Journal of proteomics

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  • Impact factor
    5.07
  • 5-year impact
    0.00
  • Cited half-life
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  • Immediacy index
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  • ISSN
    1876-7737

Publications in this journal

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    ABSTRACT: Seed priming with ascorbic acid improves salt tolerance in durum wheat. For understanding the potential mechanisms underlying this priming effect a gel-free shotgun proteomic analysis was performed comparing unprimed to ascorbate-primed wheat seed during germination under saline and non-saline conditions. Since seed germination is the result of interplay or cross-talk between embryo and embryo-surrounding tissues, we studied the variation of metabolic proteome in both tissues separately. 167 of 697 identified and 69 of 471 identified proteins increase or decrease in abundance significantly in response to priming and/or salinity compared to untreated, unstressed control in embryo and embryo-surrounding tissues, respectively. In untreated wheat embryo salt stress was accompanied by change in 129 proteins, most of which are belonging to metabolism, energy, disease/defense, protein destination and storage categories. Ascorbate pretreatment prevents and counteracts the effects of salinity upon most of these proteins and changes specifically the abundance of 35 others proteins, most of which are involved in metabolism, protein destination and storage categories. Hierarchical clustering analysis revealed three and two major clusters of protein expression in embryo and embryo-surrounding tissues, respectively. This study opens promising new avenues to understand priming-induced salt tolerance in plants.
    Journal of proteomics 05/2014;
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    ABSTRACT: During fertilization in plants, pollen grains germinate and generate a pollen tube which grows through the style tissue to the egg apparatus delivering the two sperm cells for fertilization. For this process, adaption to specific environmental conditions and communication between male and female organs are essential, requiring sensing of internal and external signals which are translated into tube growth. The plasma membrane (PM) H(+) ATPase energises the pollen plasma membrane for nutrient, ion and water uptake, but additionally, its activity directly affects the germination frequency and drives elongation of pollen tubes. A combination of in vivo cross-linking with para-formaldehyde, immunoaffinity purification of cross-linked PMH(+) ATPase complexes and subsequent mass spectrometry analysis revealed putative interaction partners of the PMH(+) ATPase of lily pollen, which are possibly involved in the perception and transduction of intra- and extracellular signals. Major interactions partners included (i) membrane-localised receptor-like kinases (RLKs) with the leucine-rich repeat RLKs (LRR-RLKs) forming the largest group, (ii) interacting protein kinases, phosphatases, WD-40 domain proteins and 14-3-3 proteins that may transduce intracellular, phosphorylation-dependent signals and (iii) specific cytosolic Ca(2+) signatures may be decoded by interacting Ca(2+) sensor proteins, calmodulin and calmodulin-like proteins, and Ca(2+)-dependent protein kinases, which were all identified as interaction partners of the PMH(+) ATPase in lily pollen. These identified interaction partners suggest new putative regulation mechanisms of the PMH(+) ATPase in general and new insights in regulating pollen tube growth rates in particular. Furthermore, the optimised experimental strategy can be applied to other non-model organisms to identify membrane protein interactions. Membrane proteomics is still very challenging due to the low abundance and poor solubility of membrane proteins. Furthermore, membrane protein interaction studies in a non-model organism like Lilium longiflorum require an unbiased preparation and detection approach. The presented strategy to identify putative interaction partners of the PMH(+) ATPase by using a combination of different biochemical techniques, i.e.in vivo crosslinking, immunoaffinity purification and mass spectrometry without the need of genetic engineering, transformation or other molecular biology techniques can be easily transferred to other protein interaction studies. The well characterised interaction of the PMH(+) ATPase with regulating 14-3-3 proteins served as an intrinsic control to proof the suitability and reliability of the presented strategy, while newly identified interaction partners may indicate novel regulation mechanisms of the PMH(+) ATPase.
    Journal of proteomics 05/2014;
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    ABSTRACT: Combined RNA-Seq and proteomics analyses reveal striking differential expression of splice isoforms of key proteins in important cancer pathways and networks. Even between primary tumor cell lines from histologically-similar inflammatory breast cancers, we find striking differences in hormone receptor-negative cell lines that are ERBB2 (Her2/neu)-amplified versus ERBB1 (EGFR) over-expressed with low ERBB2 activity. We have related these findings to protein-protein interaction networks, signaling and metabolic pathways, and methods for predicting functional variants among multiple alternative isoforms. Understanding the upstream ligands and regulators and the downstream pathways and interaction networks for ERBB receptors is certain to be important for explanation and prediction of the variable levels of expression and therapeutic responses of ERBB+tumors in the breast and in other organ sites. Alternative splicing is a remarkable evolutionary development that increases protein diversity from multi-exonic genes without requiring expansion of the genome. It is no longer sufficient to report up- or down-expression of genes and proteins without dissecting the complexity due to alternative splicing. This article is part of a Special Issue entitled: 20years of Proteomics.
    Journal of proteomics 05/2014;
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    ABSTRACT: Protein disulfide-isomerase (PDI) is a four-domain flexible protein that catalyzes the formation of disulfide bonds in the endoplasmic reticulum. Here we have analyzed native PDI purified from human placenta by chemical cross-linking followed by mass spectrometry (CXMS). In addition to PDI the sample contained soluble calnexin and ERp72. Extensive cross-linking was observed within the PDI molecule, both intra- and inter-domain, as well as between the different components in the mixture. The high sensitivity of the analysis in the current experiments, combined with a likely promiscuous interaction pattern of the involved proteins, revealed relatively densely populated cross-link heat maps. The established X-ray structure of the monomeric PDI could be confirmed; however, the dimer as presented in the existing models do not seem to be prevalent in solution as modeling on the observed cross-links revealed new models of dimeric PDI. The observed inter-protein cross-links confirmed the existence of a peptide binding area on calnexin that binds strongly both PDI and ERp72. On the other hand, interaction sites on PDI and ERp72 could not be uniquely identified, indicating a more non-specific interaction pattern. Biological significance The present work demonstrates the use of chemical cross-linking and mass spectrometry (CXMS) for the determination of a solution structure of natural human PDI and its interaction with the chaperones ERp72 and calnexin. The data shows that the dimeric structure of PDI may be more diverse than indicated by present models. We further observe that the temperature influences the cross-linking pattern of PDI, but this does not influence the overall folding pattern of the molecule.
    Journal of proteomics 05/2014;
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    ABSTRACT: Although the primary physiological effects produced by scorpion toxins are already well known, most of the secondary molecular events following scorpion neurotoxins-ion channels interactions are poorly understood and described. For this reason, we used a proteomic approach to determine the changes in relative protein abundance in F11 mouse neuroblastoma cells treated with Cn2, the major β-toxin from the venom of the scorpion Centruroides noxius Hoffmann. Here we show that the relative abundance of 24 proteins changed after Cn2 treatment. Proteins related to protection from apoptosis and cell survival, as well as those involved in cell morphology and some translation elongation factors were diminished. By contrast, proteins associated with energy metabolism were increased. Additionally, results of western immunoblots confirmed the preference of action towards some special targets. These results suggest that Cn2 could modify the neuronal structure and induce apoptosis and reduction of the proliferation and cell survival. To support this conclusion we directly measured the Cn2 effect on cell proliferation, division and apoptosis. Our results open new avenues for continuying the studies aimed at better understanding the envenomation process caused by scorpion stings. Biological significance The purpose of this work was to identify which proteins, apart from the ion-channels, are involved in the envenomation process in order to develop possible strategies to circumvent the deleterious effects caused by the toxic peptides of the venom. For this reason, we characterized the early changes in the proteome of F11 cells induced by Cn2, the major toxin of the New World scorpion Centruroides noxius Hoffmann, using 2D-PAGE and LC-MS/MS. We identified 24 proteins which relative abundance is modified after the Cn2 treatment. Among these, proteins related with apoptosis protection, cell survival, neuronal morphology and some translation elongation factors were diminished, whereas proteins associated with energy metabolism were increased.
    Journal of proteomics 05/2014;
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    ABSTRACT: The study innovatively pinpoints target proteins of carbonylation, a key PTM induced by oxidative stress, in the SHROB (genetically obese spontaneously hypertensive) rat model of metabolic syndrome (MetS). Protein carbonylation was assessed by a fluorescence-labelling proteomics approach, and complemented with biometric and biochemical markers of MetS. SHROB and healthy Wistar rats were fed two diets, soybean and linseed oil supplementations, in order to distinguish intrinsic carbonylation of SHROB animals from diet-modulated carbonylation unrelated to MetS. First exploratory data showed similar carbonylation patterns and metabolic conditions in SHROB rats fed soybean and linseed, but different from Wistar animals. A total of 18 carbonylated spots in liver, and 12 in skeletal tissue, related to pathways of lipid (29.6%), carbohydrate (25.9%) and amino acid (18.5%) metabolisms, were identified. In particular, SHROB animals present higher carbonylation in four liver proteins belonging to lipid metabolism, redox regulation and chaperone activity (ALDH2, PDI, PDIA3, PECR), and in the skeletal muscle ALDOA involved in muscle dysfunction. Conversely, SHROB rats display lower carbonylation in liver albumin, AKR1C9, ADH1 and catalase. This investigation provides a novel perspective of carbonylation in the context of metabolic disorders, and may be a starting point to characterize new redox pathways exacerbating MetS. Oxidative stress is a concomitant factor in the pathogenesis of MetS that induces oxidative PTM as carbonylation. Through the use a redox proteomics approach, we have thoroughly mapped the occurrence of protein targets of carbonylation in the genetically-induced MetS model SHROB rat. The present research brings a new insight of MetS pathogenesis and it may provide valuable information to understand the biological impact of oxidative stress in patients with MetS.
    Journal of proteomics 04/2014;
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    ABSTRACT: Comparisons of proteomic responses of closely related congeners and populations have shown which cellular processes are critical to adapt to environmental stress. For example, several proteomic species comparisons showed that increasing abundances of oxidative stress proteins indicate that reactive oxygen species (ROS) represent a ubiquitous signal and possible co-stressor of warm and cold temperature, acute hyposaline and low pH stress, possibly causing a shift from pro-oxidant NADH-producing to anti-oxidant NADPH-producing and -consuming metabolic pathways. Changes in cytoskeletal and actin-binding proteins in response to several stressors, including ROS, suggest that both are important structural and functional elements in responding to stress. Disruption of protein homeostasis, e.g., increased abundance of molecular chaperones, was severe in response to acute heat stress, inducing proteolysis, but was also observed in response to chronic heat and cold stress and was concentrated to the endoplasmic reticulum during hyposaline stress. Small GTPases affecting vesicle formation and transport, Ca(2+)-signaling and ion transport responded to salinity stress in species- and population-specific ways. Aerobic energy metabolism was in general down-regulated in response to temperature, hypoxia, hyposalinity and low pH stress, but other metabolic pathways were activated to respond to increased oxidative stress or to switch metabolic fuels. Thus, comparative proteomics is a powerful approach to identify functionally adaptive variation. This article is part of a Special Issue entitled: Proteomics of non-model organisms.
    Journal of proteomics 04/2014;
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    ABSTRACT: This communication reports the results of proteomic, transcriptomic, biochemical and electrophysiological analysis of the soluble venom and venom glands of the Mexican centipede Scolopendra viridis Say (here thereafter abbreviated S. viridis). Separation of the soluble venom permitted to obtain 54 different fractions, from which a mass finger printing analysis permitted the identification of at least 86 components, where 70% of the molecules have low molecular masses. Two-dimensional electrophoretic separation of this venom revealed the presence of about forty proteins with molecular weights ranging from 17 to 58kDa. The novo sequencing of 149 peptides obtained by LC-MS/MS from the 2D-gels showed the presence of proteins with amino acid sequences similar to several enzymes and venom allergens type 3. Furthermore, a total of 180 sequences were obtained from a cDNA library prepared with two venomous glands. From this, 155 sequences correspond to complete genes containing more than 200 base pairs each. Comparative sequence analyses of these sequences indicated the presence of different types of enzymes and toxin-like genes. Two proteins with molecular weights around 37,000 and 42,000Da were shown to contain hyaluronidase activity. Electrophysiological assays performed with soluble venom show that it decreases mammalian sodium channel currents. Animal venoms of Scolopendra species have been scarcely studied, although they have been reported to contain several bioactive compounds, some of which with potential therapeutic interest. The Mexican centipede S. viridis contains a powerful venom, capable of inflicting immediate effects on their preys. This communication is focused on the identification and description of a proteomic and transcriptomic analysis of the protein components of this venom. The presence of several amino acid sequences similar to reported enzymes are the principal components in the S. viridis venom, but also a low number of toxins were identified. This knowledge should contribute to the understanding of the pharmacological effects caused by bites of this centipide species. This article is part of a Special Issue entitled: SMP Cancun 2013.
    Journal of proteomics 04/2014;
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    ABSTRACT: To investigate quantitative differences in aberrant glycosylation of target glycoproteins between noncancerous group and patient group with adenocarcinoma lung cancer (ADLC), differential proteomic approach was developed by cooperatively using comparative lectin-capturing, targeted mass spectrometry (MRM MS), and antibody/lectin sandwich ELISA. Plasma samples comparatively prepared from 3 ADLC patients and 3 controls, with and without lectin-fractionation using fucose-specific Aleuria aurantia lectin (AAL), were trypsin-digested and analyzed for target glycoproteins, alpha-1-acid glycoprotein (AGP) and ceruloplasmin (CP), by MRM MS. From the MRM MS data the abundance levels of AAL-captured glycoforms of both targets were significantly higher in ADLC cases compared to controls, although the levels in total protein abundance were comparable between ADLC and control groups. This difference between ADLC and control groups in the fucosylated glycoform levels was originated mainly from aberrant fucosylation on the targets in ADLC plasmas rather than change in total protein abundance of the targets, and also confirmed by sandwich ELISA. AGP and CP were further verified to be biomarker candidates by MRM-based analysis of AAL-captured plasmas (30 ADLC cases, 30 controls), with AUROC 0.758 and 0.847 respectively. This differential proteomic approach can be useful for identifying and verifying biomarker candidate involved in aberrant protein glycosylation. The present paper introduces an efficient differential proteomic method to investigate quantitative differences in aberrant protein glycosylation of serological glycoproteins between noncancerous group and lung cancer patient group. This differential proteomic approach consisting of the targeted MRM MS of comparatively lectin-captured plasma fractions and the antibody/lectin sandwich ELISA-based assay was evaluated to be useful for identification of aberrantly fucosylated glycoproteins AGP and CP in lung cancer plasmas. In addition, we have demonstrated that the MRM MS-based differential proteomic approach is also useful for high-throughput verification of the aberrantly fucosylated glycoproteins AGP and CP using the large number of individual plasmas. Therefore, the present MRM MS-based differential proteomic strategy with lectin-capturing can be a powerful tool for high-throughput verification of aberrantly glycosylated biomarker candidates, identified preliminary by mass profiling experiments in proteomic fields but requiring further validation using a large number of cohorts.
    Journal of proteomics 04/2014;
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    ABSTRACT: Lipoprotein-associated proteins form an intrinsic part of the major plasma lipoprotein classes. There is increasing evidence that the quantity of these proteins per lipoprotein particle determines lipoprotein function including redox, inflammatory and thrombotic properties and may impact on lipoprotein-related risks for developing heart disease. However, only limited information on the relative quantity of these proteins has been published and no comprehensive absolute quantitative data providing the stoichiometry of proteins associated with lipoproteins is available to date. To address this, we performed extensive absolute quantification by mass spectrometry of 17 lipoprotein-associated proteins on VLDL, LDL, Lp(a) and HDL from healthy subjects. For the first time we show the exact stoichiometry of apolipoproteins on different lipoprotein classes. The most distinct differences were seen in the abundance of all apoCs, apoE and apoF. We further revealed strong variations between individual samples, which indicates the complexity of the protein complement of lipoproteins and can provide additional insights into lipoprotein-related risk factors. This approach has the potential to determine alterations in the protein profiles of lipoproteins in disease states such as CVD or diabetes and, if performed on large cohorts, to translate into a tool for identifying new candidate biomarkers for risk of disease. A more comprehensive picture about the protein complement on individual lipoprotein classes is the goal of lipoprotein proteomics analyses. Despite many such studies, there is a lack of absolute quantitative data on lipoproteins isolated from individual subjects. The stoichiometry of lipoprotein-associated proteins rather than their presence or absence could provide insights into an individual's predisposition for disease such as heart disease or diabetes. Our study provides a comprehensive overview of the absolute quantity of proteins on the major apolipoprotein classes VLDL, LDL, Lp(a) and HDL.
    Journal of proteomics 04/2014;
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    ABSTRACT: The scorpion Mesobuthus martensii is the most populous species in eastern Asian countries, and several toxic components have been identified from their venoms. Nevertheless, a complete proteomic profile of the venom of M. martensii is still not available. In this study, the venom of M. martensii was analyzed by comprehensive proteomic approaches. 153 fractions were isolated from the M. martensii venom by 2-DE, SDS-PAGE and RP-HPLC. The ESI-Q-TOF MS results of all fractions were used to search the scorpion genomic and transcriptomic databases. Totally, 227 non-redundant protein sequences were unambiguously identified, composed of 134 previously known and 93 previously unknown proteins. Among 134 previously known proteins, 115 proteins were firstly confirmed from the M. martensii crude venom and 19 toxins were confirmed once again, involving 43 typical toxins, 7 atypical toxins, 12 venom enzymes and 72 cell associated proteins. In typical toxins, 7 novel-toxin sequences were identified, including 3 Na(+)-channel toxins, 3K(+)-channel toxins and 1 no-annotation toxin. These results increased 230% (115/50) venom components compared with previous studies from the M. martensii venom, especially 50% (24/48) typical toxins. Additionally, a mass fingerprint obtained by MALDI-TOF MS indicated that the scorpion venom contained more than 200 different molecular masses components. This work firstly gave a systematic investigation of the M. martensii venom by combined proteomics strategy coupled with genomics and transcriptomics. A large number of protein components were unambiguously identified from the venom of M. martensii, most of which were confirmed for the first time. We also contributed 7 novel-toxin sequences and 93 protein sequences previously unknown to be part of the venom, for which we assigned potential biological functions. Besides, we obtained a mass fingerprint of the M. martensii venom. Together, our study not only provides the most comprehensive catalogue of the molecular diversity of the M. martensii venom at the proteomic level, but also enriches the composition information of scorpion venom.
    Journal of proteomics 04/2014;
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    ABSTRACT: Schistosomes are blood trematodes that are perfectly adapted to living in their intravascular habitat and to achieve this they have developed mechanisms enabling them to evade the immune and haemostatic responses of the host and to regulate endothelial cell function to favour their own survival. The objective of this work was to analyse the changes induced by Schistosoma bovis schistosomula in the proteome expressed by infected hamsters, over 10 and 20days, on the endothelial surface of their pulmonary vasculature. To accomplish this, we subjected the lungs of non-infected and S. bovis-infected hamsters to vascular perfusion with a biotin ester reactive. Analysis by liquid chromatography and tandem mass spectrometry analysis (LC-MS/MS) of endothelial surface proteins resulted in the identification of a total of 459 non-redundant proteins in the lung vasculature of infected and non-infected hamsters. Here we report the proteins identified, classified according to their biological function and cellular location, further analysing the differences in lung vascular proteomes between non-infected and S. bovis-infected hamsters for ten and twenty days. This work provides the first data on the vascular surface proteome of the lung after S. bovis infection and identifies some of the changes induced in it during infection. To identify the changes induced by schistosomula larvae of Schistosoma bovis in the proteome of the pulmonary vasculature of the host, we compared the proteins expressed on the vascular endothelium of the lungs of non-infected and infected hamsters over 10 and 20 days. Mass spectrometry analysis (LC-MS/MS) of the proteins isolated from the vascular endothelium resulted in the identification of a total of 459 non-redundant proteins in the lung of infected and non-infected hamsters. The proteins identified are classified according to their biological function and cellular location, further analysing the differences in lung vascular proteomes between non-infected and S. bovis-infected hamsters. This work provides the first data on the vascular surface proteome of the lung after S. bovis infection and identifies some of the changes induced in it during infection suggesting the possible involvement of these proteins during parasite infection.
    Journal of proteomics 04/2014;
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    ABSTRACT: Chronic hepatitis B virus (HBV) infection is a major risk factor for hepatocellular carcinoma (HCC), the sixth most common cancer worldwide. To explore potential biomarkers for HCC, iTRAQ coupled with mass spectrometry was used to analyze proteins in plasma from individuals with HBV-associated HCC, nonmalignant cirrhosis, chronic hepatitis B, and healthy individuals. Twenty-one aberrantly expressed proteins were identified from HCC patients as compared with nontumor controls. Overexpression of von Willebrand factor (vWF) was confirmed by Western blotting, and immunohistochemical analysis from liver biopsies and ELISA from plasma samples revealed a correlation between vWF expression and HCC clinicopathologic staging. Furthermore, siRNA-induced vWF silencing reduced HBV replication by over two-fold via the interferon-signaling pathway and impaired the invasion and migration of HCC cells in vitro. These results indicate that vWF can serve as a biomarker, and perhaps an alternative target for therapeutic intervention of HCC progression and HBV viral infection. We report comparative plasma proteome profiles of HBV-associated HCC and nonmalignant chronic liver diseases, including chronic hepatitis B and cirrhosis. The quantification of these datasets showed altered abundance of 21 proteins in HBV-related HCC and provides a reference point for future applied and basic research. In addition, we have demonstrated that the candidate protein vWF is involved in the pathogenesis of HBV infection and replication, and also associated with clinicopathologic staging of HCC patients with HBV infection. Overall these findings provide information on the mechanism of HCC development, which may assist in the development of novel cancer and HBV therapeutic drugs.
    Journal of proteomics 04/2014;
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    ABSTRACT: Extracellular vesicles are produced by many pathogenic microorganisms and have varied functions that include secretion and release of microbial factors, which contribute to virulence. Very little is known about vesicle production by Gram-positive bacteria, as well as their biogenesis and release mechanisms. In this work, we demonstrate the active production of vesicles by Streptococcus pneumoniae from the plasma membrane, rather than being a product from cell lysis. We biochemically characterized them by proteomics and fatty acid analysis, showing that these vesicles and the plasma membrane resemble in essential aspects, but have some differences: vesicles are more enriched in lipoproteins and short-chain fatty acids. We also demonstrate that these vesicles act as carriers of surface proteins and virulence factors. They are also highly immunoreactive against human sera and induce immune responses that protect against infection. Overall, this work provides insights into the biology of this important Gram-positive human pathogen and the role of extracellular vesicles in clinical applications. Pneumococcus is one of the leading causes of bacterial pneumonia worldwide in children and the elderly, being responsible for high morbidity and mortality rates in developing countries. The augment of pneumococcal disease in developed countries has raised major public health concern, since the difficulties to treat these infections due to increasing antibiotic resistance. Vaccination is still the best way to combat pneumococcal infections. One of the mechanisms that bacterial pathogens use to combat the defense responses of invaded hosts is the production and release of extracellular vesicles derived from the outer surface. Little is known about this phenomenon in Gram-positives. We show that pneumococcus produces membrane-derived vesicles particularly enriched in lipoproteins. We also show the utility of pneumococcal vesicles as a new type of vaccine, as they induce protection in immunized mice against infection with a virulent strain. This work will contribute to understand the role of these structures in important biological processes such as host-pathogen interactions and prevention of human disease.
    Journal of proteomics 04/2014;