5 years

  13. ENDF/B-5 modifications 1986

    International Nuclear Information System (INIS)

    McLaughlin, P.K.

    1986-05-01

    This document summarizes the modifications made to the ENDF/B-5 evaluated neutron data files in 1986. The new versions of the files are available from the IAEA Nuclear Data Section upon request, costfree, on magnetic tape. (author)

  14. The computational nature of memory modification.

    Science.gov (United States)

    Gershman, Samuel J; Monfils, Marie-H; Norman, Kenneth A; Niv, Yael

    2017-03-15

    Retrieving a memory can modify its influence on subsequent behavior. We develop a computational theory of memory modification, according to which modification of a memory trace occurs through classical associative learning, but which memory trace is eligible for modification depends on a structure learning mechanism that discovers the units of association by segmenting the stream of experience into statistically distinct clusters (latent causes). New memories are formed when the structure learning mechanism infers that a new latent cause underlies current sensory observations. By the same token, old memories are modified when old and new sensory observations are inferred to have been generated by the same latent cause. We derive this framework from probabilistic principles, and present a computational implementation. Simulations demonstrate that our model can reproduce the major experimental findings from studies of memory modification in the Pavlovian conditioning literature.

  15. Chemical modification and characterization of quaternized polysulfones.

    CSIR Research Space (South Africa)

    Nonjola, P

    2008-12-01

    Full Text Available Synthesis and characterization of anion-exchange membranes (AEMs) using polysulfones is described. The modification process of polysulfones involves two steps: Firstly, by introducing chloromethyl groups followed by quaternization reaction...

  16. Formal Design Review Foot Clamp Modification

    International Nuclear Information System (INIS)

    OTEN, T.C.

    2000-01-01

    This report documents the Design Review performed for the foot clamp modification. The report documents the acceptability of the design, identifies the documents that were reviewed, the scope of the review and the members of the review team

  17. The computational nature of memory modification

    Science.gov (United States)

    Gershman, Samuel J; Monfils, Marie-H; Norman, Kenneth A; Niv, Yael

    2017-01-01

    Retrieving a memory can modify its influence on subsequent behavior. We develop a computational theory of memory modification, according to which modification of a memory trace occurs through classical associative learning, but which memory trace is eligible for modification depends on a structure learning mechanism that discovers the units of association by segmenting the stream of experience into statistically distinct clusters (latent causes). New memories are formed when the structure learning mechanism infers that a new latent cause underlies current sensory observations. By the same token, old memories are modified when old and new sensory observations are inferred to have been generated by the same latent cause. We derive this framework from probabilistic principles, and present a computational implementation. Simulations demonstrate that our model can reproduce the major experimental findings from studies of memory modification in the Pavlovian conditioning literature. DOI: http://dx.doi.org/10.7554/eLife.23763.001 PMID:28294944

  18. 75 FR 3253 - Petitions for Modification

    Science.gov (United States)

    2010-01-20

    ... face equipment; maintenance) and 30 CFR 18.35 (Portable trailing cables and cords). Modification...) Limit the speed of the machine to 10 miles per hour (MPH) by permanently blocking out any gear that...

  19. Medication for Behavior Modification in Birds

    NARCIS (Netherlands)

    van Zeeland, Yvonne

    2018-01-01

    The use of behavior modifying drugs may be considered in birds with behavior problems, especially those refractory to behavior modification therapy and environmental management. To accomplish behavior change, a variety of drugs can be used, including psychoactive drugs, hormones, antihistamines,

  20. Covalent Surface Modifications of Carbon Nanotubes.

    Energy Technology Data Exchange (ETDEWEB)

    Pavia Sanders, Adriana [Sandia National Lab. (SNL-CA), Livermore, CA (United States); O' Bryan, Greg [Sandia National Lab. (SNL-CA), Livermore, CA (United States)

    2017-07-01

    A report meant to document the chemistries investigated by the author for covalent surface modification of CNTs. Oxidation, cycloaddition, and radical reactions were explored to determine their success at covalently altering the CNT surface. Characterization through infrared spectroscopy, Raman spectroscopy, and thermo gravimetric analysis was performed in order to determine the success of the chemistries employed. This report is not exhaustive and was performed for CNT surface modification exploration as it pertains to the "Next Gen" project.

  1. Measurement and modification of the EEG and related behavior

    Science.gov (United States)

    Sterman, M. B.

    1991-01-01

    Electrophysiological changes in the sensorimotor pathways were found to accompany the effect of rhythmic EEG patterns in the sensorimotor cortex. Additionally, several striking behavioral changes were seen, including in particular an enhancement of sleep and an elevation of seizure threshold to epileptogenic agents. This raised the possibility that human seizure disorders might be influenced therapeutically by similar training. Our objective in human EEG feedback training became not only the facilitation of normal rhythmic patterns, but also the suppression of abnormal activity, thus requiring complex contingencies directed to the normalization of the sensorimotor EEG. To achieve this, a multicomponent frequency analysis was developed to extract and separate normal and abnormal elements of the EEG signal. Each of these elements was transduced to a specific component of a visual display system, and these were combined through logic circuits to present the subject with a symbolic display. Variable criteria provided for the gradual shaping of EEG elements towards the desired normal pattern. Some 50-70% of patients with poorly controlled seizure disorders experienced therapeutic benefits from this approach in our laboratory, and subsequently in many others. A more recent application of this approach to the modification of human brain function in our lab has been directed to the dichotomous problems of task overload and underload in the contemporary aviation environment. At least 70% of all aviation accidents have been attributed to the impact of these kinds of problems on crew performance. The use of EEG in this context has required many technical innovations and the application of the latest advances in EEG signal analysis. Our first goal has been the identification of relevant EEG characteristics. Additionally, we have developed a portable recording and analysis system for application in this context. Findings from laboratory and in-flight studies suggest that we

  2. Capture of negative muons in magnesium oxides and crystalline modifications of phosphorus

    International Nuclear Information System (INIS)

    Zinov, V.G.; Kachalkin, A.K.; Nikityuk, L.N.; Pokrovskij, V.N.; Rybakov, V.N.; Yutlandov, I.A.

    1977-01-01

    The paper is aimed at comparing the structure of mesic K X-ray patterns of phosphorus in its crystalline modifications, comparing the structure of mesic X-ray patterns of magnesium and oxygen in compounds of MgO, MgO 2 , H 2 O and metallic magnesium, as well as comparison of propabilities of μ - atomic capture in magnesium oxides. By analyzing the mesic K X-ray patterns of red and white phosphorus it is concluded that the phosphorus crystalline modification produces the effect on the line structure, the higher series number being somewhat larger for the allotrope of phosphorus with polymeric structure. A comparison is made of the mesic X-ray series of the magnesium in oxide and metal, of the oxygen in oxide and water with the analogous data for aluminium and silicon. The data confirm the supposition that chemical bond (valence electrons) plays a substantial role in meson capture

  3. Modifications to nuclear power plants. Safety guide

    International Nuclear Information System (INIS)

    2004-01-01

    This Safety Guide was prepared under the IAEA's programme for safety standards for nuclear power plants. It supplements Section 7 of the Safety Requirements publication on Safety of Nuclear Power Plants: Operation, which establishes the safety requirements for the modification of nuclear power plants. Reasons for carrying out modifications to nuclear power plants may include: (1) maintaining or strengthening existing safety provisions and thus maintaining consistency with or improving on the current design. (2) recovering from plant faults. (3) improving the thermal performance or increasing the power rating of the plant. (4) increasing the maintainability of the plant, reducing the radiation exposure of personnel or reducing the costs of plant maintenance. And (5) extending the design life of the plant. Most modifications, made on the basis of operating experience, are intended to improve on the design or to improve operational performance and flexibility. Some are rendered necessary by new regulatory requirements, ageing of the plant or obsolescence of equipment. However, the benefits of regularly updating the plant design can be jeopardized if modifications are not kept under rigorous control throughout the lifetime of the plant. The need to reduce costs and improve efficiency, in combination with changes to the structure of the electricity generation sector of the economy in many countries, has led many companies to make changes in the structure of the operating organization for nuclear power plants. Whatever the reason for such organizational changes, consideration should be given to the effects of those changes with the aim of ensuring that they would have no impacts that would compromise the safety of the plant. The objective of this Safety Guide is to provide guidance and recommendations on controlling activities relating to modifications at nuclear power plants in order to reduce risk and to ensure that the configuration of the plant is at all times under

  4. Modifications to nuclear power plants. Safety guide

    International Nuclear Information System (INIS)

    2007-01-01

    This Safety Guide was prepared under the IAEA's programme for safety standards for nuclear power plants. It supplements Section 7 of the Safety Requirements publication on Safety of Nuclear Power Plants: Operation, which establishes the safety requirements for the modification of nuclear power plants. Reasons for carrying out modifications to nuclear power plants may include: (1) maintaining or strengthening existing safety provisions and thus maintaining consistency with or improving on the current design. (2) recovering from plant faults. (3) improving the thermal performance or increasing the power rating of the plant. (4) increasing the maintainability of the plant, reducing the radiation exposure of personnel or reducing the costs of plant maintenance. And (5) extending the design life of the plant. Most modifications, made on the basis of operating experience, are intended to improve on the design or to improve operational performance and flexibility. Some are rendered necessary by new regulatory requirements, ageing of the plant or obsolescence of equipment. However, the benefits of regularly updating the plant design can be jeopardized if modifications are not kept under rigorous control throughout the lifetime of the plant. The need to reduce costs and improve efficiency, in combination with changes to the structure of the electricity generation sector of the economy in many countries, has led many companies to make changes in the structure of the operating organization for nuclear power plants. Whatever the reason for such organizational changes, consideration should be given to the effects of those changes with the aim of ensuring that they would have no impacts that would compromise the safety of the plant. The objective of this Safety Guide is to provide guidance and recommendations on controlling activities relating to modifications at nuclear power plants in order to reduce risk and to ensure that the configuration of the plant is at all times under

  5. Dynamic O-linked N-acetylglucosamine modification of proteins affects stress responses and survival of mesothelial cells exposed to peritoneal dialysis fluids.

    Science.gov (United States)

    Herzog, Rebecca; Bender, Thorsten O; Vychytil, Andreas; Bialas, Katarzyna; Aufricht, Christoph; Kratochwill, Klaus

    2014-12-01

    The ability of cells to respond and survive stressful conditions is determined, in part, by the attachment of O-linked N-acetylglucosamine (O-GlcNAc) to proteins (O-GlcNAcylation), a post-translational modification dependent on glucose and glutamine. This study investigates the role of dynamic O-GlcNAcylation of mesothelial cell proteins in cell survival during exposure to glucose-based peritoneal dialysis fluid (PDF). Immortalized human mesothelial cells and primary mesothelial cells, cultured from human omentum or clinical effluent of PD patients, were assessed for O-GlcNAcylation under normal conditions or after exposure to PDF. The dynamic status of O-GlcNAcylation and effects on cellular survival were investigated by chemical modulation with 6-diazo-5-oxo-L-norleucine (DON) to decrease or O-(2-acetamido-2-deoxy-D-glucopyranosylidene)amino N-phenyl carbamate (PUGNAc) to increase O-GlcNAc levels. Viability was decreased by reducing O-GlcNAc levels by DON, which also led to suppressed expression of the cytoprotective heat shock protein 72. In contrast, increasing O-GlcNAc levels by PUGNAc or alanyl-glutamine led to significantly improved cell survival paralleled by higher heat shock protein 72 levels during PDF treatment. Addition of alanyl-glutamine increased O-GlcNAcylation and partly counteracted its inhibition by DON, also leading to improved cell survival. Immunofluorescent analysis of clinical samples showed that the O-GlcNAc signal primarily originates from mesothelial cells. In conclusion, this study identified O-GlcNAcylation in mesothelial cells as a potentially important molecular mechanism after exposure to PDF. Modulating O-GlcNAc levels by clinically feasible interventions might evolve as a novel therapeutic target for the preservation of peritoneal membrane integrity in PD. Copyright © 2014 by the American Society of Nephrology.

  6. Overcoming the Refractory Expression of Secreted Recombinant Proteins in Mammalian Cells through Modification of the Signal Peptide and Adjacent Amino Acids.

    Science.gov (United States)

    Güler-Gane, Gülin; Kidd, Sara; Sridharan, Sudharsan; Vaughan, Tristan J; Wilkinson, Trevor C I; Tigue, Natalie J

    2016-01-01

    The expression and subsequent purification of mammalian recombinant proteins is of critical importance to many areas of biological science. To maintain the appropriate tertiary structure and post-translational modifications of such proteins, transient mammalian expression systems are often adopted. The successful utilisation of these systems is, however, not always forthcoming and some recombinant proteins prove refractory to expression in mammalian hosts. In this study we focussed on the role of different N-terminal signal peptides and residues immediately downstream, in influencing the level of secreted recombinant protein obtained from suspension HEK293 cells. Using secreted alkaline phosphatase (SEAP) as a model protein, we identified that the +1/+2 downstream residues flanking a heterologous signal peptide significantly affect secreted levels. By incorporating these findings we conducted a comparison of different signal peptide sequences and identified the most productive as secrecon, a computationally-designed sequence. Importantly, in the context of the secrecon signal peptide and SEAP, we also demonstrated a clear preference for specific amino acid residues at the +1 position (e.g. alanine), and a detrimental effect of others (cysteine, proline, tyrosine and glutamine). When proteins that naturally contain these "undesirable" residues at the +1 position were expressed with their native signal peptide, the heterologous secrecon signal peptide, or secrecon with an additional alanine at the +1 or +1 and +2 position, the level of expression differed significantly and in an unpredictable manner. For each protein, however, at least one of the panel of signal peptide/adjacent amino acid combinations enabled successful recombinant expression. In this study, we highlight the important interplay between a signal peptide and its adjacent amino acids in enabling protein expression, and we describe a strategy that could enable recombinant proteins that have so far

  7. Post-translational regulation of Foxp3 : identification of novel molecular targets for immune modulation

    NARCIS (Netherlands)

    van Loosdregt, J.

    2011-01-01

    Maintenance of immune homeostasis is a complex process allowing the immune system to be both aggressive enough to eradicate cells that express foreign antigens, and yet provide sensitivity to tolerate cells expressing self antigens. Key modulators allowing tolerance of host antigens, thereby

  8. Mass spectrometry analysis of proteome-wide proteolytic post-translational degradation of proteins

    OpenAIRE

    Shen, Yufeng; Hixson, Kim K.; Tolić, Nikola; Camp, David G.; Purvine, Samuel O.; Moore, Ronald J.; Smith, Richard D.

    2008-01-01

    Protein proteolytic degradation is an essential component to proper cell function and its life cycle. Here, we study the protein degradation in yeast Saccharomyces cerevisiae cells on a proteome-wide scale by detection of the intermediate peptides produced from the intracellular degradation of proteins using sequencing-based tandem mass spectrometry. By tracing the detected ~1,100 peptides and their ~200 protein substrate origins we obtain evidence for new insights into the proteome-wide prot...

  9. Post-translational Analysis of Arabidopsis thaliana Proteins in Response to Cyclic Guanosine Monophosphate Treatment

    KAUST Repository

    Parrott, Brian

    2011-01-01

    and phosphorylation before analysis via mass spectrometry. Preliminary results suggest a tendency towards an increased number of phosphorylated proteins as a result of cGMP treatment. The data also showed a sharp increase in methionine oxidation in response

  10. Post-translational suppression of expression of intestinal brush border enzymes by fructose

    DEFF Research Database (Denmark)

    Danielsen, E M

    1989-01-01

    The two major dietary sugars, fructose and sucrose, were found to suppress effectively the biosynthetic renewal of brush border enzymes in the gut. When studied in cultured explants of pig small intestine mucosa, 10-50 mM concentrations of fructose completely prevented the expression of mature...... cotranslational glycosylation that in turn triggers a rapid proteolytic breakdown. Our findings suggest that renewal of digestive brush border enzymes is transiently suppressed during intake of fructose- or sucrose-rich meals....

  11. Post-translational Analysis of Arabidopsis thaliana Proteins in Response to Cyclic Guanosine Monophosphate Treatment

    KAUST Repository

    Parrott, Brian

    2011-12-12

    The introduction of mass spectrometry techniques to the field of biology has made possible the exploration of the proteome as a whole system as opposed to prior techniques, such as anti-body based assays or yeast two-hybrid studies, which were strictly limited to the study of a few proteins at a time. This practice has allowed for a systems biology approach of exploring the proteome, with the possibility of viewing entire pathways over increments of time. In this study, the effect of treating Arabidopsis thaliana suspension culture cells with 3’,5’-cyclic guanosine monophosphate (cGMP), which is a native second messenger, was examined. Samples were collected at four time points and proteins were extracted and enriched for both oxidation and phosphorylation before analysis via mass spectrometry. Preliminary results suggest a tendency towards an increased number of phosphorylated proteins as a result of cGMP treatment. The data also showed a sharp increase in methionine oxidation in response to the treatment, occurring within the first ten minutes. This finding suggests that cGMP may utilize methionine oxidation as a mechanism of signal transduction. As such, this study corroborates a growing body of evidence supporting the inclusion of methionine oxidation in intracellular signaling pathways.

  12. Co-and post-translational events in the biogenesis of pig small intestinal aminopeptidase N

    DEFF Research Database (Denmark)

    Danielsen, Erik Michael; Norén, O; Sjöström, H

    1982-01-01

    The biogenesis of pig small intestinal aminopeptidase N (EC 3. 4. 11. 2) was studied by cell-free translation of intestinal mRNA and by labelling of organ cultured intestinal explants. In cell-free translation, the primary mRNA translation product of aminopeptidase N was a polypeptide of Mr 115......,000. When translation was performed in the presence of dog pancreatic microsomes, a Mr 140,000 polypeptide was also observed. A polypeptide of Mr 115,000 was seen for the enzyme, purified from tunicamycin exposed explants. This result suggests that aminopeptidase N is co-translationally inserted...

  13. TNFα Post-Translationally Targets ZnT2 to Accumulate Zinc in Lysosomes.

    Science.gov (United States)

    Hennigar, Stephen R; Kelleher, Shannon L

    2015-10-01

    Mammary epithelial cells undergo widespread lysosomal-mediated cell death (LCD) during early mammary gland involution. Recently, we demonstrated that tumor necrosis factor-α (TNFα), a cytokine released during early involution, redistributes the zinc (Zn) transporter ZnT2 to accumulate Zn in lysosomes and activate LCD and involution. The objective of this study is to determine how TNFα retargets ZnT2 to lysosomes. We tested the hypothesis that TNFα signaling dephosphorylates ZnT2 to uncover a highly conserved dileucine motif (L294L) in the C-terminus of ZnT2, allowing adaptor protein complex-3 (AP-3) to bind and traffic ZnT2 to lysosomes. Confocal micrographs showed that TNFα redistributed wild-type (WT) ZnT2 from late endosomes (Pearson's coefficient = 0.202 ± 0.05 and 0.097 ± 0.03; Plysosomes (0.292 ± 0.03 and 0.649 ± 0.03; Plysosomal Zn (Plysosomes, increase lysosomal Zn, or activate LCD. Moreover, TNFα increased (Plysosomes and activate LCD. Our findings suggest that women with variation in the C-terminus of ZnT2 may be at risk for inadequate involution and breast disease due the inability to traffic ZnT2 to lysosomes. © 2015 Wiley Periodicals, Inc.

  14. Post-translational regulation of P2X receptor channels: modulation by phospholipids

    Directory of Open Access Journals (Sweden)

    Louis-Philippe eBernier

    2013-11-01

    Full Text Available P2X receptor channels mediate fast excitatory signaling by ATP and play major roles in sensory transduction, neuro-immune communication and inflammatory response. P2X receptors constitute a gene family of calcium-permeable ATP-gated cation channels therefore the regulation of P2X signaling is critical for both membrane potential and intracellular calcium homeostasis. Phosphoinositides (PIPn are anionic signaling phospholipids that act as functional regulators of many types of ion channels. Direct PIPn binding was demonstrated for several ligand- or voltage-gated ion channels, however no generic motif emerged to accurately predict lipid-protein binding sites. This review presents what is currently known about the modulation of the different P2X subtypes by phospholipids and about critical determinants underlying their sensitivity to PIPn levels in the plasma membrane.All functional mammalian P2X subtypes tested, with the notable exception of P2X5, have been shown to be positively modulated by PIPn, i.e. homomeric P2X1, P2X2, P2X3, P2X4, and P2X7, as well as heteromeric P2X1/5 and P2X2/3 receptors. Based on various results reported on the aforementioned subtypes including mutagenesis of the prototypical PIPn-sensitive P2X4 and PIPn-insensitive P2X5 receptor subtypes, an increasing amount of functional, biochemical and structural evidence converges on the modulatory role of a short polybasic domain located in the proximal C-terminus of P2X subunits. This linear motif, semi-conserved in the P2X family, seems necessary and sufficient for encoding direct modulation of ATP-gated channels by PIPn. Furthermore, the physiological impact of the regulation of ionotropic purinergic responses by phospholipids on pain pathways was recently revealed in the context of native crosstalks between phospholipase C-linked metabotropic receptors and P2X receptor channels in DRG sensory neurons and microglia.

  15. Prediction, production and characterization of post-translationally modified antimicrobial peptides

    NARCIS (Netherlands)

    van Heel, Auke Johan

    2016-01-01

    Pathogenic bacteria are rapidly becoming resistant to the currently used antibiotics therefore we need novel antibiotics, preferably with new mechanisms of action. One potential source are the so called antimicrobial peptides that are produced by many different organisms. To gain access to these

  16. The effects of an overnight holding of whole blood at room temperature on haemoglobin modification and in vitro markers of red blood cell aging.

    Science.gov (United States)

    Eckstein, M; Zimmermann, R; Roth, T; Hauck-Dlimi, B; Strasser, E F; Xiang, W

    2015-05-01

    Some effects of the red blood cell (RBC) storage lesion are well documented whereas others are not. Whether a period of room temperature hold (RTH) during RBC production enhances the RBC storage lesion has remained controversial. In this study, we compared whole blood (WB)-derived RBCs produced after 24-h RTH with rapidly cooled (RC) RBCs and tested them for classical metabolic markers and signs of oxidative damage. SAGM-RBCs were prepared from mixed and split pairs (n = 12) of WB units. RBCs prepared after a 24-h period of RTH on day+1 after collection (RTH-RBCs) were compared with RC-RBCs. All RBCs were stored at 4°C for 42 days with assay of in vitro variables on days+1, +15, +22, +29 and +42. The study examined standard quality parameters, glutathione, catalase and superoxide dismutase (SOD) activities, and indicative markers of oxidative cell damage including post-translational haemoglobin modification, malondialdehyde (MDA), and phosphatidylserine expression. RTH-RBCs exhibited decreased levels of potassium (1·98 ± 0·26 vs. 5·23 ± 0·65 mmol/l) and of 2,3-diphosphoglycerate (2,3-DPG) on day+1 compared with RC-RBCs. Haemolysis rate on day+42 was higher in RTH-RBCs than in RC-RBCs (0·52 ± 0·13 vs. 0·37 ± 0·12%). The phosphatidylserine expression amounted to 0·25 ± 0·20% in RTH-RBCs and 0·07 ± 0·12% in RC-RBCs. Haemoglobin modification was not different between both RBC groups. RTH-RBCs showed slightly higher MDA concentration on days +29 and +42. RC-RBCs and RTH-RBCs show only small differences of classical in vitro parameters and no relevant differences in antioxidative metabolism and oxidative haemoglobin modification. These findings do not explain the loss observed in in vivo survival studies with RBCs. © 2015 International Society of Blood Transfusion.

  17. Pattern Recognition

    Directory of Open Access Journals (Sweden)

    Aleš Procházka

    2018-05-01

    Full Text Available Multimodal signal analysis based on sophisticated sensors, efficient communicationsystems and fast parallel processing methods has a rapidly increasing range of multidisciplinaryapplications. The present paper is devoted to pattern recognition, machine learning, and the analysisof sleep stages in the detection of sleep disorders using polysomnography (PSG data, includingelectroencephalography (EEG, breathing (Flow, and electro-oculogram (EOG signals. The proposedmethod is based on the classification of selected features by a neural network system with sigmoidaland softmax transfer functions using Bayesian methods for the evaluation of the probabilities of theseparate classes. The application is devoted to the analysis of the sleep stages of 184 individualswith different diagnoses, using EEG and further PSG signals. Data analysis points to an averageincrease of the length of the Wake stage by 2.7% per 10 years and a decrease of the length of theRapid Eye Movement (REM stages by 0.8% per 10 years. The mean classification accuracy for givensets of records and single EEG and multimodal features is 88.7% ( standard deviation, STD: 2.1 and89.6% (STD:1.9, respectively. The proposed methods enable the use of adaptive learning processesfor the detection and classification of health disorders based on prior specialist experience andman–machine interaction.

  18. Loss and modification of habitat

    Science.gov (United States)

    Lemckert, Francis; Hecnar, Stephen; Pilliod, David S.; Wilkinson, John W.; Heatwole, Harold

    2012-01-01

    water balance (Thorson and Svihla 1943; Brattstrom 1963; Tracy 1976). Hence, individuals require and seek specific microhabitats that maintain their preferred body temperature while at the same time reducing water loss or allowing individuals to re-hydrate. Amphibians also possess relatively few physical attributes that protect them from predators. Although they may avoid predators behaviourally or deter them by skin toxins, amphibians lack defensive shells or hardened cuticles, do not have protective teeth or claws, and most are insufficiently fast to escape predators. Hence, they are relatively dependent on sites that conceal or protect them from predation. Most amphibians also differ significantly from other vertebrates in possessing a complex two-phase life cycle: the pre-metamorphic larval (tadpole) stage and the post-metamorphic juvenile and adult stage (Wilbur 1980, 1984). Most amphibian species have two distinct econes (Heatwole 1989), each with different habitat requirements, the larvae being aquatic and the post-metamorphic animals more terrestrial. The habitats required by the two phases can differ greatly, but both are essential to the survival of a species. However, amphibian diversity is great and exceptions to this general pattern exist. For example, some species have direct development without going through a larval stage and are fully terrestrial, whereas the larvae of other species can reach sexual maturity without going through metamorphosis (i.e., neoteny) and are fully aquatic.

  19. Sequence tagging reveals unexpected modifications in toxicoproteomics

    Science.gov (United States)

    Dasari, Surendra; Chambers, Matthew C.; Codreanu, Simona G.; Liebler, Daniel C.; Collins, Ben C.; Pennington, Stephen R.; Gallagher, William M.; Tabb, David L.

    2010-01-01

    Toxicoproteomic samples are rich in posttranslational modifications (PTMs) of proteins. Identifying these modifications via standard database searching can incur significant performance penalties. Here we describe the latest developments in TagRecon, an algorithm that leverages inferred sequence tags to identify modified peptides in toxicoproteomic data sets. TagRecon identifies known modifications more effectively than the MyriMatch database search engine. TagRecon outperformed state of the art software in recognizing unanticipated modifications from LTQ, Orbitrap, and QTOF data sets. We developed user-friendly software for detecting persistent mass shifts from samples. We follow a three-step strategy for detecting unanticipated PTMs in samples. First, we identify the proteins present in the sample with a standard database search. Next, identified proteins are interrogated for unexpected PTMs with a sequence tag-based search. Finally, additional evidence is gathered for the detected mass shifts with a refinement search. Application of this technology on toxicoproteomic data sets revealed unintended cross-reactions between proteins and sample processing reagents. Twenty five proteins in rat liver showed signs of oxidative stress when exposed to potentially toxic drugs. These results demonstrate the value of mining toxicoproteomic data sets for modifications. PMID:21214251

  20. Modifications at operating nuclear power plants

    International Nuclear Information System (INIS)

    Duffy, T.J.; Gazda, P.A.

    1985-01-01

    Modifications at operating nuclear power plants offer the structural engineer many challenges in the areas of scheduling of work, field adjustments, and engineering staff planning. The scheduling of structural modification work for operating nuclear power plants is normally closely tied to planned or unplanned outages of the plant. Coordination between the structural engineering effort, the operating plant staff, and the contractor who will be performing the modifications is essential to ensure that all work can be completed within the allotted time. Due to the inaccessibility of some areas in operating nuclear power plants or the short time available to perform the structural engineering in the case of an unscheduled outrage, field verification of a design is not always possible prior to initiating the construction of the modification. This requires the structural engineer to work closely with the contractor to promptly resolve problems due to unanticipated interferences or material procurement problems that may arise during the course of construction. The engineering staff planning for structural modifications at an operating nuclear power plant must be flexible enough to permit rapid response to the common ''fire drills,'' but controlled enough to ensure technically correct designs and to minimize the expenditure of man-hours and the resulting engineering cost

  1. Proposed Modification of the Progressive Retirement Programme

    CERN Document Server

    2003-01-01

    After discussion at TREF on 29 October 2003, the Management proposes in this document a modification affecting one of the general principles of the Progressive Retirement Programme (PRP), decided by the Council in December 1996 and introduced in April 1997. This modification would authorise the Director-General, in exceptional cases, as indicated in section 3 below, and, with the staff member's consent, to cancel his or her participation in the PRP and to reinstate the staff member in his or her original contractual situation. No modifications to the Staff Rules and Regulations are required. The proposal set out in section 3 of the present document is submitted by the Management, after consultation of the Governing Board of the Pension Fund, for recommendation by the Finance Committee to the Council and for approval by the Council, to enter into force as of 1 January 2004.

  2. Three Sisters Dam modifications and performance

    Energy Technology Data Exchange (ETDEWEB)

    Courage, L.J.R. [Monenco AGRA Inc., Calgary, AB (Canada)

    1995-12-31

    Recent modifications and maintenance carried out at the Three Sisters Dam, in the Alberta Rockies south of the town of Canmore, were described. A detailed account was given of the dam`s geological setting, its abnormally high leakage through the foundation and its sinkhole activity. Results of studies aimed at finding the cause of leakage and sinkhole occurrences were reviewed. Modifications made to the dam since 1951 were detailed, as were modifications to handle probable maximum flood levels. Three approaches for estimating failure probabilities after identification of failure modes were described. The overall conclusion was that based on constant leakage, no settlement in the dam, penstocks, or the powerhouse since construction, the Three Sisters Dam was stable. 1 ref.

  3. Nanofibrillated Cellulose Surface Modification: A Review

    Directory of Open Access Journals (Sweden)

    Julien Bras

    2013-05-01

    Full Text Available Interest in nanofibrillated cellulose (NFC has increased notably over recent decades. This bio-based nanomaterial has been used essentially in bionanocomposites or in paper thanks to its high mechanical reinforcement ability or barrier property respectively. Its nano-scale dimensions and its capacity to form a strong entangled nanoporous network have encouraged the emergence of new high-value applications. It is worth noting that chemical surface modification of this material can be a key factor to achieve a better compatibility with matrices. In order to increase the compatibility in different matrices or to add new functions, surface chemical modification of NFC appears to be the prior choice to conserve its intrinsic nanofibre properties. In this review, the authors have proposed for the first time an overview of all chemical grafting strategies used to date on nanofibrillated cellulose with focus on surface modification such as physical adsorption, molecular grafting or polymer grafting.

  4. Characteristic measurements of JRR-2 after modification

    International Nuclear Information System (INIS)

    1977-03-01

    JRR-2 was shut down in December 1973, to improve it in heavy water leakage from the metal packing between core tank and support ring, corrosion of the lower shielding plug, and fault in the control-rod mechanism. Modification was on a standing seal to stop heavy water leakage at the support ring, replacement of the reactor top shield, replacement of the control-rod mechanism, improvement of the helium system, improvement of the irradiated air system to reduce radioactive argon gas release from the stack. Works were completed in September 1975. Functional tests and characteristic measurements then followed, to show good results of the modification. Described are; modification, critical experiments, calibration of the control rods, reactor thermal power, measurements of the neutron flux distribution, core heavy water flow distribution, heavy water decomposition rate, helium leak rate and tritium concentration in the thermal shielding light water, radioactive argon gas release from the stack, and effect of the reactor top shield. (auth.)

  5. Maillard reaction versus other nonenzymatic modifications in neurodegenerative processes.

    Science.gov (United States)

    Pamplona, Reinald; Ilieva, Ekaterina; Ayala, Victoria; Bellmunt, Maria Josep; Cacabelos, Daniel; Dalfo, Esther; Ferrer, Isidre; Portero-Otin, Manuel

    2008-04-01

    Nonenzymatic protein modifications are generated from direct oxidation of amino acid side chains and from reaction of the nucleophilic side chains of specific amino acids with reactive carbonyl species. These reactions give rise to specific markers that have been analyzed in different neurodegenerative diseases sharing protein aggregation, such as Alzheimer's disease, Pick's disease, Parkinson's disease, dementia with Lewy bodies, Creutzfeldt-Jakob disease, and amyotrophic lateral sclerosis. Collectively, available data demonstrate that oxidative stress homeostasis, mitochondrial function, and energy metabolism are key factors in determining the disease-specific pattern of protein molecular damage. In addition, these findings suggest the lack of a "gold marker of oxidative stress," and, consequently, they strengthen the need for a molecular dissection of the nonenzymatic reactions underlying neurodegenerative processes.

  6. Disassembling and modification of RA-3

    International Nuclear Information System (INIS)

    Tarizzo, R.D.

    1990-01-01

    The objective of this paper is to describe the partial disassembling and modification of RA-3, called the Modernization Project. It comprises all the technical and administrative steps directly related with this task. The improvement of RA-3 is a result of the lack of 90% enriched uranium obliging a change over to 20% enriched uranium. This brought about design modifications both in fuel elements and the reactor. The presentation of documents for the licensing authority as well as are detailed separately. The modernization project was divided in 25 tasks: 1) changing fuel element support table, 2) changing heat exchanger, 3) repairing of cooling towers, 4) repairing of primary circuit valves, 5) repairing of irradiation channels, 6) construction of a new sampler, 7) changing tangential channel, 8) cleaning and disassembling of reactor (inside), 9) changing continuous demineralizer (ion exchange column), 10) detection of failure in fuel elements, 11) modification of nuclear instrumentation, 12) modification of conventional instrumentation, 13) modification of electrical system, 14) changing telemanipulators, 15) construction of mechanism bridge, 16) changing a primary circuit valve when the heat exchanger is changed too, 17) painting ground floor, hall floor, and pump room floor with epoxy resin levelling, 18) installation of fire alarm system, 19) radioactive liquid discharge, 20) modification of secondary circuit (This task involves: a) installation of a third secondary pump, b) extension of this piping, c) installation of two 12 inch valves to the present cooling towers pools independent, d) installation of filtering system), 21) optimization hot water bed, 22) changing detector support table, 23) removal, decontamination and reinstallation of shielding, 25) changing pneumatic system

  7. Generalized nucleation and looping model for epigenetic memory of histone modifications

    Science.gov (United States)

    Erdel, Fabian; Greene, Eric C.

    2016-01-01

    Histone modifications can redistribute along the genome in a sequence-independent manner, giving rise to chromatin position effects and epigenetic memory. The underlying mechanisms shape the endogenous chromatin landscape and determine its response to ectopically targeted histone modifiers. Here, we simulate linear and looping-driven spreading of histone modifications and compare both models to recent experiments on histone methylation in fission yeast. We find that a generalized nucleation-and-looping mechanism describes key observations on engineered and endogenous methylation domains including intrinsic spatial confinement, independent regulation of domain size and memory, variegation in the absence of antagonists, and coexistence of short- and long-term memory at loci with weak and strong constitutive nucleation. These findings support a straightforward relationship between the biochemical properties of chromatin modifiers and the spatiotemporal modification pattern. The proposed mechanism gives rise to a phase diagram for cellular memory that may be generally applicable to explain epigenetic phenomena across different species. PMID:27382173

  8. Selective cell culture on UV transparent polymer by F2 laser surface modification

    International Nuclear Information System (INIS)

    Hanada, Yasutaka; Sugioka, Koji; Kawano, Hiroyuki; Tsuchimoto, Takayoshi; Miyamoto, Iwao; Miyawaki, Atsushi; Midorikawa, Katsumi

    2009-01-01

    A microchip made of UV transparent polymer (CYTOP) that can perform selective cell culture has been fabricated by F 2 laser surface modification. The refractive index of CYTOP is almost the same as that of culture medium, which is essential for three-dimensional (3D) observation of cells. The F 2 laser modification of CYTOP achieves hydrophilicity only on the laser irradiated area with little deterioration of the optical properties and surface smoothness. After the laser modification, HeLa cells were successfully cultured and strongly adhered only on the modified area of CYTOP. The cells patterned on CYTOP were applied for clear 3D observation using an optical microscope in phase contrast mode.

  9. Hydrophilic Surface Modification of PDMS Microchannel for O/W and W/O/W Emulsions

    Directory of Open Access Journals (Sweden)

    Shazia Bashir

    2015-09-01

    Full Text Available A surface modification method for bonded polydimethylsiloxane (PDMS microchannels is presented herein. Polymerization of acrylic acid was performed on the surface of a microchannel using an inline atmospheric pressure dielectric barrier microplasma technique. The surface treatment changes the wettability of the microchannel from hydrophobic to hydrophilic. This is a challenging task due to the fast hydrophobic recovery of the PDMS surface after modification. This modification allows the formation of highly monodisperse oil-in-water (O/W droplets. The generation of water-in-oil-in-water (W/O/W double emulsions was successfully achieved by connecting in series a hydrophobic microchip with a modified hydrophilic microchip. An original channel blocking technique to pattern the surface wettability of a specific section of a microchip using a viscous liquid comprising a mixture of honey and glycerol, is also presented for generating W/O/W emulsions on a single chip.

  10. Pulsed electric field-assisted modification of pectin from sugar beet pulp.

    Science.gov (United States)

    Ma, Sen; Wang, Zhong-he

    2013-02-15

    This current work is concerned with the modification of sugar beet pulp (SBP) pectin assisted by pulsed electric filed (PEF) without solvent. Pectin-arachates with degree of esterification (DE) ranging from 49 to 84 were prepared in one-step modification. The results showed that the DE of pectin derivatives increased significantly with the PEF intensity from 18 to 30 kV cm((1) and total specific energy input from 124 to 345 J mL((1). Evidence of modification of pectin was provided by FT-IR, X-ray diffraction patterns and NMR spectra. Thermogravimetric investigation of modified pectin indicated a higher thermal stability than the untreated one. Results revealed that PEF technology is a promising method for industrial manufacture of pectin derivatives. Copyright © 2012 Elsevier Ltd. All rights reserved.

  11. Selective cell culture on UV transparent polymer by F{sub 2} laser surface modification

    Energy Technology Data Exchange (ETDEWEB)

    Hanada, Yasutaka, E-mail: y-hanada@riken.jp [RIKEN-Advanced Science Institute, 2-1 Hirosawa, Wako, Saitama 351-0198 (Japan); Sugioka, Koji [RIKEN-Advanced Science Institute, 2-1 Hirosawa, Wako, Saitama 351-0198 (Japan); Kawano, Hiroyuki [RIKEN-Brain Science Institute, 2-1 Hirosawa, Wako, Saitama 351-0198 (Japan); Tsuchimoto, Takayoshi [RIKEN-Advanced Science Institute, 2-1 Hirosawa, Wako, Saitama 351-0198 (Japan); Department of Applied Electronics, Faculty of Industrial Science and Technology Tokyo University of Science, 2641 Yamazaki, Noda, Chiba 278-8510 (Japan); Miyamoto, Iwao [Department of Applied Electronics, Faculty of Industrial Science and Technology Tokyo University of Science, 2641 Yamazaki, Noda, Chiba 278-8510 (Japan); Miyawaki, Atsushi [RIKEN-Brain Science Institute, 2-1 Hirosawa, Wako, Saitama 351-0198 (Japan); Midorikawa, Katsumi [RIKEN-Advanced Science Institute, 2-1 Hirosawa, Wako, Saitama 351-0198 (Japan)

    2009-09-30

    A microchip made of UV transparent polymer (CYTOP) that can perform selective cell culture has been fabricated by F{sub 2} laser surface modification. The refractive index of CYTOP is almost the same as that of culture medium, which is essential for three-dimensional (3D) observation of cells. The F{sub 2} laser modification of CYTOP achieves hydrophilicity only on the laser irradiated area with little deterioration of the optical properties and surface smoothness. After the laser modification, HeLa cells were successfully cultured and strongly adhered only on the modified area of CYTOP. The cells patterned on CYTOP were applied for clear 3D observation using an optical microscope in phase contrast mode.

  12. X-ray studies on piroxicam modifications

    International Nuclear Information System (INIS)

    Reck, G.; Bannier, G.; Hoehne, E.

    1988-01-01

    The anti-inflammatory drug piroxicam, 4-hydroxy-2-methyl-N-(2-pyridyl)2H-1,2-benzothiazine-3-car-boxamide-1,1-dioxide, crystallizes in three different forms, two anhydrates and one monohydrate. Crystal structure analyses of the monohydrate and one of the anhydrate were carried out by Bordner et al. and Kojic-Prodic et al., respectively. The results of the analysis of the third piroxicam modification and of an independently performed structure determination of the monohydrate are reported. Molecular structures and hydrogen bonding of all modifications are discussed. (author)

  13. Histone modifications in response to DNA damage

    International Nuclear Information System (INIS)

    Altaf, Mohammed; Saksouk, Nehme; Cote, Jacques

    2007-01-01

    The packaging of the eukaryotic genome into highly condensed chromatin makes it inaccessible to the factors required for gene transcription, DNA replication, recombination and repair. Eukaryotes have developed intricate mechanisms to overcome this repressive barrier imposed by chromatin. Histone modifying enzymes and ATP-dependent chromatin remodeling complexes play key roles here as they regulate many nuclear processes by altering the chromatin structure. Significantly, these activities are integral to the process of DNA repair where histone modifications act as signals and landing platforms for various repair proteins. This review summarizes the recent developments in our understanding of histone modifications and their role in the maintenance of genome integrity

  14. Modification to the lower hybrid system

    International Nuclear Information System (INIS)

    Gormezano, C.

    1989-01-01

    The main modifications which have to be done to the Lower Hybrid system are related in the change of the plasma shape in front of the grill mouth. In effect, the theoretical coupling efficiency of the LHCD grill depends only upon the density at the grill mouth and upon the launched wave index. In order to minimize the number of modifications to the launcher it is proposed to modify the length of the vacuum waveguides connecting the multijunctions to the vacuum windows. To obtain the new poloidal contour, it is proposed to recover the L1 multijunctions and to remachine their mouth. (U.K.)

  15. EG and G electron linac modifications

    International Nuclear Information System (INIS)

    Norris, N.J.; Detch, J.L.; Kocimski, S.M.; Sawyer, C.R.; Hudson, C.L.

    1986-01-01

    The electron linear accelerator at EG and G/EM, Santa Barbara Operations, installed in 1963, has been subsequently modified to produce short, intense beam pulses used in the test, calibration and development of many types of fast radiation detectors and systems. The first practical use of the single RF pulse operation, now used in many accelerators, was demonstrated on this accelerator in the late 60s. A major three-year modification, to replace obsolete equipment and bring all the subsystems up to the current state of the art, has increased the beam intensity, stability and reliability. These modifications are discussed

  16. Microscale surface modifications for heat transfer enhancement.

    Science.gov (United States)

    Bostanci, Huseyin; Singh, Virendra; Kizito, John P; Rini, Daniel P; Seal, Sudipta; Chow, Louis C

    2013-10-09

    In this experimental study, two surface modification techniques were investigated for their effect on heat transfer enhancement. One of the methods employed the particle (grit) blasting to create microscale indentations, while the other used plasma spray coating to create microscale protrusions on Al 6061 (aluminum alloy 6061) samples. The test surfaces were characterized using scanning electron microscopy (SEM) and confocal scanning laser microscopy. Because of the surface modifications, the actual surface area was increased up to 2.8× compared to the projected base area, and the arithmetic mean roughness value (Ra) was determined to vary from 0.3 μm for the reference smooth surface to 19.5 μm for the modified surfaces. Selected samples with modified surfaces along with the reference smooth surface were then evaluated for their heat transfer performance in spray cooling tests. The cooling system had vapor-atomizing nozzles and used anhydrous ammonia as the coolant in order to achieve heat fluxes up to 500 W/cm(2) representing a thermal management setting for high power systems. Experimental results showed that the microscale surface modifications enhanced heat transfer coefficients up to 76% at 500 W/cm(2) compared to the smooth surface and demonstrated the benefits of these practical surface modification techniques to enhance two-phase heat transfer process.

  17. Similarity Arguments in the Genetic Modification Debate

    DEFF Research Database (Denmark)

    Christiansen, Andreas

    2017-01-01

    In the ethical debate on genetic modification (GM), it is common to encounter the claim that some anti-GM argument would also apply an established, ethically accepted technology, and that the anti-GM argument is therefore unsuccessful. The paper discusses whether this argumentative strategy, the ...

  18. A Logic of Modification and Creation

    NARCIS (Netherlands)

    Renardel de Lavalette, Gerard R.

    2001-01-01

    In this paper, MCL (modification and creation logic) is presented, a variant of quantified dynamic logic (QDL) with enhanced expressivity. In MCL, functions and predicates can be modified by actions f := λx.t and p := λx.φ, respectively, and new objects can be created by the action Create. This

  19. Surface Modification Of Implants For Bone Surgery

    Directory of Open Access Journals (Sweden)

    Marciniak J.

    2015-09-01

    Full Text Available The study discusses the methods of surface modification methods for AISAI 316 L steel and Ti6Al4V ELI titanium alloy, dedicated to complex design implants used in bone surgery. Results of structural tests have been presented along with those evaluating the physicochemical properties of the formed surface layers. Clinical feasibility of the surface layers has also been evaluated.

  20. [Advances in microbial genome reduction and modification].

    Science.gov (United States)

    Wang, Jianli; Wang, Xiaoyuan

    2013-08-01

    Microbial genome reduction and modification are important strategies for constructing cellular chassis used for synthetic biology. This article summarized the essential genes and the methods to identify them in microorganisms, compared various strategies for microbial genome reduction, and analyzed the characteristics of some microorganisms with the minimized genome. This review shows the important role of genome reduction in constructing cellular chassis.