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Sample records for integrated proteomic analysis

  1. Integrative analysis of the mitochondrial proteome in yeast.

    Directory of Open Access Journals (Sweden)

    Holger Prokisch

    2004-06-01

    Full Text Available In this study yeast mitochondria were used as a model system to apply, evaluate, and integrate different genomic approaches to define the proteins of an organelle. Liquid chromatography mass spectrometry applied to purified mitochondria identified 546 proteins. By expression analysis and comparison to other proteome studies, we demonstrate that the proteomic approach identifies primarily highly abundant proteins. By expanding our evaluation to other types of genomic approaches, including systematic deletion phenotype screening, expression profiling, subcellular localization studies, protein interaction analyses, and computational predictions, we show that an integration of approaches moves beyond the limitations of any single approach. We report the success of each approach by benchmarking it against a reference set of known mitochondrial proteins, and predict approximately 700 proteins associated with the mitochondrial organelle from the integration of 22 datasets. We show that a combination of complementary approaches like deletion phenotype screening and mass spectrometry can identify over 75% of the known mitochondrial proteome. These findings have implications for choosing optimal genome-wide approaches for the study of other cellular systems, including organelles and pathways in various species. Furthermore, our systematic identification of genes involved in mitochondrial function and biogenesis in yeast expands the candidate genes available for mapping Mendelian and complex mitochondrial disorders in humans.

  2. Integrated proteomic and genomic analysis of colorectal cancer

    Science.gov (United States)

    Investigators who analyzed 95 human colorectal tumor samples have determined how gene alterations identified in previous analyses of the same samples are expressed at the protein level. The integration of proteomic and genomic data, or proteogenomics, pro

  3. Quantitative and qualitative proteome characteristics extracted from in-depth integrated genomics and proteomics analysis

    NARCIS (Netherlands)

    Low, T.Y.; van Heesch, S.; van den Toorn, H.; Giansanti, P.; Cristobal, A.; Toonen, P.; Schafer, S.; Hubner, N.; van Breukelen, B.; Mohammed, S.; Cuppen, E.; Heck, A.J.R.; Guryev, V.

    2013-01-01

    Quantitative and qualitative protein characteristics are regulated at genomic, transcriptomic, and posttranscriptional levels. Here, we integrated in-depth transcriptome and proteome analyses of liver tissues from two rat strains to unravel the interactions within and between these layers. We

  4. Quantitative and Qualitative Proteome Characteristics Extracted from In-Depth Integrated Genomics and Proteomics Analysis

    NARCIS (Netherlands)

    Low, Teck Yew; van Heesch, Sebastiaan; van den Toorn, Henk; Giansanti, Piero; Cristobal, Alba; Toonen, Pim; Schafer, Sebastian; Huebner, Norbert; van Breukelen, Bas; Mohammed, Shabaz; Cuppen, Edwin; Heck, Albert J. R.; Guryev, Victor

    2013-01-01

    Quantitative and qualitative protein characteristics are regulated at genomic, transcriptomic, and post-transcriptional levels. Here, we integrated in-depth transcriptome and proteome analyses of liver tissues from two rat strains to unravel the interactions within and between these layers. We

  5. Integration of gel-based and gel-free proteomic data for functional analysis of proteins through Soybean Proteome Database

    KAUST Repository

    Komatsu, Setsuko

    2017-05-10

    The Soybean Proteome Database (SPD) stores data on soybean proteins obtained with gel-based and gel-free proteomic techniques. The database was constructed to provide information on proteins for functional analyses. The majority of the data is focused on soybean (Glycine max ‘Enrei’). The growth and yield of soybean are strongly affected by environmental stresses such as flooding. The database was originally constructed using data on soybean proteins separated by two-dimensional polyacrylamide gel electrophoresis, which is a gel-based proteomic technique. Since 2015, the database has been expanded to incorporate data obtained by label-free mass spectrometry-based quantitative proteomics, which is a gel-free proteomic technique. Here, the portions of the database consisting of gel-free proteomic data are described. The gel-free proteomic database contains 39,212 proteins identified in 63 sample sets, such as temporal and organ-specific samples of soybean plants grown under flooding stress or non-stressed conditions. In addition, data on organellar proteins identified in mitochondria, nuclei, and endoplasmic reticulum are stored. Furthermore, the database integrates multiple omics data such as genomics, transcriptomics, metabolomics, and proteomics. The SPD database is accessible at http://proteome.dc.affrc.go.jp/Soybean/. Biological significanceThe Soybean Proteome Database stores data obtained from both gel-based and gel-free proteomic techniques. The gel-free proteomic database comprises 39,212 proteins identified in 63 sample sets, such as different organs of soybean plants grown under flooding stress or non-stressed conditions in a time-dependent manner. In addition, organellar proteins identified in mitochondria, nuclei, and endoplasmic reticulum are stored in the gel-free proteomics database. A total of 44,704 proteins, including 5490 proteins identified using a gel-based proteomic technique, are stored in the SPD. It accounts for approximately 80% of all

  6. Integration of gel-based and gel-free proteomic data for functional analysis of proteins through Soybean Proteome Database.

    Science.gov (United States)

    Komatsu, Setsuko; Wang, Xin; Yin, Xiaojian; Nanjo, Yohei; Ohyanagi, Hajime; Sakata, Katsumi

    2017-06-23

    The Soybean Proteome Database (SPD) stores data on soybean proteins obtained with gel-based and gel-free proteomic techniques. The database was constructed to provide information on proteins for functional analyses. The majority of the data is focused on soybean (Glycine max 'Enrei'). The growth and yield of soybean are strongly affected by environmental stresses such as flooding. The database was originally constructed using data on soybean proteins separated by two-dimensional polyacrylamide gel electrophoresis, which is a gel-based proteomic technique. Since 2015, the database has been expanded to incorporate data obtained by label-free mass spectrometry-based quantitative proteomics, which is a gel-free proteomic technique. Here, the portions of the database consisting of gel-free proteomic data are described. The gel-free proteomic database contains 39,212 proteins identified in 63 sample sets, such as temporal and organ-specific samples of soybean plants grown under flooding stress or non-stressed conditions. In addition, data on organellar proteins identified in mitochondria, nuclei, and endoplasmic reticulum are stored. Furthermore, the database integrates multiple omics data such as genomics, transcriptomics, metabolomics, and proteomics. The SPD database is accessible at http://proteome.dc.affrc.go.jp/Soybean/. The Soybean Proteome Database stores data obtained from both gel-based and gel-free proteomic techniques. The gel-free proteomic database comprises 39,212 proteins identified in 63 sample sets, such as different organs of soybean plants grown under flooding stress or non-stressed conditions in a time-dependent manner. In addition, organellar proteins identified in mitochondria, nuclei, and endoplasmic reticulum are stored in the gel-free proteomics database. A total of 44,704 proteins, including 5490 proteins identified using a gel-based proteomic technique, are stored in the SPD. It accounts for approximately 80% of all predicted proteins from

  7. Integration of gel-based and gel-free proteomic data for functional analysis of proteins through Soybean Proteome Database

    KAUST Repository

    Komatsu, Setsuko; Wang, Xin; Yin, Xiaojian; Nanjo, Yohei; Ohyanagi, Hajime; Sakata, Katsumi

    2017-01-01

    The Soybean Proteome Database (SPD) stores data on soybean proteins obtained with gel-based and gel-free proteomic techniques. The database was constructed to provide information on proteins for functional analyses. The majority of the data is focused on soybean (Glycine max ‘Enrei’). The growth and yield of soybean are strongly affected by environmental stresses such as flooding. The database was originally constructed using data on soybean proteins separated by two-dimensional polyacrylamide gel electrophoresis, which is a gel-based proteomic technique. Since 2015, the database has been expanded to incorporate data obtained by label-free mass spectrometry-based quantitative proteomics, which is a gel-free proteomic technique. Here, the portions of the database consisting of gel-free proteomic data are described. The gel-free proteomic database contains 39,212 proteins identified in 63 sample sets, such as temporal and organ-specific samples of soybean plants grown under flooding stress or non-stressed conditions. In addition, data on organellar proteins identified in mitochondria, nuclei, and endoplasmic reticulum are stored. Furthermore, the database integrates multiple omics data such as genomics, transcriptomics, metabolomics, and proteomics. The SPD database is accessible at http://proteome.dc.affrc.go.jp/Soybean/. Biological significanceThe Soybean Proteome Database stores data obtained from both gel-based and gel-free proteomic techniques. The gel-free proteomic database comprises 39,212 proteins identified in 63 sample sets, such as different organs of soybean plants grown under flooding stress or non-stressed conditions in a time-dependent manner. In addition, organellar proteins identified in mitochondria, nuclei, and endoplasmic reticulum are stored in the gel-free proteomics database. A total of 44,704 proteins, including 5490 proteins identified using a gel-based proteomic technique, are stored in the SPD. It accounts for approximately 80% of all

  8. Integrative Genomic and Proteomic Analysis of the Response of Lactobacillus casei Zhang to Glucose Restriction.

    Science.gov (United States)

    Yu, Jie; Hui, Wenyan; Cao, Chenxia; Pan, Lin; Zhang, Heping; Zhang, Wenyi

    2018-03-02

    Nutrient starvation is an important survival challenge for bacteria during industrial production of functional foods. As next-generation sequencing technology has greatly advanced, we performed proteomic and genomic analysis to investigate the response of Lactobacillus casei Zhang to a glucose-restricted environment. L. casei Zhang strains were permitted to evolve in glucose-restricted or normal medium from a common ancestor over a 3 year period, and they were sampled at 1000, 2000, 3000, 4000, 5000, 6000, 7000, and 8000 generations and subjected to proteomic and genomic analyses. Genomic resequencing data revealed different point mutations and other mutational events in each selected generation of L. casei Zhang under glucose restriction stress. The differentially expressed proteins induced by glucose restriction were mostly related to fructose and mannose metabolism, carbohydrate metabolic processes, lyase activity, and amino-acid-transporting ATPase activity. Integrative proteomic and genomic analysis revealed that the mutations protected L. casei Zhang against glucose starvation by regulating other cellular carbohydrate, fatty acid, and amino acid catabolism; phosphoenolpyruvate system pathway activation; glycogen synthesis; ATP consumption; pyruvate metabolism; and general stress-response protein expression. The results help reveal the mechanisms of adapting to glucose starvation and provide new strategies for enhancing the industrial utility of L. casei Zhang.

  9. iTRAQ-Based Proteomics Analysis and Network Integration for Kernel Tissue Development in Maize

    Science.gov (United States)

    Dong, Yongbin; Wang, Qilei; Du, Chunguang; Xiong, Wenwei; Li, Xinyu; Zhu, Sailan; Li, Yuling

    2017-01-01

    Grain weight is one of the most important yield components and a developmentally complex structure comprised of two major compartments (endosperm and pericarp) in maize (Zea mays L.), however, very little is known concerning the coordinated accumulation of the numerous proteins involved. Herein, we used isobaric tags for relative and absolute quantitation (iTRAQ)-based comparative proteomic method to analyze the characteristics of dynamic proteomics for endosperm and pericarp during grain development. Totally, 9539 proteins were identified for both components at four development stages, among which 1401 proteins were non-redundant, 232 proteins were specific in pericarp and 153 proteins were specific in endosperm. A functional annotation of the identified proteins revealed the importance of metabolic and cellular processes, and binding and catalytic activities for the tissue development. Three and 76 proteins involved in 49 Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways were integrated for the specific endosperm and pericarp proteins, respectively, reflecting their complex metabolic interactions. In addition, four proteins with important functions and different expression levels were chosen for gene cloning and expression analysis. Different concordance between mRNA level and the protein abundance was observed across different proteins, stages, and tissues as in previous research. These results could provide useful message for understanding the developmental mechanisms in grain development in maize. PMID:28837076

  10. Network analysis of epidermal growth factor signaling using integrated genomic, proteomic and phosphorylation data.

    Directory of Open Access Journals (Sweden)

    Katrina M Waters

    Full Text Available To understand how integration of multiple data types can help decipher cellular responses at the systems level, we analyzed the mitogenic response of human mammary epithelial cells to epidermal growth factor (EGF using whole genome microarrays, mass spectrometry-based proteomics and large-scale western blots with over 1000 antibodies. A time course analysis revealed significant differences in the expression of 3172 genes and 596 proteins, including protein phosphorylation changes measured by western blot. Integration of these disparate data types showed that each contributed qualitatively different components to the observed cell response to EGF and that varying degrees of concordance in gene expression and protein abundance measurements could be linked to specific biological processes. Networks inferred from individual data types were relatively limited, whereas networks derived from the integrated data recapitulated the known major cellular responses to EGF and exhibited more highly connected signaling nodes than networks derived from any individual dataset. While cell cycle regulatory pathways were altered as anticipated, we found the most robust response to mitogenic concentrations of EGF was induction of matrix metalloprotease cascades, highlighting the importance of the EGFR system as a regulator of the extracellular environment. These results demonstrate the value of integrating multiple levels of biological information to more accurately reconstruct networks of cellular response.

  11. Network Analysis of Epidermal Growth Factor Signaling using Integrated Genomic, Proteomic and Phosphorylation Data

    Energy Technology Data Exchange (ETDEWEB)

    Waters, Katrina M.; Liu, Tao; Quesenberry, Ryan D.; Willse, Alan R.; Bandyopadhyay, Somnath; Kathmann, Loel E.; Weber, Thomas J.; Smith, Richard D.; Wiley, H. S.; Thrall, Brian D.

    2012-03-29

    To understand how integration of multiple data types can help decipher cellular responses at the systems level, we analyzed the mitogenic response of human mammary epithelial cells to epidermal growth factor (EGF) using whole genome microarrays, mass spectrometry-based proteomics and large-scale western blots with over 1000 antibodies. A time course analysis revealed significant differences in the expression of 3172 genes and 596 proteins, including protein phosphorylation changes measured by western blot. Integration of these disparate data types showed that each contributed qualitatively different components to the observed cell response to EGF and that varying degrees of concordance in gene expression and protein abundance measurements could be linked to specific biological processes. Networks inferred from individual data types were relatively limited, whereas networks derived from the integrated data recapitulated the known major cellular responses to EGF and exhibited more highly connected signaling nodes than networks derived from any individual dataset. While cell cycle regulatory pathways were altered as anticipated, we found the most robust response to mitogenic concentrations of EGF was induction of matrix metalloprotease cascades, highlighting the importance of the EGFR system as a regulator of the extracellular environment. These results demonstrate the value of integrating multiple levels of biological information to more accurately reconstruct networks of cellular response.

  12. A graphical user interface for RAId, a knowledge integrated proteomics analysis suite with accurate statistics.

    Science.gov (United States)

    Joyce, Brendan; Lee, Danny; Rubio, Alex; Ogurtsov, Aleksey; Alves, Gelio; Yu, Yi-Kuo

    2018-03-15

    RAId is a software package that has been actively developed for the past 10 years for computationally and visually analyzing MS/MS data. Founded on rigorous statistical methods, RAId's core program computes accurate E-values for peptides and proteins identified during database searches. Making this robust tool readily accessible for the proteomics community by developing a graphical user interface (GUI) is our main goal here. We have constructed a graphical user interface to facilitate the use of RAId on users' local machines. Written in Java, RAId_GUI not only makes easy executions of RAId but also provides tools for data/spectra visualization, MS-product analysis, molecular isotopic distribution analysis, and graphing the retrieval versus the proportion of false discoveries. The results viewer displays and allows the users to download the analyses results. Both the knowledge-integrated organismal databases and the code package (containing source code, the graphical user interface, and a user manual) are available for download at https://www.ncbi.nlm.nih.gov/CBBresearch/Yu/downloads/raid.html .

  13. Integrative Analysis of Subcellular Quantitative Proteomics Studies Reveals Functional Cytoskeleton Membrane-Lipid Raft Interactions in Cancer.

    Science.gov (United States)

    Shah, Anup D; Inder, Kerry L; Shah, Alok K; Cristino, Alexandre S; McKie, Arthur B; Gabra, Hani; Davis, Melissa J; Hill, Michelle M

    2016-10-07

    Lipid rafts are dynamic membrane microdomains that orchestrate molecular interactions and are implicated in cancer development. To understand the functions of lipid rafts in cancer, we performed an integrated analysis of quantitative lipid raft proteomics data sets modeling progression in breast cancer, melanoma, and renal cell carcinoma. This analysis revealed that cancer development is associated with increased membrane raft-cytoskeleton interactions, with ∼40% of elevated lipid raft proteins being cytoskeletal components. Previous studies suggest a potential functional role for the raft-cytoskeleton in the action of the putative tumor suppressors PTRF/Cavin-1 and Merlin. To extend the observation, we examined lipid raft proteome modulation by an unrelated tumor suppressor opioid binding protein cell-adhesion molecule (OPCML) in ovarian cancer SKOV3 cells. In agreement with the other model systems, quantitative proteomics revealed that 39% of OPCML-depleted lipid raft proteins are cytoskeletal components, with microfilaments and intermediate filaments specifically down-regulated. Furthermore, protein-protein interaction network and simulation analysis showed significantly higher interactions among cancer raft proteins compared with general human raft proteins. Collectively, these results suggest increased cytoskeleton-mediated stabilization of lipid raft domains with greater molecular interactions as a common, functional, and reversible feature of cancer cells.

  14. Intrauterine Growth Restriction Programs the Hypothalamus of Adult Male Rats: Integrated Analysis of Proteomic and Metabolomic Data.

    Science.gov (United States)

    Pedroso, Amanda P; Souza, Adriana P; Dornellas, Ana P S; Oyama, Lila M; Nascimento, Cláudia M O; Santos, Gianni M S; Rosa, José C; Bertolla, Ricardo P; Klawitter, Jelena; Christians, Uwe; Tashima, Alexandre K; Ribeiro, Eliane B

    2017-04-07

    Programming of hypothalamic functions regulating energy homeostasis may play a role in intrauterine growth restriction (IUGR)-induced adulthood obesity. The present study investigated the effects of IUGR on the hypothalamus proteome and metabolome of adult rats submitted to 50% protein-energy restriction throughout pregnancy. Proteomic and metabolomic analyzes were performed by data independent acquisition mass spectrometry and multiple reaction monitoring, respectively. At age 4 months, the restricted rats showed elevated adiposity, increased leptin and signs of insulin resistance. 1356 proteins were identified and 348 quantified while 127 metabolites were quantified. The restricted hypothalamus showed down-regulation of 36 proteins and 5 metabolites and up-regulation of 21 proteins and 9 metabolites. Integrated pathway analysis of the proteomics and metabolomics data indicated impairment of hypothalamic glucose metabolism, increased flux through the hexosamine pathway, deregulation of TCA cycle and the respiratory chain, and alterations in glutathione metabolism. The data suggest IUGR modulation of energy metabolism and redox homeostasis in the hypothalamus of male adult rats. The present results indicated deleterious consequences of IUGR on hypothalamic pathways involved in pivotal physiological functions. These results provide guidance for future mechanistic studies assessing the role of intrauterine malnutrition in the development of metabolic diseases later in life.

  15. Integrated transcriptomic and proteomic analysis of the bile stress response in probiotic Lactobacillus salivarius LI01.

    Science.gov (United States)

    Lv, Long-Xian; Yan, Ren; Shi, Hai-Yan; Shi, Ding; Fang, Dai-Qiong; Jiang, Hui-Yong; Wu, Wen-Rui; Guo, Fei-Fei; Jiang, Xia-Wei; Gu, Si-Lan; Chen, Yun-Bo; Yao, Jian; Li, Lan-Juan

    2017-01-06

    Lactobacillus salivarius LI01, isolated from healthy humans, has demonstrated probiotic properties in the prevention and treatment of liver failure. Tolerance to bile stress is crucial to allow lactobacilli to survive in the gastrointestinal tract and exert their benefits. In this work, we used a Digital Gene Expression transcriptomic and iTRAQ LC-MS/MS proteomic approach to examine the characteristics of LI01 in response to bile stress. Using culture medium with or without 0.15% ox bile, 591 differentially transcribed genes and 347 differentially expressed proteins were detected in LI01. Overall, we found the bile resistance of LI01 to be based on a highly remodeled cell envelope and a reinforced bile efflux system rather than on the activity of bile salt hydrolases. Additionally, some differentially expressed genes related to regulatory systems, the general stress response and central metabolism processes, also play roles in stress sensing, bile-induced damage prevention and energy efficiency. Moreover, bile salts appear to enhance proteolysis and amino acid uptake (especially aromatic amino acids) by LI01, which may support the liver protection properties of this strain. Altogether, this study establishes a model of global response mechanism to bile stress in L. salivarius LI01. L. salivarius strain LI01 exhibits not only antibacterial and antifungal properties but also exerts a good health-promoting effect in acute liver failure. As a potential probiotic strain, the bile-tolerance trait of strain LI01 is important, though this has not yet been explored. In this study, an analysis based on DGE and iTRAQ was performed to investigate the gene expression in strain LI01 under bile stress at the mRNA and protein levels, respectively. To our knowledge, this work also represents the first combined transcriptomic and proteomic analysis of the bile stress response mechanism in L. salivarius. Copyright © 2016. Published by Elsevier B.V.

  16. An integrated metagenome and -proteome analysis of the microbial community residing in a biogas production plant.

    Science.gov (United States)

    Ortseifen, Vera; Stolze, Yvonne; Maus, Irena; Sczyrba, Alexander; Bremges, Andreas; Albaum, Stefan P; Jaenicke, Sebastian; Fracowiak, Jochen; Pühler, Alfred; Schlüter, Andreas

    2016-08-10

    To study the metaproteome of a biogas-producing microbial community, fermentation samples were taken from an agricultural biogas plant for microbial cell and protein extraction and corresponding metagenome analyses. Based on metagenome sequence data, taxonomic community profiling was performed to elucidate the composition of bacterial and archaeal sub-communities. The community's cytosolic metaproteome was represented in a 2D-PAGE approach. Metaproteome databases for protein identification were compiled based on the assembled metagenome sequence dataset for the biogas plant analyzed and non-corresponding biogas metagenomes. Protein identification results revealed that the corresponding biogas protein database facilitated the highest identification rate followed by other biogas-specific databases, whereas common public databases yielded insufficient identification rates. Proteins of the biogas microbiome identified as highly abundant were assigned to the pathways involved in methanogenesis, transport and carbon metabolism. Moreover, the integrated metagenome/-proteome approach enabled the examination of genetic-context information for genes encoding identified proteins by studying neighboring genes on the corresponding contig. Exemplarily, this approach led to the identification of a Methanoculleus sp. contig encoding 16 methanogenesis-related gene products, three of which were also detected as abundant proteins within the community's metaproteome. Thus, metagenome contigs provide additional information on the genetic environment of identified abundant proteins. Copyright © 2016 Elsevier B.V. All rights reserved.

  17. Integrated analysis of transcriptome and proteome changes related to the Ogura cytoplasmic male sterility in cabbage.

    Directory of Open Access Journals (Sweden)

    Miaomiao Xing

    Full Text Available Cabbage (Brassica oleracea L. var. capitata, an important vegetable crop in the Brassicaceae family, is economically important worldwide. In the process of hybrid seed production, Ogura cytoplasmic male sterility (OguCMS, controlled by the mitochondrial gene orf138, has been extensively used for cabbage hybrid production with complete and stable male sterility. To identify the critical genes and pathways involved in the sterility and to better understand the underlying molecular mechanisms, the anther of OguCMS line R2P2CMS and the fertile line R2P2 were used for RNA-seq and iTRAQ (Isobaric Tags for Relative and Absolute Quantitation proteome analysis. RNA-seq analysis generated 13,037,109 to 13,066,594 SE50-clean reads, from the sterile and fertile lines, which were assembled into 36,890 unigenes. Among them, 1,323 differentially expressed genes (DEGs were identified, consisting of 307 up- and 1016 down-regulated genes. For ITRAQ analysis, a total of 7,147 unique proteins were identified, and 833 were differentially expressed including 538 up- and 295 down-regulated proteins. These were mainly annotated to the ribosome, spliceosome and mRNA surveillance pathways. Combined transcriptomic and proteomic analyses identified 22 and 70 genes with the same and opposite expression profiles, respectively. Using KEGG analysis of DEGs, gibberellin mediated signaling pathways regulating tapetum programmed cell death and four different pathways involved in sporopollenin synthesis were identified. Secretion and translocation of the sporopollenin precursors were identified, and the key genes participating in these pathways were all significantly down-regulated in R2P2CMS. Light and transmission electron (TE microscopy revealed fat abnormal tapetum rather than vacuolization and degradation at the tetrad and microspore stages of the OguCMS line. This resulted in the failed deposition of sporopollenin on the pollen resulting in sterility. This study provides a

  18. Integration of two-dimensional LC-MS with multivariate statistics for comparative analysis of proteomic samples

    NARCIS (Netherlands)

    Gaspari, M.; Verhoeckx, K.C.M.; Verheij, E.R.; Greef, J. van der

    2006-01-01

    LC-MS-based proteomics requires methods with high peak capacity and a high degree of automation, integrated with data-handling tools able to cope with the massive data produced and able to quantitatively compare them. This paper describes an off-line two-dimensional (2D) LC-MS method and its

  19. A graphical user interface for RAId, a knowledge integrated proteomics analysis suite with accurate statistics

    OpenAIRE

    Joyce, Brendan; Lee, Danny; Rubio, Alex; Ogurtsov, Aleksey; Alves, Gelio; Yu, Yi-Kuo

    2018-01-01

    Abstract Objective RAId is a software package that has been actively developed for the past 10 years for computationally and visually analyzing MS/MS data. Founded on rigorous statistical methods, RAId’s core program computes accurate E-values for peptides and proteins identified during database searches. Making this robust tool readily accessible for the proteomics community by developing a graphical user interface (GUI) is our main goa...

  20. Complex and extensive post-transcriptional regulation revealed by integrative proteomic and transcriptomic analysis of metabolite stress response in Clostridium acetobutylicum.

    Science.gov (United States)

    Venkataramanan, Keerthi P; Min, Lie; Hou, Shuyu; Jones, Shawn W; Ralston, Matthew T; Lee, Kelvin H; Papoutsakis, E Terry

    2015-01-01

    Clostridium acetobutylicum is a model organism for both clostridial biology and solvent production. The organism is exposed to its own toxic metabolites butyrate and butanol, which trigger an adaptive stress response. Integrative analysis of proteomic and RNAseq data may provide novel insights into post-transcriptional regulation. The identified iTRAQ-based quantitative stress proteome is made up of 616 proteins with a 15 % genome coverage. The differentially expressed proteome correlated poorly with the corresponding differential RNAseq transcriptome. Up to 31 % of the differentially expressed proteins under stress displayed patterns opposite to those of the transcriptome, thus suggesting significant post-transcriptional regulation. The differential proteome of the translation machinery suggests that cells employ a different subset of ribosomal proteins under stress. Several highly upregulated proteins but with low mRNA levels possessed mRNAs with long 5'UTRs and strong RBS scores, thus supporting the argument that regulatory elements on the long 5'UTRs control their translation. For example, the oxidative stress response rubrerythrin was upregulated only at the protein level up to 40-fold without significant mRNA changes. We also identified many leaderless transcripts, several displaying different transcriptional start sites, thus suggesting mRNA-trimming mechanisms under stress. Downregulation of Rho and partner proteins pointed to changes in transcriptional elongation and termination under stress. The integrative proteomic-transcriptomic analysis demonstrated complex expression patterns of a large fraction of the proteome. Such patterns could not have been detected with one or the other omic analyses. Our analysis proposes the involvement of specific molecular mechanisms of post-transcriptional regulation to explain the observed complex stress response.

  1. High throughput and accurate serum proteome profiling by integrated sample preparation technology and single-run data independent mass spectrometry analysis.

    Science.gov (United States)

    Lin, Lin; Zheng, Jiaxin; Yu, Quan; Chen, Wendong; Xing, Jinchun; Chen, Chenxi; Tian, Ruijun

    2018-03-01

    Mass spectrometry (MS)-based serum proteome analysis is extremely challenging due to its high complexity and dynamic range of protein abundances. Developing high throughput and accurate serum proteomic profiling approach capable of analyzing large cohorts is urgently needed for biomarker discovery. Herein, we report a streamlined workflow for fast and accurate proteomic profiling from 1μL of blood serum. The workflow combined an integrated technique for highly sensitive and reproducible sample preparation and a new data-independent acquisition (DIA)-based MS method. Comparing with standard data dependent acquisition (DDA) approach, the optimized DIA method doubled the number of detected peptides and proteins with better reproducibility. Without protein immunodepletion and prefractionation, the single-run DIA analysis enables quantitative profiling of over 300 proteins with 50min gradient time. The quantified proteins span more than five orders of magnitude of abundance range and contain over 50 FDA-approved disease markers. The workflow allowed us to analyze 20 serum samples per day, with about 358 protein groups per sample being identified. A proof-of-concept study on renal cell carcinoma (RCC) serum samples confirmed the feasibility of the workflow for large scale serum proteomic profiling and disease-related biomarker discovery. Blood serum or plasma is the predominant specimen for clinical proteomic studies while the analysis is extremely challenging for its high complexity. Many efforts had been made in the past for serum proteomics for maximizing protein identifications, whereas few have been concerned with throughput and reproducibility. Here, we establish a rapid, robust and high reproducible DIA-based workflow for streamlined serum proteomic profiling from 1μL serum. The workflow doesn't need protein depletion and pre-fractionation, while still being able to detect disease-relevant proteins accurately. The workflow is promising in clinical application

  2. PIQMIe: A web server for semi-quantitative proteomics data management and analysis

    NARCIS (Netherlands)

    A. Kuzniar (Arnold); R. Kanaar (Roland)

    2014-01-01

    textabstractWe present the Proteomics Identifications and Quantitations Data Management and Integration Service or PIQMIe that aids in reliable and scalable data management, analysis and visualization of semi-quantitative mass spectrometry based proteomics experiments. PIQMIe readily integrates

  3. Integrating cell biology and proteomic approaches in plants.

    Science.gov (United States)

    Takáč, Tomáš; Šamajová, Olga; Šamaj, Jozef

    2017-10-03

    Significant improvements of protein extraction, separation, mass spectrometry and bioinformatics nurtured advancements of proteomics during the past years. The usefulness of proteomics in the investigation of biological problems can be enhanced by integration with other experimental methods from cell biology, genetics, biochemistry, pharmacology, molecular biology and other omics approaches including transcriptomics and metabolomics. This review aims to summarize current trends integrating cell biology and proteomics in plant science. Cell biology approaches are most frequently used in proteomic studies investigating subcellular and developmental proteomes, however, they were also employed in proteomic studies exploring abiotic and biotic stress responses, vesicular transport, cytoskeleton and protein posttranslational modifications. They are used either for detailed cellular or ultrastructural characterization of the object subjected to proteomic study, validation of proteomic results or to expand proteomic data. In this respect, a broad spectrum of methods is employed to support proteomic studies including ultrastructural electron microscopy studies, histochemical staining, immunochemical localization, in vivo imaging of fluorescently tagged proteins and visualization of protein-protein interactions. Thus, cell biological observations on fixed or living cell compartments, cells, tissues and organs are feasible, and in some cases fundamental for the validation and complementation of proteomic data. Validation of proteomic data by independent experimental methods requires development of new complementary approaches. Benefits of cell biology methods and techniques are not sufficiently highlighted in current proteomic studies. This encouraged us to review most popular cell biology methods used in proteomic studies and to evaluate their relevance and potential for proteomic data validation and enrichment of purely proteomic analyses. We also provide examples of

  4. Integrated Analysis of Genetic and Proteomic Data Identifies Biomarkers Associated with Adverse Events Following Smallpox Vaccination

    Science.gov (United States)

    Complex clinical outcomes, such as adverse reaction to vaccination, arise from the concerted interactions among the myriad components of a biological system. Therefore, comprehensive etiological models can be developed only through the integrated study of multiple types of experi...

  5. BioinformatiqTM - integrating data types for proteomic discovery

    International Nuclear Information System (INIS)

    Arthur, J.W.; Harrison, M.; Manoharan, A.; Traini, M.; Shaw, E.; Wilkins, M.

    2001-01-01

    Proteomics (Wilkins et al. 1997) involves the large-scale analysis of expressed proteins. At each stage of the discovery process the researcher accumulates large volumes of data. These include: clinical or biological data about the sample being studied; details of sample purification and separation; images of 2D gels and associated information; MALDI mass spectra; MS/MS and PSD spectra; as well as meta-data relating to the projects undertaken and experiments performed. All this must be combined with existing databases of protein and EST sequences, post-translational modifications, and protein glycosylation, then processed with sophisticated bioinformatics tools in order to extract meaningful answers to questions of biological, clinical, and agricultural significance. BioinformatlQ TM is a web-based application for the storage, management, and automated bioinformatic analysis of proteomic information. This poster will demonstrate the integration of these disparate data sources in proteomics

  6. A system for integrative and post-planned analysis of 2-DE/MS centered proteomics data

    Directory of Open Access Journals (Sweden)

    Wang Juhui

    2005-12-01

    Full Text Available Proteomic analysis is intrinsically an iterative, incremental process. Information is usually acquired gradually by researchers, and in different projects. At the same time, there are relatively few examples of biological data management systems which take into account this reality, most of them usually treat the experiment generated data as static and unchangeable: data are never reconsidered, or seldom, whereas technology becomes more powerful or that other researchers have brought information on data correction. And yet, post-planned analysis [21] which involves multiple iterations and subsequent re-investigations of previously prepared data might bring tremendous benefits.

  7. Proteomic analysis of the dysferlin protein complex unveils its importance for sarcolemmal maintenance and integrity.

    Directory of Open Access Journals (Sweden)

    Antoine de Morrée

    Full Text Available Dysferlin is critical for repair of muscle membranes after damage. Mutations in dysferlin lead to a progressive muscular dystrophy. Recent studies suggest additional roles for dysferlin. We set out to study dysferlin's protein-protein interactions to obtain comprehensive knowledge of dysferlin functionalities in a myogenic context. We developed a robust and reproducible method to isolate dysferlin protein complexes from cells and tissue. We analyzed the composition of these complexes in cultured myoblasts, myotubes and skeletal muscle tissue by mass spectrometry and subsequently inferred potential protein functions through bioinformatics analyses. Our data confirm previously reported interactions and support a function for dysferlin as a vesicle trafficking protein. In addition novel potential functionalities were uncovered, including phagocytosis and focal adhesion. Our data reveal that the dysferlin protein complex has a dynamic composition as a function of myogenic differentiation. We provide additional experimental evidence and show dysferlin localization to, and interaction with the focal adhesion protein vinculin at the sarcolemma. Finally, our studies reveal evidence for cross-talk between dysferlin and its protein family member myoferlin. Together our analyses show that dysferlin is not only a membrane repair protein but also important for muscle membrane maintenance and integrity.

  8. Proteomic Analysis of Chinese Hamster Ovary Cells

    DEFF Research Database (Denmark)

    Baycin-Hizal, Deniz; Tabb, David L.; Chaerkady, Raghothama

    2012-01-01

    To complement the recent genomic sequencing of Chinese hamster ovary (CHO) cells, proteomic analysis was performed on CHO cells including the cellular proteome, secretome, and glycoproteome using tandem mass spectrometry (MS/MS) of multiple fractions obtained from gel electrophoresis, multidimens......To complement the recent genomic sequencing of Chinese hamster ovary (CHO) cells, proteomic analysis was performed on CHO cells including the cellular proteome, secretome, and glycoproteome using tandem mass spectrometry (MS/MS) of multiple fractions obtained from gel electrophoresis...

  9. System-Level and Granger Network Analysis of Integrated Proteomic and Metabolomic Dynamics Identifies Key Points of Grape Berry Development at the Interface of Primary and Secondary Metabolism

    Directory of Open Access Journals (Sweden)

    Lei Wang

    2017-06-01

    Full Text Available Grapevine is a fruit crop with worldwide economic importance. The grape berry undergoes complex biochemical changes from fruit set until ripening. This ripening process and production processes define the wine quality. Thus, a thorough understanding of berry ripening is crucial for the prediction of wine quality. For a systemic analysis of grape berry development we applied mass spectrometry based platforms to analyse the metabolome and proteome of Early Campbell at 12 stages covering major developmental phases. Primary metabolites involved in central carbon metabolism, such as sugars, organic acids and amino acids together with various bioactive secondary metabolites like flavonols, flavan-3-ols and anthocyanins were annotated and quantified. At the same time, the proteomic analysis revealed the protein dynamics of the developing grape berries. Multivariate statistical analysis of the integrated metabolomic and proteomic dataset revealed the growth trajectory and corresponding metabolites and proteins contributing most to the specific developmental process. K-means clustering analysis revealed 12 highly specific clusters of co-regulated metabolites and proteins. Granger causality network analysis allowed for the identification of time-shift correlations between metabolite-metabolite, protein- protein and protein-metabolite pairs which is especially interesting for the understanding of developmental processes. The integration of metabolite and protein dynamics with their corresponding biochemical pathways revealed an energy-linked metabolism before veraison with high abundances of amino acids and accumulation of organic acids, followed by protein and secondary metabolite synthesis. Anthocyanins were strongly accumulated after veraison whereas other flavonoids were in higher abundance at early developmental stages and decreased during the grape berry developmental processes. A comparison of the anthocyanin profile of Early Campbell to other

  10. System-Level and Granger Network Analysis of Integrated Proteomic and Metabolomic Dynamics Identifies Key Points of Grape Berry Development at the Interface of Primary and Secondary Metabolism.

    Science.gov (United States)

    Wang, Lei; Sun, Xiaoliang; Weiszmann, Jakob; Weckwerth, Wolfram

    2017-01-01

    Grapevine is a fruit crop with worldwide economic importance. The grape berry undergoes complex biochemical changes from fruit set until ripening. This ripening process and production processes define the wine quality. Thus, a thorough understanding of berry ripening is crucial for the prediction of wine quality. For a systemic analysis of grape berry development we applied mass spectrometry based platforms to analyse the metabolome and proteome of Early Campbell at 12 stages covering major developmental phases. Primary metabolites involved in central carbon metabolism, such as sugars, organic acids and amino acids together with various bioactive secondary metabolites like flavonols, flavan-3-ols and anthocyanins were annotated and quantified. At the same time, the proteomic analysis revealed the protein dynamics of the developing grape berries. Multivariate statistical analysis of the integrated metabolomic and proteomic dataset revealed the growth trajectory and corresponding metabolites and proteins contributing most to the specific developmental process. K-means clustering analysis revealed 12 highly specific clusters of co-regulated metabolites and proteins. Granger causality network analysis allowed for the identification of time-shift correlations between metabolite-metabolite, protein- protein and protein-metabolite pairs which is especially interesting for the understanding of developmental processes. The integration of metabolite and protein dynamics with their corresponding biochemical pathways revealed an energy-linked metabolism before veraison with high abundances of amino acids and accumulation of organic acids, followed by protein and secondary metabolite synthesis. Anthocyanins were strongly accumulated after veraison whereas other flavonoids were in higher abundance at early developmental stages and decreased during the grape berry developmental processes. A comparison of the anthocyanin profile of Early Campbell to other cultivars revealed

  11. Integrative Analysis Using Proteome and Transcriptome Data From Yeast to Unravel Regulatory Patterns at Post-Transcriptional Level

    DEFF Research Database (Denmark)

    Olivares Hernandez, Roberto; Usaite, Renata; Nielsen, Jens

    2010-01-01

    In this stud) we combined proteome and transcriptome data from six different published dataset to identify patterns that can provide new insight into the reasons for these deviations By using a categorization method and integrating genome-scale information we found that the relation between protein and mRNA...... is related to the gene function We could further identify that for genes belonging to amino acid biosynthetic pathways there is no translational regulation, meaning that there is generally a good correlation between mRNA and protein levels We also found that there is generally translational control for large...... proteins and there also evidence for a role of conserved motifs m the 3' untranslated regions in the mRNA-protein correlation, probably by controlling the level of mRNA Biotechnol Bioeng 2010,107 865-875...

  12. Analysis of Peanut Leaf Proteome

    DEFF Research Database (Denmark)

    Ramesh, R.; Suravajhala, Prashanth; Pechan, T.

    2010-01-01

    Peanut (Arachis hypogaea) is one of the most important sources of plant protein. Current selection of genotypes requires molecular characterization of available populations. Peanut genome database has several EST cDNAs which can be used to analyze gene expression. Analysis of proteins is a direct...... approach to define function of their associated genes. Proteome analysis linked to genome sequence information is critical for functional genomics. However, the available protein expression data is extremely inadequate. Proteome analysis of peanut leaf was conducted using two-dimensional gel...... electrophoresis in combination with sequence identification using MALDI/TOF to determine their identity and function related to growth, development and responses to stresses. Peanut leaf proteins were resolved into 300 polypeptides with pI values between 3.5 and 8.0 and relative molecular masses from 12 to 100 k...

  13. Proteome analysis of Aspergillus ochraceus.

    Science.gov (United States)

    Rizwan, Muhammad; Miller, Ingrid; Tasneem, Fareeha; Böhm, Josef; Gemeiner, Manfred; Razzazi-Fazeli, Ebrahim

    2010-08-01

    Genome sequencing for many important fungi has begun during recent years; however, there is still some deficiency in proteome profiling of aspergilli. To obtain a comprehensive overview of proteins and their expression, a proteomic approach based on 2D gel electrophoresis and MALDI-TOF/TOF mass spectrometry was used to investigate A. ochraceus. The cell walls of fungi are exceptionally resistant to destruction, therefore two lysis protocols were tested: (1) lysis via manual grinding using liquid nitrogen, and (2) mechanical lysis via rapid agitation with glass beads using MagNalyser. Mechanical grinding with mortar and pestle using liquid nitrogen was found to be a more efficient extraction method for our purpose, resulting in extracts with higher protein content and a clear band pattern in SDS-PAGE. Two-dimensional electrophoresis gave a complex spot pattern comprising proteins of a broad range of isoelectric points and molecular masses. The most abundant spots were subjected to mass spectrometric analysis. We could identify 31 spots representing 26 proteins, most of them involved in metabolic processes and response to stress. Seventeen spots were identified by de novo sequencing due to a lack of DNA and protein database sequences of A. ochraceus. The proteins identified in our study have been reported for the first time in A. ochraceus and this represents the first proteomic approach with identification of major proteins, when the fungus was grown under submerged culture.

  14. Analysis of mass spectrometry data in proteomics

    DEFF Research Database (Denmark)

    Matthiesen, Rune; Jensen, Ole N

    2008-01-01

    The systematic study of proteins and protein networks, that is, proteomics, calls for qualitative and quantitative analysis of proteins and peptides. Mass spectrometry (MS) is a key analytical technology in current proteomics and modern mass spectrometers generate large amounts of high-quality data...... that in turn allow protein identification, annotation of secondary modifications, and determination of the absolute or relative abundance of individual proteins. Advances in mass spectrometry-driven proteomics rely on robust bioinformatics tools that enable large-scale data analysis. This chapter describes...... some of the basic concepts and current approaches to the analysis of MS and MS/MS data in proteomics....

  15. Integration analysis of quantitative proteomics and transcriptomics data identifies potential targets of frizzled-8 protein-related antiproliferative factor in vivo.

    Science.gov (United States)

    Yang, Wei; Kim, Yongsoo; Kim, Taek-Kyun; Keay, Susan K; Kim, Kwang Pyo; Steen, Hanno; Freeman, Michael R; Hwang, Daehee; Kim, Jayoung

    2012-12-01

    What's known on the subject? and What does the study add? Interstitial cystitis (IC) is a prevalent and debilitating pelvic disorder generally accompanied by chronic pain combined with chronic urinating problems. Over one million Americans are affected, especially middle-aged women. However, its aetiology or mechanism remains unclear. No efficient drug has been provided to patients. Several urinary biomarker candidates have been identified for IC; among the most promising is antiproliferative factor (APF), whose biological activity is detectable in urine specimens from >94% of patients with both ulcerative and non-ulcerative IC. The present study identified several important mediators of the effect of APF on bladder cell physiology, suggesting several candidate drug targets against IC. In an attempt to identify potential proteins and genes regulated by APF in vivo, and to possibly expand the APF-regulated network identified by stable isotope labelling by amino acids in cell culture (SILAC), we performed an integration analysis of our own SILAC data and the microarray data of Gamper et al. (2009) BMC Genomics 10: 199. Notably, two of the proteins (i.e. MAPKSP1 and GSPT1) that are down-regulated by APF are involved in the activation of mTORC1, suggesting that the mammalian target of rapamycin (mTOR) pathway is potentially a critical pathway regulated by APF in vivo. Several components of the mTOR pathway are currently being studied as potential therapeutic targets in other diseases. Our analysis suggests that this pathway might also be relevant in the design of diagnostic tools and medications targeting IC. • To enhance our understanding of the interstitial cystitis urine biomarker antiproliferative factor (APF), as well as interstitial cystitis biology more generally at the systems level, we reanalyzed recently published large-scale quantitative proteomics and in vivo transcriptomics data sets using an integration analysis tool that we have developed. • To

  16. Proteomic analysis of the Theileria annulata schizont

    Science.gov (United States)

    Witschi, M.; Xia, D.; Sanderson, S.; Baumgartner, M.; Wastling, J.M.; Dobbelaere, D.A.E.

    2013-01-01

    The apicomplexan parasite, Theileria annulata, is the causative agent of tropical theileriosis, a devastating lymphoproliferative disease of cattle. The schizont stage transforms bovine leukocytes and provides an intriguing model to study host/pathogen interactions. The genome of T. annulata has been sequenced and transcriptomic data are rapidly accumulating. In contrast, little is known about the proteome of the schizont, the pathogenic, transforming life cycle stage of the parasite. Using one-dimensional (1-D) gel LC-MS/MS, a proteomic analysis of purified T. annulata schizonts was carried out. In whole parasite lysates, 645 proteins were identified. Proteins with transmembrane domains (TMDs) were under-represented and no proteins with more than four TMDs could be detected. To tackle this problem, Triton X-114 treatment was applied, which facilitates the extraction of membrane proteins, followed by 1-D gel LC-MS/MS. This resulted in the identification of an additional 153 proteins. Half of those had one or more TMD and 30 proteins with more than four TMDs were identified. This demonstrates that Triton X-114 treatment can provide a valuable additional tool for the identification of new membrane proteins in proteomic studies. With two exceptions, all proteins involved in glycolysis and the citric acid cycle were identified. For at least 29% of identified proteins, the corresponding transcripts were not present in the existing expressed sequence tag databases. The proteomics data were integrated into the publicly accessible database resource at EuPathDB (www.eupathdb.org) so that mass spectrometry-based protein expression evidence for T. annulata can be queried alongside transcriptional and other genomics data available for these parasites. PMID:23178997

  17. A community proposal to integrate proteomics activities in ELIXIR

    NARCIS (Netherlands)

    Vizcaíno, Juan Antonio; Walzer, Mathias; Jiménez, Rafael C; Bittremieux, Wout; Bouyssié, David; Carapito, Christine; Corrales, Fernando; Ferro, Myriam; Heck, Albert J R; Horvatovich, Peter; Hubalek, Martin; Lane, Lydie; Laukens, Kris; Levander, Fredrik; Lisacek, Frederique; Novak, Petr; Palmblad, Magnus; Piovesan, Damiano; Pühler, Alfred; Schwämmle, Veit; Valkenborg, Dirk; van Rijswijk, Merlijn; Vondrasek, Jiri; Eisenacher, Martin; Martens, Lennart; Kohlbacher, Oliver

    2017-01-01

    Computational approaches have been major drivers behind the progress of proteomics in recent years. The aim of this white paper is to provide a framework for integrating computational proteomics into ELIXIR in the near future, and thus to broaden the portfolio of omics technologies supported by this

  18. Integrative Temporo-Spatial, Mineralogic, Spectroscopic, and Proteomic Analysis of Postnatal Enamel Development in Teeth with Limited Growth

    Directory of Open Access Journals (Sweden)

    Mirali Pandya

    2017-10-01

    Full Text Available Tooth amelogenesis is a complex process beginning with enamel organ cell differentiation and enamel matrix secretion, transitioning through changes in ameloblast polarity, cytoskeletal, and matrix organization, that affects crucial biomineralization events such as mineral nucleation, enamel crystal growth, and enamel prism organization. Here we have harvested the enamel organ including the pliable enamel matrix of postnatal first mandibular mouse molars during the first 8 days of tooth enamel development to conduct a step-wise cross-sectional analysis of the changes in the mineral and protein phase. Mineral phase diffraction pattern analysis using single-crystal, powder sample X-ray diffraction analysis indicated conversion of calcium phosphate precursors to partially fluoride substituted hydroxyapatite from postnatal day 4 (4 dpn onwards. Attenuated total reflectance spectra (ATR revealed a substantial elevation in phosphate and carbonate incorporation as well as structural reconfiguration between postnatal days 6 and 8. Nanoscale liquid chromatography coupled with tandem mass spectrometry (nanoLC-MS/MS demonstrated highest protein counts for ECM/cell surface proteins, stress/heat shock proteins, and alkaline phosphatase on postnatal day 2, high counts for ameloblast cytoskeletal proteins such as tubulin β5, tropomyosin, β-actin, and vimentin on postnatal day 4, and elevated levels of cofilin-1, calmodulin, and peptidyl-prolyl cis-trans isomerase on day 6. Western blot analysis of hydrophobic enamel proteins illustrated continuously increasing amelogenin levels from 1 dpn until 8 dpn, while enamelin peaked on days 1 and 2 dpn, and ameloblastin on days 1–5 dpn. In summary, these data document the substantial changes in the enamel matrix protein and mineral phase that take place during postnatal mouse molar amelogenesis from a systems biological perspective, including (i relatively high levels of matrix protein expression during the early

  19. The Role of Lipid Droplets in Mortierella alpina Aging Revealed by Integrative Subcellular and Whole-Cell Proteome Analysis.

    Science.gov (United States)

    Yu, Yadong; Li, Tao; Wu, Na; Jiang, Ling; Ji, Xiaojun; Huang, He

    2017-03-07

    Lipid droplets (LDs) participate in many cellular processes in oleaginous microorganisms. However, the exact function of LDs in the Mortierella alpina aging process remains elusive. Herein, subcellular proteomics was employed to unveil the composition and dynamics of the LD proteome in the aging M. alpina for the first time. More than 400 proteins were detected in LDs and 62 of them changed expression significantly during aging. By combining the LD proteomic data with whole-cell data, we found that the carbohydrate metabolism and de novo lipid biosynthesis were all inhibited during aging of M. alpina mycelia. The up-regulation of fructose metabolism-related enzymes in LDs might imply that LDs facilitated the fructose metabolism, which in turn might cause pyruvate to accumulate and enter malate-pyruvate cycle, and ultimately, provide additional NADPH for the synthesis of arachidonic acid (ARA). Lysophospholipase and lecithinase were up-regulated in LDs during the aging process, suggesting that the phospholipids and lecithin were starting to be hydrolyzed, in order to release fatty acids for the cells. The impairment of the anti-oxidant system might lead to the accumulation of ROS and consequently cause the up-regulation of autophagy-related proteins in LDs, which further induces the M. alpina mycelia to activate the autophagy process.

  20. Network-based analysis of proteomic profiles

    KAUST Repository

    Wong, Limsoon

    2016-01-26

    Mass spectrometry (MS)-based proteomics is a widely used and powerful tool for profiling systems-wide protein expression changes. It can be applied for various purposes, e.g. biomarker discovery in diseases and study of drug responses. Although RNA-based high-throughput methods have been useful in providing glimpses into the underlying molecular processes, the evidences they provide are indirect. Furthermore, RNA and corresponding protein levels have been known to have poor correlation. On the other hand, MS-based proteomics tend to have consistency issues (poor reproducibility and inter-sample agreement) and coverage issues (inability to detect the entire proteome) that need to be urgently addressed. In this talk, I will discuss how these issues can be addressed by proteomic profile analysis techniques that use biological networks (especially protein complexes) as the biological context. In particular, I will describe several techniques that we have been developing for network-based analysis of proteomics profile. And I will present evidence that these techniques are useful in identifying proteomics-profile analysis results that are more consistent, more reproducible, and more biologically coherent, and that these techniques allow expansion of the detected proteome to uncover and/or discover novel proteins.

  1. Proteomic analysis of human oral verrucous carcinoma

    African Journals Online (AJOL)

    Jane

    2011-10-05

    Oct 5, 2011 ... This study is about proteomic analysis of oral verrucous carcinoma (OVC). The total proteins ..... receptor protein (recoverin) through autoimmunity ..... chromosome 8q21.1 and overexpressed in human prostate cancer. Cancer ...

  2. Babelomics: an integrative platform for the analysis of transcriptomics, proteomics and genomic data with advanced functional profiling

    Science.gov (United States)

    Medina, Ignacio; Carbonell, José; Pulido, Luis; Madeira, Sara C.; Goetz, Stefan; Conesa, Ana; Tárraga, Joaquín; Pascual-Montano, Alberto; Nogales-Cadenas, Ruben; Santoyo, Javier; García, Francisco; Marbà, Martina; Montaner, David; Dopazo, Joaquín

    2010-01-01

    Babelomics is a response to the growing necessity of integrating and analyzing different types of genomic data in an environment that allows an easy functional interpretation of the results. Babelomics includes a complete suite of methods for the analysis of gene expression data that include normalization (covering most commercial platforms), pre-processing, differential gene expression (case-controls, multiclass, survival or continuous values), predictors, clustering; large-scale genotyping assays (case controls and TDTs, and allows population stratification analysis and correction). All these genomic data analysis facilities are integrated and connected to multiple options for the functional interpretation of the experiments. Different methods of functional enrichment or gene set enrichment can be used to understand the functional basis of the experiment analyzed. Many sources of biological information, which include functional (GO, KEGG, Biocarta, Reactome, etc.), regulatory (Transfac, Jaspar, ORegAnno, miRNAs, etc.), text-mining or protein–protein interaction modules can be used for this purpose. Finally a tool for the de novo functional annotation of sequences has been included in the system. This provides support for the functional analysis of non-model species. Mirrors of Babelomics or command line execution of their individual components are now possible. Babelomics is available at http://www.babelomics.org. PMID:20478823

  3. A community proposal to integrate proteomics activities in ELIXIR.

    Science.gov (United States)

    Vizcaíno, Juan Antonio; Walzer, Mathias; Jiménez, Rafael C; Bittremieux, Wout; Bouyssié, David; Carapito, Christine; Corrales, Fernando; Ferro, Myriam; Heck, Albert J R; Horvatovich, Peter; Hubalek, Martin; Lane, Lydie; Laukens, Kris; Levander, Fredrik; Lisacek, Frederique; Novak, Petr; Palmblad, Magnus; Piovesan, Damiano; Pühler, Alfred; Schwämmle, Veit; Valkenborg, Dirk; van Rijswijk, Merlijn; Vondrasek, Jiri; Eisenacher, Martin; Martens, Lennart; Kohlbacher, Oliver

    2017-01-01

    Computational approaches have been major drivers behind the progress of proteomics in recent years. The aim of this white paper is to provide a framework for integrating computational proteomics into ELIXIR in the near future, and thus to broaden the portfolio of omics technologies supported by this European distributed infrastructure. This white paper is the direct result of a strategy meeting on 'The Future of Proteomics in ELIXIR' that took place in March 2017 in Tübingen (Germany), and involved representatives of eleven ELIXIR nodes. These discussions led to a list of priority areas in computational proteomics that would complement existing activities and close gaps in the portfolio of tools and services offered by ELIXIR so far. We provide some suggestions on how these activities could be integrated into ELIXIR's existing platforms, and how it could lead to a new ELIXIR use case in proteomics. We also highlight connections to the related field of metabolomics, where similar activities are ongoing. This white paper could thus serve as a starting point for the integration of computational proteomics into ELIXIR. Over the next few months we will be working closely with all stakeholders involved, and in particular with other representatives of the proteomics community, to further refine this paper.

  4. Comprehensive proteomic analysis of human pancreatic juice

    DEFF Research Database (Denmark)

    Grønborg, Mads; Bunkenborg, Jakob; Kristiansen, Troels Zakarias

    2004-01-01

    Proteomic technologies provide an excellent means for analysis of body fluids for cataloging protein constituents and identifying biomarkers for early detection of cancers. The biomarkers currently available for pancreatic cancer, such as CA19-9, lack adequate sensitivity and specificity...... contributing to late diagnosis of this deadly disease. In this study, we carried out a comprehensive characterization of the "pancreatic juice proteome" in patients with pancreatic adenocarcinoma. Pancreatic juice was first fractionated by 1-dimensional gel electrophoresis and subsequently analyzed by liquid...... in this study could be directly assessed for their potential as biomarkers for pancreatic cancer by quantitative proteomics methods or immunoassays....

  5. Identification Of Protein Vaccine Candidates Using Comprehensive Proteomic Analysis Strategies

    Science.gov (United States)

    2007-12-01

    that fascinating fungus known as Coccidioides. I also want to thank the UA Mass Spectrometry Facility and the UA Proteomics Consortium, especially...W. & N. N. Kav. 2006. The proteome of the phytopathogenic fungus Sclerotinia sclerotiorum. Proteomics 6: 5995-6007. 127. de Godoy, L. M., J. V...IDENTIFICATION OF PROTEIN VACCINE CANDIDATES USING COMPREHENSIVE PROTEOMIC ANALYSIS STRATEGIES by James G. Rohrbough

  6. PACOM: A Versatile Tool for Integrating, Filtering, Visualizing, and Comparing Multiple Large Mass Spectrometry Proteomics Data Sets.

    Science.gov (United States)

    Martínez-Bartolomé, Salvador; Medina-Aunon, J Alberto; López-García, Miguel Ángel; González-Tejedo, Carmen; Prieto, Gorka; Navajas, Rosana; Salazar-Donate, Emilio; Fernández-Costa, Carolina; Yates, John R; Albar, Juan Pablo

    2018-04-06

    Mass-spectrometry-based proteomics has evolved into a high-throughput technology in which numerous large-scale data sets are generated from diverse analytical platforms. Furthermore, several scientific journals and funding agencies have emphasized the storage of proteomics data in public repositories to facilitate its evaluation, inspection, and reanalysis. (1) As a consequence, public proteomics data repositories are growing rapidly. However, tools are needed to integrate multiple proteomics data sets to compare different experimental features or to perform quality control analysis. Here, we present a new Java stand-alone tool, Proteomics Assay COMparator (PACOM), that is able to import, combine, and simultaneously compare numerous proteomics experiments to check the integrity of the proteomic data as well as verify data quality. With PACOM, the user can detect source of errors that may have been introduced in any step of a proteomics workflow and that influence the final results. Data sets can be easily compared and integrated, and data quality and reproducibility can be visually assessed through a rich set of graphical representations of proteomics data features as well as a wide variety of data filters. Its flexibility and easy-to-use interface make PACOM a unique tool for daily use in a proteomics laboratory. PACOM is available at https://github.com/smdb21/pacom .

  7. A community proposal to integrate proteomics activities in ELIXIR

    DEFF Research Database (Denmark)

    Vizcaíno, Juan Antonio; Walzer, Mathias; Jiménez, Rafael C.

    2017-01-01

    in computational proteomics that would complement existing activities and close gaps in the portfolio of tools and services offered by ELIXIR so far. We provide some suggestions on how these activities could be integrated into ELIXIR's existing platforms, and how it could lead to a new ELIXIR use case...... involved, and in particular with other representatives of the proteomics community, to further refine this paper....

  8. Integration of a central protein repository into a standard data processing application for mining proteomics data

    DEFF Research Database (Denmark)

    Fritzemeier, Kai; Kristensen, Jakob; Larsen, Martin Røssel

    efficient data mining and categorizing of large data sets. Methods All samples were analyzed on an Orbitrap mass Spectrometer coupled to a nano Easy LC. The proteomics repository database is built using the Sun Java technology and the Microsoft mySQL database technology for optimal performance. Proteome......Novel Aspect All major protein repositories integrated into a central domain for direct analyses and interpretation in a standard proteomics data analysis software. Introduction Modern proteomics must face the challenge of performing bioinformatics analysis and comparison of large datasets....... It is a time consuming and at times nearly impossible task to distinguish known proteins from novel proteins in these data sets without proper annotation and comparison with literature sources. Tools are needed that can handle the complexity of these data including: redundancy (same protein but different...

  9. Clinical proteomic analysis of scrub typhus infection.

    Science.gov (United States)

    Park, Edmond Changkyun; Lee, Sang-Yeop; Yun, Sung Ho; Choi, Chi-Won; Lee, Hayoung; Song, Hyun Seok; Jun, Sangmi; Kim, Gun-Hwa; Lee, Chang-Seop; Kim, Seung Il

    2018-01-01

    Scrub typhus is an acute and febrile infectious disease caused by the Gram-negative α-proteobacterium Orientia tsutsugamushi from the family Rickettsiaceae that is widely distributed in Northern, Southern and Eastern Asia. In the present study, we analysed the serum proteome of scrub typhus patients to investigate specific clinical protein patterns in an attempt to explain pathophysiology and discover potential biomarkers of infection. Serum samples were collected from three patients (before and after treatment with antibiotics) and three healthy subjects. One-dimensional sodium dodecyl sulphate-polyacrylamide gel electrophoresis followed by liquid chromatography-tandem mass spectrometry was performed to identify differentially abundant proteins using quantitative proteomic approaches. Bioinformatic analysis was then performed using Ingenuity Pathway Analysis. Proteomic analysis identified 236 serum proteins, of which 32 were differentially expressed in normal subjects, naive scrub typhus patients and patients treated with antibiotics. Comparative bioinformatic analysis of the identified proteins revealed up-regulation of proteins involved in immune responses, especially complement system, following infection with O. tsutsugamushi , and normal expression was largely rescued by antibiotic treatment. This is the first proteomic study of clinical serum samples from scrub typhus patients. Proteomic analysis identified changes in protein expression upon infection with O. tsutsugamushi and following antibiotic treatment. Our results provide valuable information for further investigation of scrub typhus therapy and diagnosis.

  10. Inspection, visualisation and analysis of quantitative proteomics data

    OpenAIRE

    Gatto, Laurent

    2016-01-01

    Material Quantitative Proteomics and Data Analysis Course. 4 - 5 April 2016, Queen Hotel, Chester, UK Table D - Inspection, visualisation and analysis of quantitative proteomics data, Laurent Gatto (University of Cambridge)

  11. System-Level and Granger Network Analysis of Integrated Proteomic and Metabolomic Dynamics Identifies Key Points of Grape Berry Development at the Interface of Primary and Secondary Metabolism

    OpenAIRE

    Wang, Lei; Sun, Xiaoliang; Weiszmann, Jakob; Weckwerth, Wolfram

    2017-01-01

    Grapevine is a fruit crop with worldwide economic importance. The grape berry undergoes complex biochemical changes from fruit set until ripening. This ripening process and production processes define the wine quality. Thus, a thorough understanding of berry ripening is crucial for the prediction of wine quality. For a systemic analysis of grape berry development we applied mass spectrometry based platforms to analyse the metabolome and proteome of Early Campbell at 12 stages covering major d...

  12. Global Proteome Analysis of the NCI-60 Cell Line Panel

    Directory of Open Access Journals (Sweden)

    Amin Moghaddas Gholami

    2013-08-01

    Full Text Available The NCI-60 cell line collection is a very widely used panel for the study of cellular mechanisms of cancer in general and in vitro drug action in particular. It is a model system for the tissue types and genetic diversity of human cancers and has been extensively molecularly characterized. Here, we present a quantitative proteome and kinome profile of the NCI-60 panel covering, in total, 10,350 proteins (including 375 protein kinases and including a core cancer proteome of 5,578 proteins that were consistently quantified across all tissue types. Bioinformatic analysis revealed strong cell line clusters according to tissue type and disclosed hundreds of differentially regulated proteins representing potential biomarkers for numerous tumor properties. Integration with public transcriptome data showed considerable similarity between mRNA and protein expression. Modeling of proteome and drug-response profiles for 108 FDA-approved drugs identified known and potential protein markers for drug sensitivity and resistance. To enable community access to this unique resource, we incorporated it into a public database for comparative and integrative analysis (http://wzw.tum.de/proteomics/nci60.

  13. Integrated transcriptomics and proteomics analysis of storage protein composition in developing barley grain to improve nutritional profile

    DEFF Research Database (Denmark)

    Kaczmarczyk, Agnieszka Ewa; Dionisio, Giuseppe; Renaut, Jenny

    2012-01-01

    The aim of the study was to understand the molecular and biochemical mechanisms underpinning the effect of nitrogen (N) on barley (Hordeum vulgare) storage protein production (hordeins) during grain filling. Using a combination of advanced biochemistry methods, we could comprehensively describe......-regimes caused significant differences in both quantity and quality of the storage proteins transcripts. Principal Component Analysis of the amino acid (AA) profiles also indicated dissimilarity in individual AA percentages, correlated to hordein content. The abundance values of proteins of interest confirmed...

  14. Proteomic analysis of human tooth pulp: proteomics of human tooth.

    Science.gov (United States)

    Eckhardt, Adam; Jágr, Michal; Pataridis, Statis; Mikšík, Ivan

    2014-12-01

    The unique pulp-dentin complex demonstrates strong regenerative potential, which enables it to respond to disease and traumatic injury. Identifying the proteins of the pulp-dentin complex is crucial to understanding the mechanisms of regeneration, tissue calcification, defense processes, and the reparation of dentin by dental pulp. The lack of knowledge of these proteins limits the development of more efficient therapies. The proteomic profile of human tooth pulp was investigated and compared with the proteome of human dentin and blood. The samples of tooth pulp were obtained from 5 sound permanent human third molars of 5 adults (n = 5). The extracted proteins were separated by 2-dimensional gel electrophoresis, analyzed by nano-liquid chromatography tandem mass spectrometry, and identified by correlating mass spectra to the proteomic databases. A total of 342 proteins were identified with high confidence, and 2 proteins were detected for the first time in an actual human sample. The identified tooth pulp proteins have a variety of functions: structural, catalytic, transporter, protease activity, immune response, and many others. In a comparison with dentin and blood plasma, 140 (pulp/dentin) shared proteins were identified, 37 of which were not observed in plasma. It can be suggested that they might participate in the unique pulp-dentin complex. This proteomic investigation of human tooth pulp, together with the previously published study of human dentin, is one of the most comprehensive proteome lists of human teeth to date. Copyright © 2014 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

  15. Isolation and proteomic analysis of Chlamydomonas centrioles.

    Science.gov (United States)

    Keller, Lani C; Marshall, Wallace F

    2008-01-01

    Centrioles are barrel-shaped cytoskeletal organelles composed of nine triplet microtubules blades arranged in a pinwheel-shaped array. Centrioles are required for recruitment of pericentriolar material (PCM) during centrosome formation, and they act as basal bodies, which are necessary for the outgrowth of cilia and flagella. Despite being described over a hundred years ago, centrioles are still among the most enigmatic organelles in all of cell biology. To gain molecular insights into the function and assembly of centrioles, we sought to determine the composition of the centriole proteome. Here, we describe a method that allows for the isolation of virtually "naked" centrioles, with little to no obscuring PCM, from the green alga, Chlamydomonas. Proteomic analysis of this material provided evidence that multiple human disease gene products encode protein components of the centriole, including genes involved in Meckel syndrome and Oral-Facial-Digital syndrome. Isolated centrioles can be used in combination with a wide variety of biochemical assays in addition to being utilized as a source for proteomic analysis.

  16. A Combined Metabolomic and Proteomic Analysis of Gestational Diabetes Mellitus

    Directory of Open Access Journals (Sweden)

    Joanna Hajduk

    2015-12-01

    Full Text Available The aim of this pilot study was to apply a novel combined metabolomic and proteomic approach in analysis of gestational diabetes mellitus. The investigation was performed with plasma samples derived from pregnant women with diagnosed gestational diabetes mellitus (n = 18 and a matched control group (n = 13. The mass spectrometry-based analyses allowed to determine 42 free amino acids and low molecular-weight peptide profiles. Different expressions of several peptides and altered amino acid profiles were observed in the analyzed groups. The combination of proteomic and metabolomic data allowed obtaining the model with a high discriminatory power, where amino acids ethanolamine, l-citrulline, l-asparagine, and peptide ions with m/z 1488.59; 4111.89 and 2913.15 had the highest contribution to the model. The sensitivity (94.44% and specificity (84.62%, as well as the total group membership classification value (90.32% calculated from the post hoc classification matrix of a joint model were the highest when compared with a single analysis of either amino acid levels or peptide ion intensities. The obtained results indicated a high potential of integration of proteomic and metabolomics analysis regardless the sample size. This promising approach together with clinical evaluation of the subjects can also be used in the study of other diseases.

  17. Identification Of Protein Vaccine Candidates Using Comprehensive Proteomic Analysis Strategies

    National Research Council Canada - National Science Library

    Rohrbough, James G

    2007-01-01

    Presented in this dissertation are proteomic analysis studies focused on identifying proteins to be used as vaccine candidates against Coccidioidomycosis, a potentially fatal human pulmonary disease...

  18. Proteomic Analysis of the Human Olfactory Bulb.

    Science.gov (United States)

    Dammalli, Manjunath; Dey, Gourav; Madugundu, Anil K; Kumar, Manish; Rodrigues, Benvil; Gowda, Harsha; Siddaiah, Bychapur Gowrishankar; Mahadevan, Anita; Shankar, Susarla Krishna; Prasad, Thottethodi Subrahmanya Keshava

    2017-08-01

    The importance of olfaction to human health and disease is often underappreciated. Olfactory dysfunction has been reported in association with a host of common complex diseases, including neurological diseases such as Alzheimer's disease and Parkinson's disease. For health, olfaction or the sense of smell is also important for most mammals, for optimal engagement with their environment. Indeed, animals have developed sophisticated olfactory systems to detect and interpret the rich information presented to them to assist in day-to-day activities such as locating food sources, differentiating food from poisons, identifying mates, promoting reproduction, avoiding predators, and averting death. In this context, the olfactory bulb is a vital component of the olfactory system receiving sensory information from the axons of the olfactory receptor neurons located in the nasal cavity and the first place that processes the olfactory information. We report in this study original observations on the human olfactory bulb proteome in healthy subjects, using a high-resolution mass spectrometry-based proteomic approach. We identified 7750 nonredundant proteins from human olfactory bulbs. Bioinformatics analysis of these proteins showed their involvement in biological processes associated with signal transduction, metabolism, transport, and olfaction. These new observations provide a crucial baseline molecular profile of the human olfactory bulb proteome, and should assist the future discovery of biomarker proteins and novel diagnostics associated with diseases characterized by olfactory dysfunction.

  19. Integrated transcriptomic and proteomic evaluation of gentamicin nephrotoxicity in rats

    International Nuclear Information System (INIS)

    Com, Emmanuelle; Boitier, Eric; Marchandeau, Jean-Pierre; Brandenburg, Arnd; Schroeder, Susanne; Hoffmann, Dana; Mally, Angela; Gautier, Jean-Charles

    2012-01-01

    Gentamicin is an aminoglycoside antibiotic, which induces renal tubular necrosis in rats. In the context of the European InnoMed PredTox project, transcriptomic and proteomic studies were performed to provide new insights into the molecular mechanisms of gentamicin-induced nephrotoxicity. Male Wistar rats were treated with 25 and 75 mg/kg/day subcutaneously for 1, 3 and 14 days. Histopathology observations showed mild tubular degeneration/necrosis and regeneration and moderate mononuclear cell infiltrate after long-term treatment. Transcriptomic data indicated a strong treatment-related gene expression modulation in kidney and blood cells at the high dose after 14 days of treatment, with the regulation of 463 and 3241 genes, respectively. Of note, the induction of NF-kappa B pathway via the p38 MAPK cascade in the kidney, together with the activation of T-cell receptor signaling in blood cells were suggestive of inflammatory processes in relation with the recruitment of mononuclear cells in the kidney. Proteomic results showed a regulation of 163 proteins in kidney at the high dose after 14 days of treatment. These protein modulations were suggestive of a mitochondrial dysfunction with impairment of cellular energy production, induction of oxidative stress, an effect on protein biosynthesis and on cellular assembly and organization. Proteomic results also provided clues for potential nephrotoxicity biomarkers such as AGAT and PRBP4 which were strongly modulated in the kidney. Transcriptomic and proteomic data turned out to be complementary and their integration gave a more comprehensive insight into the putative mode of nephrotoxicity of gentamicin which was in accordance with histopathological findings. -- Highlights: ► Gentamicin induces renal tubular necrosis in rats. ► The mechanisms of gentamicin nephrotoxicity remain still elusive. ► Transcriptomic and proteomic analyses were performed to study this toxicity in rats. ► Transcriptomic and proteomic

  20. Proteomic Analysis of Human Tooth Pulp: Proteomics of Human Tooth

    Czech Academy of Sciences Publication Activity Database

    Eckhardt, Adam; Jágr, Michal; Pataridis, Statis; Mikšík, Ivan

    2014-01-01

    Roč. 40, č. 12 (2014), s. 1961-1966 ISSN 0099-2399 R&D Projects: GA ČR(CZ) GA13-17224S; GA ČR(CZ) GAP206/12/0453; GA MZd(CZ) NT14324 Institutional support: RVO:67985823 Keywords : dentin * human pulp * tandem mass spectrometry * tooth proteome * 2-dimensional gel electrophoresis Subject RIV: FF - HEENT, Dentistry Impact factor: 3.375, year: 2014

  1. Proteomic Analysis of Bovine Nucleolus

    Institute of Scientific and Technical Information of China (English)

    Amrutlal K.Patel; Doug Olson; Suresh K. Tikoo

    2010-01-01

    Nucleolus is the most prominent subnuclear structure, which performs a wide variety of functions in the eu-karyotic cellular processes. In order to understand the structural and functional role of the nucleoli in bovine cells,we analyzed the proteomie composition of the bovine nueleoli. The nucleoli were isolated from Madin Darby bo-vine kidney cells and subjected to proteomie analysis by LC-MS/MS after fractionation by SDS-PAGE and strongcation exchange chromatography. Analysis of the data using the Mascot database search and the GPM databasesearch identified 311 proteins in the bovine nucleoli, which contained 22 proteins previously not identified in theproteomic analysis of human nucleoli. Analysis of the identified proteins using the GoMiner software suggestedthat the bovine nueleoli contained proteins involved in ribosomal biogenesis, cell cycle control, transcriptional,translational and post-translational regulation, transport, and structural organization.

  2. Integration of genomics, proteomics, and imaging for cardiac stem cell therapy

    International Nuclear Information System (INIS)

    Chun, Hyung J.; Wilson, Kitch O.; Huang, Mei; Wu, Joseph C.

    2007-01-01

    Cardiac stem cell therapy is beginning to mature as a valid treatment for heart disease. As more clinical trials utilizing stem cells emerge, it is imperative to establish the mechanisms by which stem cells confer benefit in cardiac diseases. In this paper, we review three methods - molecular cellular imaging, gene expression profiling, and proteomic analysis - that can be integrated to provide further insights into the role of this emerging therapy. (orig.)

  3. Data from proteomic characterization and comparison of mammalian milk fat globule proteomes by iTRAQ analysis

    Directory of Open Access Journals (Sweden)

    Yongxin Yang

    2015-06-01

    Full Text Available Milk fat globules memebrane (MFGM-enriched proteomes from Holstein, Jersey, yak, buffalo, goat, camel, horse, and human were extracted and identified by an iTRAQ quantification proteomic approach. Proteomes data were analyzed by bioinformatic and multivariate statistical analysis and used to present the characteristic traits of the MFGM proteins among the studied mammals. The data of this study are also related to the research article “Proteomic characterization and comparison of mammalian milk fat globule proteomes by iTRAQ analysis” in the Journal of Proteomics [1].

  4. Urine sample preparation for proteomic analysis.

    Science.gov (United States)

    Olszowy, Pawel; Buszewski, Boguslaw

    2014-10-01

    Sample preparation for both environmental and more importantly biological matrices is a bottleneck of all kinds of analytical processes. In the case of proteomic analysis this element is even more important due to the amount of cross-reactions that should be taken into consideration. The incorporation of new post-translational modifications, protein hydrolysis, or even its degradation is possible as side effects of proteins sample processing. If protocols are evaluated appropriately, then identification of such proteins does not bring difficulties. However, if structural changes are provided without sufficient attention then protein sequence coverage will be reduced or even identification of such proteins could be impossible. This review summarizes obstacles and achievements in protein sample preparation of urine for proteome analysis using different tools for mass spectrometry analysis. The main aim is to present comprehensively the idea of urine application as a valuable matrix. This article is dedicated to sample preparation and application of urine mainly in novel cancer biomarkers discovery. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  5. Proteome analysis in the assessment of ageing.

    Science.gov (United States)

    Nkuipou-Kenfack, Esther; Koeck, Thomas; Mischak, Harald; Pich, Andreas; Schanstra, Joost P; Zürbig, Petra; Schumacher, Björn

    2014-11-01

    Based on demographic trends, the societies in many developed countries are facing an increasing number and proportion of people over the age of 65. The raise in elderly populations along with improved health-care will be concomitant with an increased prevalence of ageing-associated chronic conditions like cardiovascular, renal, and respiratory diseases, arthritis, dementia, and diabetes mellitus. This is expected to pose unprecedented challenges both for individuals and societies and their health care systems. An ultimate goal of ageing research is therefore the understanding of physiological ageing and the achievement of 'healthy' ageing by decreasing age-related pathologies. However, on a molecular level, ageing is a complex multi-mechanistic process whose contributing factors may vary individually, partly overlap with pathological alterations, and are often poorly understood. Proteome analysis potentially allows modelling of these multifactorial processes. This review summarises recent proteomic research on age-related changes identified in animal models and human studies. We combined this information with pathway analysis to identify molecular mechanisms associated with ageing. We identified some molecular pathways that are affected in most or even all organs and others that are organ-specific. However, appropriately powered studies are needed to confirm these findings based in in silico evaluation. Copyright © 2014 Elsevier B.V. All rights reserved.

  6. Microfluidics: Functionality Integration for Proteomic Applications

    DEFF Research Database (Denmark)

    Petersen, Daria

    2006-01-01

    , fabrikation og karakterisering af integrerede mikrochips til protein analyser med forøget detektionsgrænse; 2) en ny samleproces af glas-mikrosystemer til ’miniaturized total analysis systems’ (-TAS); og 3) design samt fabrikation af glas mikrosystemer til kapillær elektroforese (CE) separering integreret...... demonstration af ’on-chip’ protein koncentrering integreret med ’on-chip’, ikke-kovalent mærkning. I-kolonne og efter-kolonne protein mærkningsstrategier påvises i rapporten at være forenelig med protein koncentrering, ved brug af et ’sol-gel’ filter, efterfulgt af en ’sieving’ elektroforese (CSE...

  7. Proteome analysis of the hypercholestrolemic rat, RICO

    International Nuclear Information System (INIS)

    Cho, S.Y.; Park, K.-S.; Paik, Y.-K.; Seong, J.-K.

    2001-01-01

    In an attempt to develop novel markers for hypercholesterolemia, hepatic tissues and serum prepared from hypeicholesterolemic rat (i e RICO) were analyzed by two-dimensional electrophoresis (2DE) and matrix-assisted laser desorption ionization mass spectrometry (MALDI-ToF). Results were compared to those of paired inbreed rat (WKY). Comparative analysis of the respective spot patterns in 2DE revealed that the numbers of differential expression proteins were identified in serum and liver tissues of RICO. Some of the representative proteins annotated in 2DE were apolipoprotein family and numerous lipid metabolism related proteins. Especially, we found that protein disulfide isomerase subunits (ER-60) in 2DE have differential post-translational modification pattern by MALDI-ToF analysis. Our results suggest that the proteomic analysis of these proteins might be a novel approach to identify the molecular events in detail during lipid disorder such atherosclerosis

  8. Integrated genomics and proteomics of the Torpedo californica electric organ: concordance with the mammalian neuromuscular junction

    Directory of Open Access Journals (Sweden)

    Mate Suzanne E

    2011-05-01

    Full Text Available Abstract Background During development, the branchial mesoderm of Torpedo californica transdifferentiates into an electric organ capable of generating high voltage discharges to stun fish. The organ contains a high density of cholinergic synapses and has served as a biochemical model for the membrane specialization of myofibers, the neuromuscular junction (NMJ. We studied the genome and proteome of the electric organ to gain insight into its composition, to determine if there is concordance with skeletal muscle and the NMJ, and to identify novel synaptic proteins. Results Of 435 proteins identified, 300 mapped to Torpedo cDNA sequences with ≥2 peptides. We identified 14 uncharacterized proteins in the electric organ that are known to play a role in acetylcholine receptor clustering or signal transduction. In addition, two human open reading frames, C1orf123 and C6orf130, showed high sequence similarity to electric organ proteins. Our profile lists several proteins that are highly expressed in skeletal muscle or are muscle specific. Synaptic proteins such as acetylcholinesterase, acetylcholine receptor subunits, and rapsyn were present in the electric organ proteome but absent in the skeletal muscle proteome. Conclusions Our integrated genomic and proteomic analysis supports research describing a muscle-like profile of the organ. We show that it is a repository of NMJ proteins but we present limitations on its use as a comprehensive model of the NMJ. Finally, we identified several proteins that may become candidates for signaling proteins not previously characterized as components of the NMJ.

  9. Proteomic analysis of Aspergillus fumigatus - clinical implications.

    Science.gov (United States)

    Moloney, Nicola M; Owens, Rebecca A; Doyle, Sean

    2016-07-01

    Aspergillus fumigatus is a ubiquitous saprophytic fungus capable of producing small airborne spores, which are frequently inhaled by humans. In healthy individuals, the fungus is rapidly cleared by innate mechanisms, including immune cells. However, in individuals with impaired lung function or immunosuppression the spores can germinate and prompt severe allergic responses, and disease with limited or extensive invasiveness. The traits that make A. fumigatus a successful colonizer and pathogen of humans are multi-factorial. Thus, a global investigative approach is required to elucidate the mechanisms utilized by the fungus to cause disease. Expert commentary: In doing so, a better understanding of disease pathology can be achieved with improved therapeutic/diagnostic solutions, thereby improving patient outcome. Proteomic analysis permits such investigations and recent work has yielded insight into these mechanisms.

  10. In silico proteome analysis to facilitate proteomics experiments using mass spectrometry

    Directory of Open Access Journals (Sweden)

    Lindo Micheal

    2003-08-01

    Full Text Available Abstract Proteomics experiments typically involve protein or peptide separation steps coupled to the identification of many hundreds to thousands of peptides by mass spectrometry. Development of methodology and instrumentation in this field is proceeding rapidly, and effective software is needed to link the different stages of proteomic analysis. We have developed an application, proteogest, written in Perl that generates descriptive and statistical analyses of the biophysical properties of multiple (e.g. thousands protein sequences submitted by the user, for instance protein sequences inferred from the complete genome sequence of a model organism. The application also carries out in silico proteolytic digestion of the submitted proteomes, or subsets thereof, and the distribution of biophysical properties of the resulting peptides is presented. proteogest is customizable, the user being able to select many options, for instance the cleavage pattern of the digestion treatment or the presence of modifications to specific amino acid residues. We show how proteogest can be used to compare the proteomes and digested proteome products of model organisms, to examine the added complexity generated by modification of residues, and to facilitate the design of proteomics experiments for optimal representation of component proteins.

  11. Global Proteome Analysis of Leptospira interrogans

    Science.gov (United States)

    Comparative global proteome analyses were performed on Leptospira interrogans serovar Copenhageni grown under conventional in vitro conditions and those mimicking in vivo conditions (iron limitation and serum presence). Proteomic analyses were conducted using iTRAQ and LC-ESI-tandem mass spectrometr...

  12. Proteome analysis of Saccharomyces cerevisiae: a methodological outline

    DEFF Research Database (Denmark)

    Fey, S J; Nawrocki, A; Görg, A

    1997-01-01

    Proteome analysis offers a unique means of identifying important proteins, characterizing their modifications and beginning to describe their function. This is achieved through the combination of two technologies: protein separation and selection by two-dimensional gel electrophoresis, and protei...

  13. Prediction of acute coronary syndromes by urinary proteome analysis.

    Directory of Open Access Journals (Sweden)

    Nay M Htun

    Full Text Available Identification of individuals who are at risk of suffering from acute coronary syndromes (ACS may allow to introduce preventative measures. We aimed to identify ACS-related urinary peptides, that combined as a pattern can be used as prognostic biomarker. Proteomic data of 252 individuals enrolled in four prospective studies from Australia, Europe and North America were analyzed. 126 of these had suffered from ACS within a period of up to 5 years post urine sampling (cases. Proteomic analysis of 84 cases and 84 matched controls resulted in the discovery of 75 ACS-related urinary peptides. Combining these to a peptide pattern, we established a prognostic biomarker named Acute Coronary Syndrome Predictor 75 (ACSP75. ACSP75 demonstrated reasonable prognostic discrimination (c-statistic = 0.664, which was similar to Framingham risk scoring (c-statistics = 0.644 in a validation cohort of 42 cases and 42 controls. However, generating by a composite algorithm named Acute Coronary Syndrome Composite Predictor (ACSCP, combining the biomarker pattern ACSP75 with the previously established urinary proteomic biomarker CAD238 characterizing coronary artery disease as the underlying aetiology, and age as a risk factor, further improved discrimination (c-statistic = 0.751 resulting in an added prognostic value over Framingham risk scoring expressed by an integrated discrimination improvement of 0.273 ± 0.048 (P < 0.0001 and net reclassification improvement of 0.405 ± 0.113 (P = 0.0007. In conclusion, we demonstrate that urinary peptide biomarkers have the potential to predict future ACS events in asymptomatic patients. Further large scale studies are warranted to determine the role of urinary biomarkers in clinical practice.

  14. PROTEINCHALLENGE: Crowd sourcing in proteomics analysis and software development

    DEFF Research Database (Denmark)

    Martin, Sarah F.; Falkenberg, Heiner; Dyrlund, Thomas Franck

    2013-01-01

    , including arguments for community-wide open source software development and “big data” compatible solutions for the future. For the meantime, we have laid out ten top tips for data processing. With these at hand, a first large-scale proteomics analysis hopefully becomes less daunting to navigate.......However there is clearly a real need for robust tools, standard operating procedures and general acceptance of best practises. Thus we submit to the proteomics community a call for a community-wide open set of proteomics analysis challenges—PROTEINCHALLENGE—that directly target and compare data analysis workflows......In large-scale proteomics studies there is a temptation, after months of experimental work, to plug resulting data into a convenient—if poorly implemented—set of tools, which may neither do the data justice nor help answer the scientific question. In this paper we have captured key concerns...

  15. Proteomics wants cRacker: automated standardized data analysis of LC-MS derived proteomic data.

    Science.gov (United States)

    Zauber, Henrik; Schulze, Waltraud X

    2012-11-02

    The large-scale analysis of thousands of proteins under various experimental conditions or in mutant lines has gained more and more importance in hypothesis-driven scientific research and systems biology in the past years. Quantitative analysis by large scale proteomics using modern mass spectrometry usually results in long lists of peptide ion intensities. The main interest for most researchers, however, is to draw conclusions on the protein level. Postprocessing and combining peptide intensities of a proteomic data set requires expert knowledge, and the often repetitive and standardized manual calculations can be time-consuming. The analysis of complex samples can result in very large data sets (lists with several 1000s to 100,000 entries of different peptides) that cannot easily be analyzed using standard spreadsheet programs. To improve speed and consistency of the data analysis of LC-MS derived proteomic data, we developed cRacker. cRacker is an R-based program for automated downstream proteomic data analysis including data normalization strategies for metabolic labeling and label free quantitation. In addition, cRacker includes basic statistical analysis, such as clustering of data, or ANOVA and t tests for comparison between treatments. Results are presented in editable graphic formats and in list files.

  16. Plasma proteome analysis of cervical intraepithelial neoplasia

    Indian Academy of Sciences (India)

    ... Malaysia and University of Malaya Centre For Proteomics Research (UMCPR), Universiti Kebangsaan Malaysia, Kuala Lumpur, Malaysia; Department of Clinical Oral Biology, Faculty of Dentistry; Universiti Kebangsaan Malaysia, Kuala Lumpur, Malaysia; Department of Obstetrics and Gynecology; Universiti Kebangsaan ...

  17. Proteome-wide analysis and diel proteomic profiling of the cyanobacterium Arthrospira platensis PCC 8005.

    Directory of Open Access Journals (Sweden)

    Sabine Matallana-Surget

    Full Text Available The filamentous cyanobacterium Arthrospira platensis has a long history of use as a food supply and it has been used by the European Space Agency in the MELiSSA project, an artificial microecosystem which supports life during long-term manned space missions. This study assesses progress in the field of cyanobacterial shotgun proteomics and light/dark diurnal cycles by focusing on Arthrospira platensis. Several fractionation workflows including gel-free and gel-based protein/peptide fractionation procedures were used and combined with LC-MS/MS analysis, enabling the overall identification of 1306 proteins, which represents 21% coverage of the theoretical proteome. A total of 30 proteins were found to be significantly differentially regulated under light/dark growth transition. Interestingly, most of the proteins showing differential abundance were related to photosynthesis, the Calvin cycle and translation processes. A novel aspect and major achievement of this work is the successful improvement of the cyanobacterial proteome coverage using a 3D LC-MS/MS approach, based on an immobilized metal affinity chromatography, a suitable tool that enabled us to eliminate the most abundant protein, the allophycocyanin. We also demonstrated that cell growth follows a light/dark cycle in A. platensis. This preliminary proteomic study has highlighted new characteristics of the Arthrospira platensis proteome in terms of diurnal regulation.

  18. Comprehensive analysis of temporal alterations in cellular proteome of Bacillus subtilis under curcumin treatment.

    Directory of Open Access Journals (Sweden)

    Panga Jaipal Reddy

    Full Text Available Curcumin is a natural dietary compound with antimicrobial activity against various gram positive and negative bacteria. This study aims to investigate the proteome level alterations in Bacillus subtilis due to curcumin treatment and identification of its molecular/cellular targets to understand the mechanism of action. We have performed a comprehensive proteomic analysis of B. subtilis AH75 strain at different time intervals of curcumin treatment (20, 60 and 120 min after the drug exposure, three replicates to compare the protein expression profiles using two complementary quantitative proteomic techniques, 2D-DIGE and iTRAQ. To the best of our knowledge, this is the first comprehensive longitudinal investigation describing the effect of curcumin treatment on B. subtilis proteome. The proteomics analysis revealed several interesting targets such UDP-N-acetylglucosamine 1-carboxyvinyltransferase 1, putative septation protein SpoVG and ATP-dependent Clp protease proteolytic subunit. Further, in silico pathway analysis using DAVID and KOBAS has revealed modulation of pathways related to the fatty acid metabolism and cell wall synthesis, which are crucial for cell viability. Our findings revealed that curcumin treatment lead to inhibition of the cell wall and fatty acid synthesis in addition to differential expression of many crucial proteins involved in modulation of bacterial metabolism. Findings obtained from proteomics analysis were further validated using 5-cyano-2,3-ditolyl tetrazolium chloride (CTC assay for respiratory activity, resazurin assay for metabolic activity and membrane integrity assay by potassium and inorganic phosphate leakage measurement. The gene expression analysis of selected cell wall biosynthesis enzymes has strengthened the proteomics findings and indicated the major effect of curcumin on cell division.

  19. MPLEx: a Robust and Universal Protocol for Single-Sample Integrative Proteomic, Metabolomic, and Lipidomic Analyses

    Energy Technology Data Exchange (ETDEWEB)

    Nakayasu, Ernesto S.; Nicora, Carrie D.; Sims, Amy C.; Burnum-Johnson, Kristin E.; Kim, Young-Mo; Kyle, Jennifer E.; Matzke, Melissa M.; Shukla, Anil K.; Chu, Rosalie K.; Schepmoes, Athena A.; Jacobs, Jon M.; Baric, Ralph S.; Webb-Robertson, Bobbie-Jo; Smith, Richard D.; Metz, Thomas O.; Chia, Nicholas

    2016-05-03

    ABSTRACT

    Integrative multi-omics analyses can empower more effective investigation and complete understanding of complex biological systems. Despite recent advances in a range of omics analyses, multi-omic measurements of the same sample are still challenging and current methods have not been well evaluated in terms of reproducibility and broad applicability. Here we adapted a solvent-based method, widely applied for extracting lipids and metabolites, to add proteomics to mass spectrometry-based multi-omics measurements. Themetabolite,protein, andlipidextraction (MPLEx) protocol proved to be robust and applicable to a diverse set of sample types, including cell cultures, microbial communities, and tissues. To illustrate the utility of this protocol, an integrative multi-omics analysis was performed using a lung epithelial cell line infected with Middle East respiratory syndrome coronavirus, which showed the impact of this virus on the host glycolytic pathway and also suggested a role for lipids during infection. The MPLEx method is a simple, fast, and robust protocol that can be applied for integrative multi-omic measurements from diverse sample types (e.g., environmental,in vitro, and clinical).

    IMPORTANCEIn systems biology studies, the integration of multiple omics measurements (i.e., genomics, transcriptomics, proteomics, metabolomics, and lipidomics) has been shown to provide a more complete and informative view of biological pathways. Thus, the prospect of extracting different types of molecules (e.g., DNAs, RNAs, proteins, and metabolites) and performing multiple omics measurements on single samples is very attractive, but such studies are challenging due to the fact that the extraction conditions differ according to the molecule type. Here, we adapted an organic solvent-based extraction method that demonstrated

  20. f-divergence cutoff index to simultaneously identify differential expression in the integrated transcriptome and proteome

    OpenAIRE

    Tang, Shaojun; Hemberg, Martin; Cansizoglu, Ertugrul; Belin, Stephane; Kosik, Kenneth; Kreiman, Gabriel; Steen, Hanno; Steen, Judith

    2016-01-01

    The ability to integrate 'omics' (i.e., transcriptomics and proteomics) is becoming increasingly important to the understanding of regulatory mechanisms. There are currently no tools available to identify differentially expressed genes (DEGs)across different 'omics'data types or multi-dimensional data including time courses. We present a model capable of simultaneously identifying DEGs from continuous and discrete transcriptomic, proteomic and integrated proteogenomic data. We show that...

  1. An Innovative Approach for The Integration of Proteomics and Metabolomics Data In Severe Septic Shock Patients Stratified for Mortality.

    Science.gov (United States)

    Cambiaghi, Alice; Díaz, Ramón; Martinez, Julia Bauzá; Odena, Antonia; Brunelli, Laura; Caironi, Pietro; Masson, Serge; Baselli, Giuseppe; Ristagno, Giuseppe; Gattinoni, Luciano; de Oliveira, Eliandre; Pastorelli, Roberta; Ferrario, Manuela

    2018-04-27

    In this work, we examined plasma metabolome, proteome and clinical features in patients with severe septic shock enrolled in the multicenter ALBIOS study. The objective was to identify changes in the levels of metabolites involved in septic shock progression and to integrate this information with the variation occurring in proteins and clinical data. Mass spectrometry-based targeted metabolomics and untargeted proteomics allowed us to quantify absolute metabolites concentration and relative proteins abundance. We computed the ratio D7/D1 to take into account their variation from day 1 (D1) to day 7 (D7) after shock diagnosis. Patients were divided into two groups according to 28-day mortality. Three different elastic net logistic regression models were built: one on metabolites only, one on metabolites and proteins and one to integrate metabolomics and proteomics data with clinical parameters. Linear discriminant analysis and Partial least squares Discriminant Analysis were also implemented. All the obtained models correctly classified the observations in the testing set. By looking at the variable importance (VIP) and the selected features, the integration of metabolomics with proteomics data showed the importance of circulating lipids and coagulation cascade in septic shock progression, thus capturing a further layer of biological information complementary to metabolomics information.

  2. A community proposal to integrate proteomics activities in ELIXIR [version 1; referees: 2 approved

    Directory of Open Access Journals (Sweden)

    Juan Antonio Vizcaíno

    2017-06-01

    Full Text Available Computational approaches have been major drivers behind the progress of proteomics in recent years. The aim of this white paper is to provide a framework for integrating computational proteomics into ELIXIR in the near future, and thus to broaden the portfolio of omics technologies supported by this European distributed infrastructure. This white paper is the direct result of a strategy meeting on ‘The Future of Proteomics in ELIXIR’ that took place in March 2017 in Tübingen (Germany, and involved representatives of eleven ELIXIR nodes.   These discussions led to a list of priority areas in computational proteomics that would complement existing activities and close gaps in the portfolio of tools and services offered by ELIXIR so far. We provide some suggestions on how these activities could be integrated into ELIXIR’s existing platforms, and how it could lead to a new ELIXIR use case in proteomics. We also highlight connections to the related field of metabolomics, where similar activities are ongoing. This white paper could thus serve as a starting point for the integration of computational proteomics into ELIXIR. Over the next few months we will be working closely with all stakeholders involved, and in particular with other representatives of the proteomics community, to further refine this paper.

  3. Proteome Analysis of Rheumatoid Arthritis Gut Mucosa

    DEFF Research Database (Denmark)

    Bennike, Tue Bjerg; Ellingsen, Torkell; Glerup, Henning

    2017-01-01

    Rheumatoid arthritis (RA) is an inflammatory joint disease leading to cartilage damage and ultimately impaired joint function. To gain new insight into the systemic immune manifestations of RA, we characterized the colon mucosa proteome from 11 RA-patients and 10 healthy controls. The biopsies were...

  4. Proteomic Analysis of Chicken Skeletal Muscle during Embryonic Development

    Directory of Open Access Journals (Sweden)

    Hongjia Ouyang

    2017-05-01

    Full Text Available Embryonic growth and development of skeletal muscle is a major determinant of muscle mass, and has a significant effect on meat production in chicken. To assess the protein expression profiles during embryonic skeletal muscle development, we performed a proteomics analysis using isobaric tags for relative and absolute quantification (iTRAQ in leg muscle tissues of female Xinghua chicken at embryonic age (E 11, E16, and 1-day post hatch (D1. We identified 3,240 proteins in chicken embryonic muscle and 491 of them were differentially expressed (fold change ≥ 1.5 or ≤ 0.666 and p < 0.05. There were 19 up- and 32 down-regulated proteins in E11 vs. E16 group, 238 up- and 227 down-regulated proteins in E11 vs. D1 group, and 13 up- and 5 down-regulated proteins in E16 vs. D1 group. Protein interaction network analyses indicated that these differentially expressed proteins were mainly involved in the pathway of protein synthesis, muscle contraction, and oxidative phosphorylation. Integrative analysis of proteome and our previous transcriptome data found 189 differentially expressed proteins that correlated with their mRNA level. The interactions between these proteins were also involved in muscle contraction and oxidative phosphorylation pathways. The lncRNA-protein interaction network found four proteins DMD, MYL3, TNNI2, and TNNT3 that are all involved in muscle contraction and may be lncRNA regulated. These results provide several candidate genes for further investigation into the molecular mechanisms of chicken embryonic muscle development, and enable us to better understanding their regulation networks and biochemical pathways.

  5. Proteomic analysis of endothelial cold-adaptation

    Directory of Open Access Journals (Sweden)

    Zieger Michael AJ

    2011-12-01

    Full Text Available Abstract Background Understanding how human cells in tissue culture adapt to hypothermia may aid in developing new clinical procedures for improved ischemic and hypothermic protection. Human coronary artery endothelial cells grown to confluence at 37°C and then transferred to 25°C become resistant over time to oxidative stress and injury induced by 0°C storage and rewarming. This protection correlates with an increase in intracellular glutathione at 25°C. To help understand the molecular basis of endothelial cold-adaptation, isolated proteins from cold-adapted (25°C/72 h and pre-adapted cells were analyzed by quantitative proteomic methods and differentially expressed proteins were categorized using the DAVID Bioinformatics Resource. Results Cells adapted to 25°C expressed changes in the abundance of 219 unique proteins representing a broad range of categories such as translation, glycolysis, biosynthetic (anabolic processes, NAD, cytoskeletal organization, RNA processing, oxidoreductase activity, response-to-stress and cell redox homeostasis. The number of proteins that decreased significantly with cold-adaptation exceeded the number that increased by 2:1. Almost half of the decreases were associated with protein metabolic processes and a third were related to anabolic processes including protein, DNA and fatty acid synthesis. Changes consistent with the suppression of cytoskeletal dynamics provided further evidence that cold-adapted cells are in an energy conserving state. Among the specific changes were increases in the abundance and activity of redox proteins glutathione S-transferase, thioredoxin and thioredoxin reductase, which correlated with a decrease in oxidative stress, an increase in protein glutathionylation, and a recovery of reduced protein thiols during rewarming from 0°C. Increases in S-adenosylhomocysteine hydrolase and nicotinamide phosphoribosyltransferase implicate a central role for the methionine

  6. freeQuant: A Mass Spectrometry Label-Free Quantification Software Tool for Complex Proteome Analysis.

    Science.gov (United States)

    Deng, Ning; Li, Zhenye; Pan, Chao; Duan, Huilong

    2015-01-01

    Study of complex proteome brings forward higher request for the quantification method using mass spectrometry technology. In this paper, we present a mass spectrometry label-free quantification tool for complex proteomes, called freeQuant, which integrated quantification with functional analysis effectively. freeQuant consists of two well-integrated modules: label-free quantification and functional analysis with biomedical knowledge. freeQuant supports label-free quantitative analysis which makes full use of tandem mass spectrometry (MS/MS) spectral count, protein sequence length, shared peptides, and ion intensity. It adopts spectral count for quantitative analysis and builds a new method for shared peptides to accurately evaluate abundance of isoforms. For proteins with low abundance, MS/MS total ion count coupled with spectral count is included to ensure accurate protein quantification. Furthermore, freeQuant supports the large-scale functional annotations for complex proteomes. Mitochondrial proteomes from the mouse heart, the mouse liver, and the human heart were used to evaluate the usability and performance of freeQuant. The evaluation showed that the quantitative algorithms implemented in freeQuant can improve accuracy of quantification with better dynamic range.

  7. Analysis of high accuracy, quantitative proteomics data in the MaxQB database.

    Science.gov (United States)

    Schaab, Christoph; Geiger, Tamar; Stoehr, Gabriele; Cox, Juergen; Mann, Matthias

    2012-03-01

    MS-based proteomics generates rapidly increasing amounts of precise and quantitative information. Analysis of individual proteomic experiments has made great strides, but the crucial ability to compare and store information across different proteome measurements still presents many challenges. For example, it has been difficult to avoid contamination of databases with low quality peptide identifications, to control for the inflation in false positive identifications when combining data sets, and to integrate quantitative data. Although, for example, the contamination with low quality identifications has been addressed by joint analysis of deposited raw data in some public repositories, we reasoned that there should be a role for a database specifically designed for high resolution and quantitative data. Here we describe a novel database termed MaxQB that stores and displays collections of large proteomics projects and allows joint analysis and comparison. We demonstrate the analysis tools of MaxQB using proteome data of 11 different human cell lines and 28 mouse tissues. The database-wide false discovery rate is controlled by adjusting the project specific cutoff scores for the combined data sets. The 11 cell line proteomes together identify proteins expressed from more than half of all human genes. For each protein of interest, expression levels estimated by label-free quantification can be visualized across the cell lines. Similarly, the expression rank order and estimated amount of each protein within each proteome are plotted. We used MaxQB to calculate the signal reproducibility of the detected peptides for the same proteins across different proteomes. Spearman rank correlation between peptide intensity and detection probability of identified proteins was greater than 0.8 for 64% of the proteome, whereas a minority of proteins have negative correlation. This information can be used to pinpoint false protein identifications, independently of peptide database

  8. Proteomic analysis of blastema formation in regenerating axolotl limbs

    Directory of Open Access Journals (Sweden)

    Nye Holly LD

    2009-11-01

    Full Text Available Abstract Background Following amputation, urodele salamander limbs reprogram somatic cells to form a blastema that self-organizes into the missing limb parts to restore the structure and function of the limb. To help understand the molecular basis of blastema formation, we used quantitative label-free liquid chromatography-mass spectrometry/mass spectrometry (LC-MS/MS-based methods to analyze changes in the proteome that occurred 1, 4 and 7 days post amputation (dpa through the mid-tibia/fibula of axolotl hind limbs. Results We identified 309 unique proteins with significant fold change relative to controls (0 dpa, representing 10 biological process categories: (1 signaling, (2 Ca2+ binding and translocation, (3 transcription, (4 translation, (5 cytoskeleton, (6 extracellular matrix (ECM, (7 metabolism, (8 cell protection, (9 degradation, and (10 cell cycle. In all, 43 proteins exhibited exceptionally high fold changes. Of these, the ecotropic viral integrative factor 5 (EVI5, a cell cycle-related oncoprotein that prevents cells from entering the mitotic phase of the cell cycle prematurely, was of special interest because its fold change was exceptionally high throughout blastema formation. Conclusion Our data were consistent with previous studies indicating the importance of inositol triphosphate and Ca2+ signaling in initiating the ECM and cytoskeletal remodeling characteristic of histolysis and cell dedifferentiation. In addition, the data suggested that blastema formation requires several mechanisms to avoid apoptosis, including reduced metabolism, differential regulation of proapoptotic and antiapoptotic proteins, and initiation of an unfolded protein response (UPR. Since there is virtually no mitosis during blastema formation, we propose that high levels of EVI5 function to arrest dedifferentiated cells somewhere in the G1/S/G2 phases of the cell cycle until they have accumulated under the wound epidermis and enter mitosis in response to

  9. Comprehensive data analysis of human ureter proteome

    Directory of Open Access Journals (Sweden)

    Sameh Magdeldin

    2016-03-01

    Full Text Available Comprehensive human ureter proteome dataset was generated from OFFGel fractionated ureter samples. Our result showed that among 2217 non-redundant ureter proteins, 751 protein candidates (33.8% were detected in urine as urinary protein/polypeptide or exosomal protein. On the other hand, comparing ureter protein hits (48 that are not shown in corresponding databases to urinary bladder and prostate human protein atlas databases pinpointed 21 proteins that might be unique to ureter tissue. In conclusion, this finding offers future perspectives for possible identification of ureter disease-associated biomarkers such as ureter carcinoma. In addition, Cytoscape GO annotation was examined on the final ureter dataset to better understand proteins molecular function, biological processes, and cellular component. The ureter proteomic dataset published in this article will provide a valuable resource for researchers working in the field of urology and urine biomarker discovery.

  10. Data from proteome analysis of Lasiodiplodia theobromae (Botryosphaeriaceae

    Directory of Open Access Journals (Sweden)

    Carla C. Uranga

    2017-08-01

    Full Text Available Trunk disease fungi are a global problem affecting many economically important fruiting trees. The Botryosphaeriaceae are a family of trunk disease fungi that require detailed biochemical characterization in order to gain insight into their pathogenicity. The application of a modified Folch extraction to protein extraction from the Botryosphaeriaceae Lasiodiplodia theobromae generated an unprecedented data set of protein identifications from fragmentation analysis and de novo peptide sequencing of its proteome. This article contains data from protein identifications obtained from a database-dependent fragmentation analysis using three different proteomics algorithms (MSGF, Comet and X! Tandem via the SearchGUI proteomics pipeline program and de novo peptide sequencing. Included are data sets of gene ontology annotations using an all-Uniprot ontology database, as well as a Saccharomyces cerevisiae-only and a Candida albicans-only ontology database, in order to discern between those proteins involved in common functions with S. cerevisiae and those in common with the pathogenic yeast C. albicans. Our results reveal the proteome of L. theobromae contains more ontological categories in common to C. albicans, yet possesses a much wider metabolic repertoire than any of the yeasts studied in this work. Many novel proteins of interest were identified for further biochemical characterization and annotation efforts, as further discussed in the article referencing this article (1. Interactive Cytoscape networks of molecular functions of identified peptides using an all-Uniprot ontological database are included. Data, including raw data, are available via ProteomeXchange with identifier PXD005283.

  11. Proteomic analysis of the Theileria annulata schizont.

    OpenAIRE

    Witschi Marc; Xia D; Sanderson Sandy; Baumgartner Martin; Wastling Jonathan; Dobbelaere Dirk

    2013-01-01

    The apicomplexan parasite, Theileria annulata, is the causative agent of tropical theileriosis, a devastating lymphoproliferative disease of cattle. The schizont stage transforms bovine leukocytes and provides an intriguing model to study host/pathogen interactions. The genome of T. annulata has been sequenced and transcriptomic data are rapidly accumulating. In contrast, little is known about the proteome of the schizont, the pathogenic, transforming life cycle stage of the parasite. Using o...

  12. Comprehensive proteomic analysis of human dentin

    Czech Academy of Sciences Publication Activity Database

    Jágr, Michal; Eckhardt, Adam; Pataridis, Statis; Mikšík, Ivan

    2012-01-01

    Roč. 120, č. 4 (2012), s. 259-268 ISSN 0909-8836 R&D Projects: GA ČR(CZ) GA203/08/1428; GA ČR(CZ) GAP206/12/0453 Institutional research plan: CEZ:AV0Z50110509 Institutional support: RVO:67985823 Keywords : dentin * mass spectrometry * proteomics * tooth * two-dimensional gel electrophoresis Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 1.420, year: 2012

  13. Redefining the Breast Cancer Exosome Proteome by Tandem Mass Tag Quantitative Proteomics and Multivariate Cluster Analysis.

    Science.gov (United States)

    Clark, David J; Fondrie, William E; Liao, Zhongping; Hanson, Phyllis I; Fulton, Amy; Mao, Li; Yang, Austin J

    2015-10-20

    Exosomes are microvesicles of endocytic origin constitutively released by multiple cell types into the extracellular environment. With evidence that exosomes can be detected in the blood of patients with various malignancies, the development of a platform that uses exosomes as a diagnostic tool has been proposed. However, it has been difficult to truly define the exosome proteome due to the challenge of discerning contaminant proteins that may be identified via mass spectrometry using various exosome enrichment strategies. To better define the exosome proteome in breast cancer, we incorporated a combination of Tandem-Mass-Tag (TMT) quantitative proteomics approach and Support Vector Machine (SVM) cluster analysis of three conditioned media derived fractions corresponding to a 10 000g cellular debris pellet, a 100 000g crude exosome pellet, and an Optiprep enriched exosome pellet. The quantitative analysis identified 2 179 proteins in all three fractions, with known exosomal cargo proteins displaying at least a 2-fold enrichment in the exosome fraction based on the TMT protein ratios. Employing SVM cluster analysis allowed for the classification 251 proteins as "true" exosomal cargo proteins. This study provides a robust and vigorous framework for the future development of using exosomes as a potential multiprotein marker phenotyping tool that could be useful in breast cancer diagnosis and monitoring disease progression.

  14. PROTEINCHALLENGE: Crowd sourcing in proteomics analysis and software development

    DEFF Research Database (Denmark)

    Martin, Sarah F.; Falkenberg, Heiner; Dyrlund, Thomas Franck

    2013-01-01

    , including arguments for community-wide open source software development and “big data” compatible solutions for the future. For the meantime, we have laid out ten top tips for data processing. With these at hand, a first large-scale proteomics analysis hopefully becomes less daunting to navigate......, with the aim of setting a community-driven gold standard for data handling, reporting and sharing. This article is part of a Special Issue entitled: New Horizons and Applications for Proteomics [EuPA 2012].......In large-scale proteomics studies there is a temptation, after months of experimental work, to plug resulting data into a convenient—if poorly implemented—set of tools, which may neither do the data justice nor help answer the scientific question. In this paper we have captured key concerns...

  15. Proteomic analysis of tissue samples in translational breast cancer research

    DEFF Research Database (Denmark)

    Gromov, Pavel; Moreira, José; Gromova, Irina

    2014-01-01

    In the last decade, many proteomic technologies have been applied, with varying success, to the study of tissue samples of breast carcinoma for protein expression profiling in order to discover protein biomarkers/signatures suitable for: characterization and subtyping of tumors; early diagnosis...... the translation of basic discoveries into the daily breast cancer clinical practice. In particular, we address major issues in experimental design by reviewing the strengths and weaknesses of current proteomic strategies in the context of the analysis of human breast tissue specimens....

  16. Proteomic and genomic analysis of cardiovascular disease

    National Research Council Canada - National Science Library

    Van Eyk, Jennifer; Dunn, M. J

    2003-01-01

    ... to cardiovascular disease. By exploring the various strategies and technical aspects of both, using examples from cardiac or vascular biology, the limitations and the potential of these methods can be clearly seen. The book is divided into three sections: the first focuses on genomics, the second on proteomics, and the third provides an overview of the importance of these two scientific disciplines in drug and diagnostic discovery. The goal of this book is the transfer of their hard-earned lessons to the growing num...

  17. A proteomic analysis of human bile

    DEFF Research Database (Denmark)

    Kristiansen, Troels Zakarias; Bunkenborg, Jakob; Gronborg, Mads

    2004-01-01

    We have carried out a comprehensive characterization of human bile to define the bile proteome. Our approach involved fractionation of bile by one-dimensional gel electrophoresis and lectin affinity chromatography followed by liquid chromatography tandem mass spectrometry. Overall, we identified 87...... unique proteins, including several novel proteins as well as known proteins whose functions are unknown. A large majority of the identified proteins have not been previously described in bile. Using lectin affinity chromatography and enzymatically labeling of asparagine residues carrying glycan moieties...

  18. Quantitative Proteomic Analysis of Sulfolobus solfataricus Membrane Proteins

    NARCIS (Netherlands)

    Pham, T.K.; Sierocinski, P.; Oost, van der J.; Wright, P.C.

    2010-01-01

    A quantitative proteomic analysis of the membrane of the archaeon Sulfolobus solfataricus P2 using iTRAQ was successfully demonstrated in this technical note. The estimated number of membrane proteins of this organism is 883 (predicted based on Gravy score), corresponding to 30 % of the total

  19. Proteomic analysis of cell lines to identify the irinotecan resistance ...

    Indian Academy of Sciences (India)

    MADHU

    was selected from the wild-type LoVo cell line by chronic exposure to irinotecan ... dose–effect curves of anticancer drugs were drawn on semilogarithm .... alcohol metabolites daunorubicinol (Forrest and Gonzalez. 2000; Mordente et al. ..... Chen L, Huang C and Wei Y 2007 Proteomic analysis of liver cancer cells treated ...

  20. Proteomic analysis of Arabidopsis seed germination and priming

    NARCIS (Netherlands)

    Gallardo, K.; Job, C.; Groot, S.P.C.; Puype, M.; Demol, H.; Vandekerckhove, J.; Job, D.

    2001-01-01

    To better understand seed germination, a complex developmental process, we developed a proteome analysis of the model plant Arabidopsis for which complete genome sequence is now available. Among about 1,300 total seed proteins resolved in two-dimensional gels, changes in the abundance (up- and

  1. Data from quantitative label free proteomics analysis of rat spleen

    Directory of Open Access Journals (Sweden)

    Khadar Dudekula

    2016-09-01

    Full Text Available The dataset presented in this work has been obtained using a label-free quantitative proteomic analysis of rat spleen. A robust method for extraction of proteins from rat spleen tissue and LC-MS-MS analysis was developed using a urea and SDS-based buffer. Different fractionation methods were compared. A total of 3484 different proteins were identified from the pool of all experiments run in this study (a total of 2460 proteins with at least two peptides. A total of 1822 proteins were identified from nine non-fractionated pulse gels, 2288 proteins and 2864 proteins were identified by SDS-PAGE fractionation into three and five fractions respectively. The proteomics data are deposited in ProteomeXchange Consortium via PRIDE PXD003520, Progenesis and Maxquant output are presented in the supported information. The generated list of proteins under different regimes of fractionation allow assessing the nature of the identified proteins; variability in the quantitative analysis associated with the different sampling strategy and allow defining a proper number of replicates for future quantitative analysis. Keywords: Spleen, Rat, Protein extraction, Label-free quantitative proteomics

  2. Proteomic analysis of the Arabidopsis nucleolus suggests novel nucleolar functions

    DEFF Research Database (Denmark)

    Pendle, Alison F; Clark, Gillian P; Boon, Reinier

    2005-01-01

    The eukaryotic nucleolus is involved in ribosome biogenesis and a wide range of other RNA metabolism and cellular functions. An important step in the functional analysis of the nucleolus is to determine the complement of proteins of this nuclear compartment. Here, we describe the first proteomic ...

  3. Proteomic analysis of cold stress responses in tobacco seedlings ...

    African Journals Online (AJOL)

    Cold stress is one of the major abiotic stresses limiting the productivity and the geographical distribution of many important crops. To gain a better understanding of cold stress responses in tobacco (Nicotiana tabacum), we carried out a comparative proteomic analysis. Five-week-old tobacco seedlings were treated at 4°C ...

  4. Noninvasive diagnosis of chronic kidney diseases using urinary proteome analysis

    DEFF Research Database (Denmark)

    Siwy, Justyna; Zürbig, Petra; Argilés, Angel

    2017-01-01

    BACKGROUND: In spite of its invasive nature and risks, kidney biopsy is currently required for precise diagnosis of many chronic kidney diseases (CKDs). Here, we explored the hypothesis that analysis of the urinary proteome can discriminate different types of CKD irrespective of the underlying me...

  5. SILK FIBRE DEGRADATION AND ANALYSIS BY PROTEOMICS

    Directory of Open Access Journals (Sweden)

    YUKSELOGLU S.Muge

    2016-05-01

    Full Text Available Silk is one of the promising natural fibres and has a long established history in textile production throughout the centuries. Silk is produced by cultured silk worms, spiders, scorpions, mites and flies. It is extracellular proteinaceous fibres which consist of highly crystalline and insoluble proteins, the fibroins glued with sericin and an amourphous protein. On the other hand, understanding and controlling the degradation of protein materials are important for determining quality and the value of appearance retention in textiles. Hence, for silk textiles, appearance retention is critical value for the quality. And this is one of the key properties directly related to the degree and nature of protein degradation. It is therefore necessary to understand the silk composition and damage to obtain good conservation treatments and long-term preservation especially for the historical silk fabrics. In this study, silk fibre and its properties are briefly introduced along with images on their fibre damages. Additionally, proteomics method which helps to understand the degradation at the molecular level in textiles is introduced. Finally, proteomic evaluation of silk is summarized according to the researchers carried out in the literature.

  6. Integrated transcriptomic and proteomic analysis of the molecular cargo of extracellular vesicles derived from porcine adipose tissue-derived mesenchymal stem cells

    OpenAIRE

    Eirin, Alfonso; Zhu, Xiang-Yang; Puranik, Amrutesh S.; Woollard, John R.; Tang, Hui; Dasari, Surendra; Lerman, Amir; van Wijnen, Andre J.; Lerman, Lilach O.

    2017-01-01

    Background Mesenchymal stromal/stem cell (MSC) transplantation is a promising therapy for tissue regeneration. Extracellular vesicles (EVs) released by MSCs act as their paracrine effectors by delivering proteins and genetic material to recipient cells. To assess how their cargo mediates biological processes that drive their therapeutic effects, we integrated miRNA, mRNA, and protein expression data of EVs from porcine adipose tissue-derived MSCs. Methods Simultaneous expression profiles of m...

  7. Analysis of the variability of human normal urine by 2D-GE reveals a "public" and a "private" proteome.

    Science.gov (United States)

    Molina, Laurence; Salvetat, Nicolas; Ameur, Randa Ben; Peres, Sabine; Sommerer, Nicolas; Jarraya, Fayçal; Ayadi, Hammadi; Molina, Franck; Granier, Claude

    2011-12-10

    The characterization of the normal urinary proteome is steadily progressing and represents a major interest in the assessment of clinical urinary biomarkers. To estimate quantitatively the variability of the normal urinary proteome, urines of 20 healthy people were collected. We first evaluated the impact of the sample conservation temperature on urine proteome integrity. Keeping the urine sample at RT or at +4°C until storage at -80°C seems the best way for long-term storage of samples for 2D-GE analysis. The quantitative variability of the normal urinary proteome was estimated on the 20 urines mapped by 2D-GE. The occurrence of the 910 identified spots was analysed throughout the gels and represented in a virtual 2D gel. Sixteen percent of the spots were found to occur in all samples and 23% occurred in at least 90% of urines. About 13% of the protein spots were present only in 10% or less of the samples, thus representing the most variable part of the normal urinary proteome. Twenty proteins corresponding to a fraction of the fully conserved spots were identified by mass spectrometry. In conclusion, a "public" urinary proteome, common to healthy individuals, seems to coexist with a "private" urinary proteome, which is more specific to each individual. Copyright © 2011 Elsevier B.V. All rights reserved.

  8. Transcriptome and proteomic analysis of mango (Mangifera indica Linn) fruits.

    Science.gov (United States)

    Wu, Hong-xia; Jia, Hui-min; Ma, Xiao-wei; Wang, Song-biao; Yao, Quan-sheng; Xu, Wen-tian; Zhou, Yi-gang; Gao, Zhong-shan; Zhan, Ru-lin

    2014-06-13

    Here we used Illumina RNA-seq technology for transcriptome sequencing of a mixed fruit sample from 'Zill' mango (Mangifera indica Linn) fruit pericarp and pulp during the development and ripening stages. RNA-seq generated 68,419,722 sequence reads that were assembled into 54,207 transcripts with a mean length of 858bp, including 26,413 clusters and 27,794 singletons. A total of 42,515(78.43%) transcripts were annotated using public protein databases, with a cut-off E-value above 10(-5), of which 35,198 and 14,619 transcripts were assigned to gene ontology terms and clusters of orthologous groups respectively. Functional annotation against the Kyoto Encyclopedia of Genes and Genomes database identified 23,741(43.79%) transcripts which were mapped to 128 pathways. These pathways revealed many previously unknown transcripts. We also applied mass spectrometry-based transcriptome data to characterize the proteome of ripe fruit. LC-MS/MS analysis of the mango fruit proteome was using tandem mass spectrometry (MS/MS) in an LTQ Orbitrap Velos (Thermo) coupled online to the HPLC. This approach enabled the identification of 7536 peptides that matched 2754 proteins. Our study provides a comprehensive sequence for a systemic view of transcriptome during mango fruit development and the most comprehensive fruit proteome to date, which are useful for further genomics research and proteomic studies. Our study provides a comprehensive sequence for a systemic view of both the transcriptome and proteome of mango fruit, and a valuable reference for further research on gene expression and protein identification. This article is part of a Special Issue entitled: Proteomics of non-model organisms. Copyright © 2014 Elsevier B.V. All rights reserved.

  9. A community proposal to integrate proteomics activities in ELIXIR

    Czech Academy of Sciences Publication Activity Database

    Vizcaíno, J. A.; Walzer, M.; Jiménez, R. C.; Bittremieux, W.; Bouyssié, D.; Carapito, C.; Corrales, F.; Ferro, M.; Heck, A. J. R.; Horvatovich, P.; Hubálek, Martin; Lane, L.; Laukens, K.; Levander, F.; Lisacek, F.; Novák, Petr; Palmblad, M.; Piovesan, D.; Pühler, A.; Schwämmle, V.; Valkenborg, D.; van Rijswijk, M.; Vondrášek, Jiří; Eisenacher, M.; Martens, L.; Kohlbacher, O.

    2017-01-01

    Roč. 6, Jun 13 (2017), č. článku 875. ISSN 2046-1402 Institutional support: RVO:61388963 ; RVO:61388971 Keywords : proteomics * bioinformatics * ELIXIR Subject RIV: CE - Biochemistry OBOR OECD: Biochemistry and molecular biology https://f1000research.com/articles/6-875/v1

  10. Proteomic analysis reveals the diversity and complexity of membrane proteins in chickpea (Cicer arietinum L.

    Directory of Open Access Journals (Sweden)

    Jaiswal Dinesh Kumar

    2012-10-01

    Full Text Available Abstract Background Compartmentalization is a unique feature of eukaryotes that helps in maintaining cellular homeostasis not only in intra- and inter-organellar context, but also between the cells and the external environment. Plant cells are highly compartmentalized with a complex metabolic network governing various cellular events. The membranes are the most important constituents in such compartmentalization, and membrane-associated proteins play diverse roles in many cellular processes besides being part of integral component of many signaling cascades. Results To obtain valuable insight into the dynamic repertoire of membrane proteins, we have developed a proteome reference map of a grain legume, chickpea, using two-dimensional gel electrophoresis. MALDI-TOF/TOF and LC-ESI-MS/MS analysis led to the identification of 91 proteins involved in a variety of cellular functions viz., bioenergy, stress-responsive and signal transduction, metabolism, protein synthesis and degradation, among others. Significantly, 70% of the identified proteins are putative integral membrane proteins, possessing transmembrane domains. Conclusions The proteomic analysis revealed many resident integral membrane proteins as well as membrane-associated proteins including those not reported earlier. To our knowledge, this is the first report of membrane proteome from aerial tissues of a crop plant. The findings may provide a better understanding of the biochemical machinery of the plant membranes at the molecular level that might help in functional genomics studies of different developmental pathways and stress-responses.

  11. Translational plant proteomics: a perspective.

    Science.gov (United States)

    Agrawal, Ganesh Kumar; Pedreschi, Romina; Barkla, Bronwyn J; Bindschedler, Laurence Veronique; Cramer, Rainer; Sarkar, Abhijit; Renaut, Jenny; Job, Dominique; Rakwal, Randeep

    2012-08-03

    Translational proteomics is an emerging sub-discipline of the proteomics field in the biological sciences. Translational plant proteomics aims to integrate knowledge from basic sciences to translate it into field applications to solve issues related but not limited to the recreational and economic values of plants, food security and safety, and energy sustainability. In this review, we highlight the substantial progress reached in plant proteomics during the past decade which has paved the way for translational plant proteomics. Increasing proteomics knowledge in plants is not limited to model and non-model plants, proteogenomics, crop improvement, and food analysis, safety, and nutrition but to many more potential applications. Given the wealth of information generated and to some extent applied, there is the need for more efficient and broader channels to freely disseminate the information to the scientific community. This article is part of a Special Issue entitled: Translational Proteomics. Copyright © 2012 Elsevier B.V. All rights reserved.

  12. Next-generation technologies for spatial proteomics: Integrating ultra-high speed MALDI-TOF and high mass resolution MALDI FTICR imaging mass spectrometry for protein analysis.

    Science.gov (United States)

    Spraggins, Jeffrey M; Rizzo, David G; Moore, Jessica L; Noto, Michael J; Skaar, Eric P; Caprioli, Richard M

    2016-06-01

    MALDI imaging mass spectrometry is a powerful analytical tool enabling the visualization of biomolecules in tissue. However, there are unique challenges associated with protein imaging experiments including the need for higher spatial resolution capabilities, improved image acquisition rates, and better molecular specificity. Here we demonstrate the capabilities of ultra-high speed MALDI-TOF and high mass resolution MALDI FTICR IMS platforms as they relate to these challenges. High spatial resolution MALDI-TOF protein images of rat brain tissue and cystic fibrosis lung tissue were acquired at image acquisition rates >25 pixels/s. Structures as small as 50 μm were spatially resolved and proteins associated with host immune response were observed in cystic fibrosis lung tissue. Ultra-high speed MALDI-TOF enables unique applications including megapixel molecular imaging as demonstrated for lipid analysis of cystic fibrosis lung tissue. Additionally, imaging experiments using MALDI FTICR IMS were shown to produce data with high mass accuracy (z 5000) for proteins up to ∼20 kDa. Analysis of clear cell renal cell carcinoma using MALDI FTICR IMS identified specific proteins localized to healthy tissue regions, within the tumor, and also in areas of increased vascularization around the tumor. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  13. Analysis of biostimulated microbial communities from two field experiments reveals temporal and spatial differences in proteome profiles

    Energy Technology Data Exchange (ETDEWEB)

    Callister, S.J.; Wilkins, M.J.; Nicora, C.D.; Williams, K.H.; Banfield, J.F.; VerBerkmoes, N.C.; Hettich, R.L.; NGuessan, A.L.; Mouser, P.J.; Elifantz, H.; Smith, R.D.; Lovley, D.R.; Lipton, M.S.; Long, P.E.

    2010-07-15

    Stimulated by an acetate-amendment field experiment conducted in 2007, anaerobic microbial populations in the aquifer at the Rifle Integrated Field Research Challenge site in Colorado reduced mobile U(VI) to insoluble U(IV). During this experiment, planktonic biomass was sampled at various time points to quantitatively evaluate proteomes. In 2008, an acetate-amended field experiment was again conducted in a similar manner to the 2007 experiment. As there was no comprehensive metagenome sequence available for use in proteomics analysis, we systematically evaluated 12 different organism genome sequences to generate sets of aggregate genomes, or “pseudo-metagenomes”, for supplying relative quantitative peptide and protein identifications. Proteomics results support previous observations of the dominance of Geobacteraceae during biostimulation using acetate as sole electron donor, and revealed a shift from an early stage of iron reduction to a late stage of iron reduction. Additionally, a shift from iron reduction to sulfate reduction was indicated by changes in the contribution of proteome information contributed by different organism genome sequences within the aggregate set. In addition, the comparison of proteome measurements made between the 2007 field experiment and 2008 field experiment revealed differences in proteome profiles. These differences may be the result of alterations in abundance and population structure within the planktonic biomass samples collected for analysis.

  14. Partitioning the proteome: phase separation for targeted analysis of membrane proteins in human post-mortem brain.

    Directory of Open Access Journals (Sweden)

    Jane A English

    Full Text Available Neuroproteomics is a powerful platform for targeted and hypothesis driven research, providing comprehensive insights into cellular and sub-cellular disease states, Gene × Environmental effects, and cellular response to medication effects in human, animal, and cell culture models. Analysis of sub-proteomes is becoming increasingly important in clinical proteomics, enriching for otherwise undetectable proteins that are possible markers for disease. Membrane proteins are one such sub-proteome class that merit in-depth targeted analysis, particularly in psychiatric disorders. As membrane proteins are notoriously difficult to analyse using traditional proteomics methods, we evaluate a paradigm to enrich for and study membrane proteins from human post-mortem brain tissue. This is the first study to extensively characterise the integral trans-membrane spanning proteins present in human brain. Using Triton X-114 phase separation and LC-MS/MS analysis, we enriched for and identified 494 membrane proteins, with 194 trans-membrane helices present, ranging from 1 to 21 helices per protein. Isolated proteins included glutamate receptors, G proteins, voltage gated and calcium channels, synaptic proteins, and myelin proteins, all of which warrant quantitative proteomic investigation in psychiatric and neurological disorders. Overall, our sub-proteome analysis reduced sample complexity and enriched for integral membrane proteins by 2.3 fold, thus allowing for more manageable, reproducible, and targeted proteomics in case vs. control biomarker studies. This study provides a valuable reference for future neuroproteomic investigations of membrane proteins, and validates the use Triton X-114 detergent phase extraction on human post mortem brain.

  15. Differential proteome analysis of chikungunya virus infection on host cells.

    Directory of Open Access Journals (Sweden)

    Christina Li-Ping Thio

    Full Text Available BACKGROUND: Chikungunya virus (CHIKV is an emerging mosquito-borne alphavirus that has caused multiple unprecedented and re-emerging outbreaks in both tropical and temperate countries. Despite ongoing research efforts, the underlying factors involved in facilitating CHIKV replication during early infection remains ill-characterized. The present study serves to identify host proteins modulated in response to early CHIKV infection using a proteomics approach. METHODOLOGY AND PRINCIPAL FINDINGS: The whole cell proteome profiles of CHIKV-infected and mock control WRL-68 cells were compared and analyzed using two-dimensional gel electrophoresis (2-DGE. Fifty-three spots were found to be differentially modulated and 50 were successfully identified by MALDI-TOF/TOF. Eight were significantly up-regulated and 42 were down-regulated. The mRNA expressions of 15 genes were also found to correlate with the corresponding protein expression. STRING network analysis identified several biological processes to be affected, including mRNA processing, translation, energy production and cellular metabolism, ubiquitin-proteasome pathway (UPP and cell cycle regulation. CONCLUSION/SIGNIFICANCE: This study constitutes a first attempt to investigate alteration of the host cellular proteome during early CHIKV infection. Our proteomics data showed that during early infection, CHIKV affected the expression of proteins that are involved in mRNA processing, host metabolic machinery, UPP, and cyclin-dependent kinase 1 (CDK1 regulation (in favour of virus survival, replication and transmission. While results from this study complement the proteomics results obtained from previous late host response studies, functional characterization of these proteins is warranted to reinforce our understanding of their roles during early CHIKV infection in humans.

  16. Data from quantitative label free proteomics analysis of rat spleen.

    Science.gov (United States)

    Dudekula, Khadar; Le Bihan, Thierry

    2016-09-01

    The dataset presented in this work has been obtained using a label-free quantitative proteomic analysis of rat spleen. A robust method for extraction of proteins from rat spleen tissue and LC-MS-MS analysis was developed using a urea and SDS-based buffer. Different fractionation methods were compared. A total of 3484 different proteins were identified from the pool of all experiments run in this study (a total of 2460 proteins with at least two peptides). A total of 1822 proteins were identified from nine non-fractionated pulse gels, 2288 proteins and 2864 proteins were identified by SDS-PAGE fractionation into three and five fractions respectively. The proteomics data are deposited in ProteomeXchange Consortium via PRIDE PXD003520, Progenesis and Maxquant output are presented in the supported information. The generated list of proteins under different regimes of fractionation allow assessing the nature of the identified proteins; variability in the quantitative analysis associated with the different sampling strategy and allow defining a proper number of replicates for future quantitative analysis.

  17. Statistics in experimental design, preprocessing, and analysis of proteomics data.

    Science.gov (United States)

    Jung, Klaus

    2011-01-01

    High-throughput experiments in proteomics, such as 2-dimensional gel electrophoresis (2-DE) and mass spectrometry (MS), yield usually high-dimensional data sets of expression values for hundreds or thousands of proteins which are, however, observed on only a relatively small number of biological samples. Statistical methods for the planning and analysis of experiments are important to avoid false conclusions and to receive tenable results. In this chapter, the most frequent experimental designs for proteomics experiments are illustrated. In particular, focus is put on studies for the detection of differentially regulated proteins. Furthermore, issues of sample size planning, statistical analysis of expression levels as well as methods for data preprocessing are covered.

  18. Mechanism of cisplatin proximal tubule toxicity revealed by integrating transcriptomics, proteomics, metabolomics and biokinetics

    NARCIS (Netherlands)

    Wilmes, Anja; Bielow, Chris; Ranninger, Christina; Bellwon, Patricia; Aschauer, Lydia; Limonciel, Alice; Chassaigne, Hubert; Kristl, Theresa; Aiche, Stephan; Huber, Christian G; Guillou, Claude; Hewitt, Philipp; Leonard, Martin O; Dekant, Wolfgang; Bois, Frederic Y; Jennings, Paul

    2015-01-01

    Cisplatin is one of the most widely used chemotherapeutic agents for the treatment of solid tumours. The major dose-limiting factor is nephrotoxicity, in particular in the proximal tubule. Here, we use an integrated omics approach, including transcriptomics, proteomics and metabolomics coupled to

  19. A Combined Metabolomic and Proteomic Analysis of Gestational Diabetes Mellitus

    OpenAIRE

    Hajduk, Joanna; Klupczynska, Agnieszka; Dereziński, Paweł; Matysiak, Jan; Kokot, Piotr; Nowak, Dorota; Gajęcka, Marzena; Nowak-Markwitz, Ewa; Kokot, Zenon

    2015-01-01

    The aim of this pilot study was to apply a novel combined metabolomic and proteomic approach in analysis of gestational diabetes mellitus. The investigation was performed with plasma samples derived from pregnant women with diagnosed gestational diabetes mellitus (n = 18) and a matched control group (n = 13). The mass spectrometry-based analyses allowed to determine 42 free amino acids and low molecular-weight peptide profiles. Different expressions of several peptides and altered amino acid ...

  20. Characteristics of the tomato chromoplast revealed by proteomic analysis

    OpenAIRE

    Barsan, Cristina; Sanchez-Bel, Paloma; Rombaldi, César Valmor; Egea, Isabel; Rossignol, Michel; Kuntz, Marcel; Zouine, Mohamed; Latché, Alain; Bouzayen, Mondher; Pech, Jean-Claude

    2010-01-01

    Chromoplasts are non-photosynthetic specialized plastids that are important in ripening tomato fruit (Solanum lycopersicum) since, among other functions, they are the site of accumulation of coloured compounds. Analysis of the proteome of red fruit chromoplasts revealed the presence of 988 proteins corresponding to 802 Arabidopsis unigenes, among which 209 had not been listed so far in plastidial databanks. These data revealed several features of the chromoplast. Proteins of lipid metabolism ...

  1. YPED: an integrated bioinformatics suite and database for mass spectrometry-based proteomics research.

    Science.gov (United States)

    Colangelo, Christopher M; Shifman, Mark; Cheung, Kei-Hoi; Stone, Kathryn L; Carriero, Nicholas J; Gulcicek, Erol E; Lam, TuKiet T; Wu, Terence; Bjornson, Robert D; Bruce, Can; Nairn, Angus C; Rinehart, Jesse; Miller, Perry L; Williams, Kenneth R

    2015-02-01

    We report a significantly-enhanced bioinformatics suite and database for proteomics research called Yale Protein Expression Database (YPED) that is used by investigators at more than 300 institutions worldwide. YPED meets the data management, archival, and analysis needs of a high-throughput mass spectrometry-based proteomics research ranging from a single laboratory, group of laboratories within and beyond an institution, to the entire proteomics community. The current version is a significant improvement over the first version in that it contains new modules for liquid chromatography-tandem mass spectrometry (LC-MS/MS) database search results, label and label-free quantitative proteomic analysis, and several scoring outputs for phosphopeptide site localization. In addition, we have added both peptide and protein comparative analysis tools to enable pairwise analysis of distinct peptides/proteins in each sample and of overlapping peptides/proteins between all samples in multiple datasets. We have also implemented a targeted proteomics module for automated multiple reaction monitoring (MRM)/selective reaction monitoring (SRM) assay development. We have linked YPED's database search results and both label-based and label-free fold-change analysis to the Skyline Panorama repository for online spectra visualization. In addition, we have built enhanced functionality to curate peptide identifications into an MS/MS peptide spectral library for all of our protein database search identification results. Copyright © 2015 The Authors. Production and hosting by Elsevier Ltd.. All rights reserved.

  2. Plastid Proteomic Analysis in Tomato Fruit Development.

    Directory of Open Access Journals (Sweden)

    Miho Suzuki

    Full Text Available To better understand the mechanism of plastid differentiation from chloroplast to chromoplast, we examined proteome and plastid changes over four distinct developmental stages of 'Micro-Tom' fruit. Additionally, to discover more about the relationship between fruit color and plastid differentiation, we also analyzed and compared 'Micro-Tom' results with those from two other varieties, 'Black' and 'White Beauty'. We confirmed that proteins related to photosynthesis remain through the orange maturity stage of 'Micro-Tom', and also learned that thylakoids no longer exist at this stage. These results suggest that at a minimum there are changes in plastid morphology occurring before all related proteins change. We also compared 'Micro-Tom' fruits with 'Black' and 'White Beauty' using two-dimensional gel electrophoresis. We found a decrease of CHRC (plastid-lipid-associated protein and HrBP1 (harpin binding protein-1 in the 'Black' and 'White Beauty' varieties. CHRC is involved in carotenoid accumulation and stabilization. HrBP1 in Arabidopsis has a sequence similar to proteins in the PAP/fibrillin family. These proteins have characteristics and functions similar to lipocalin, an example of which is the transport of hydrophobic molecules. We detected spots of TIL (temperature-induced lipocalin in 2D-PAGE results, however the number of spots and their isoelectric points differed between 'Micro-Tom' and 'Black'/'White Beauty'. Lipocalin has various functions including those related to environmental stress response, apoptosis induction, membrane formation and fixation, regulation of immune response, cell growth, and metabolism adjustment. Lipocalin related proteins such as TIL and HrBP1 could be related to the accumulation of carotenoids, fruit color and the differentiation of chromoplast.

  3. Systems biology integration of proteomic data in rodent models of depression reveals involvement of the immune response and glutamatergic signaling.

    Science.gov (United States)

    Carboni, Lucia; Nguyen, Thanh-Phuong; Caberlotto, Laura

    2016-12-01

    The pathophysiological basis of major depression is incompletely understood. Recently, numerous proteomic studies have been performed in rodent models of depression to investigate the molecular underpinnings of depressive-like behaviours with an unbiased approach. The objective of the study is to integrate the results of these proteomic studies in depression models to shed light on the most relevant molecular pathways involved in the disease. Network analysis is performed integrating preexisting proteomic data from rodent models of depression. The IntAct mouse and the HRPD are used as reference protein-protein interaction databases. The functionality analyses of the networks are then performed by testing overrepresented GO biological process terms and pathways. Functional enrichment analyses of the networks revealed an association with molecular processes related to depression in humans, such as those involved in the immune response. Pathways impacted by clinically effective antidepressants are modulated, including glutamatergic signaling and neurotrophic responses. Moreover, dysregulations of proteins regulating energy metabolism and circadian rhythms are implicated. The comparison with protein pathways modulated in depressive patients revealed significant overlapping. This systems biology study supports the notion that animal models can contribute to the research into the biology and therapeutics of depression. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  4. Proteomics analysis of ram sperm by heavy ion radiation

    International Nuclear Information System (INIS)

    He Yuxuan; Li Hongyan; Zhang Hong

    2013-01-01

    The objective of this study was to investigate the proteome changes induced by heavy ion radiation using irradiated ram sperm by a two-dimensional electrophoresis (2-DE) analysis. The 2D gels were stained with Coomassie Brilliant Blue. Differentially expressed proteins were detected by PDQuest 8.0 software and subjected to ion trap mass spectrometer equipped with a surveyor HPLC system, and differential protein spots were identified. Results showed there are five differential protein spots in irradiated sperm gels, four up-regulated protein spots and one spot missed. The differentially expressed protein spots were identified to be two up-regulated proteins including enolase, and enolase 1. It was concluded there was proteome changes induced by heavy ion radiation in ram sperm, which may be useful to clarify the physiology state of ram sperm in heavy ion radiation and provide a theoretical basis for radiation ram breeding. (authors)

  5. Proteomics-driven analysis of ovine whey colostrum.

    Directory of Open Access Journals (Sweden)

    Domenica Scumaci

    Full Text Available The aim of this study was to shed light in to the complexity of the ovine colostrum proteome, with a specific focus on the low abundance proteins. The ovine colostrum is characterized by a few dominating proteins, as the immunoglobulins, but it also contains less represented protein species, equally important for the correct development of neonates. Ovine colostrum, collected immediately after lambing, was separated by 1D SDS-PAGE. Proteins bands were digested with trypsin and the resulting peptides were analyzed by LC-MS/MS. On the basis of the Swiss-Prot database, a total of 343 unique proteins were identified. To our knowledge, this study represents the most comprehensive analysis of ovine colostrum proteome.

  6. Integration of cardiac proteome biology and medicine by a specialized knowledgebase.

    Science.gov (United States)

    Zong, Nobel C; Li, Haomin; Li, Hua; Lam, Maggie P Y; Jimenez, Rafael C; Kim, Christina S; Deng, Ning; Kim, Allen K; Choi, Jeong Ho; Zelaya, Ivette; Liem, David; Meyer, David; Odeberg, Jacob; Fang, Caiyun; Lu, Hao-Jie; Xu, Tao; Weiss, James; Duan, Huilong; Uhlen, Mathias; Yates, John R; Apweiler, Rolf; Ge, Junbo; Hermjakob, Henning; Ping, Peipei

    2013-10-12

    Omics sciences enable a systems-level perspective in characterizing cardiovascular biology. Integration of diverse proteomics data via a computational strategy will catalyze the assembly of contextualized knowledge, foster discoveries through multidisciplinary investigations, and minimize unnecessary redundancy in research efforts. The goal of this project is to develop a consolidated cardiac proteome knowledgebase with novel bioinformatics pipeline and Web portals, thereby serving as a new resource to advance cardiovascular biology and medicine. We created Cardiac Organellar Protein Atlas Knowledgebase (COPaKB; www.HeartProteome.org), a centralized platform of high-quality cardiac proteomic data, bioinformatics tools, and relevant cardiovascular phenotypes. Currently, COPaKB features 8 organellar modules, comprising 4203 LC-MS/MS experiments from human, mouse, drosophila, and Caenorhabditis elegans, as well as expression images of 10,924 proteins in human myocardium. In addition, the Java-coded bioinformatics tools provided by COPaKB enable cardiovascular investigators in all disciplines to retrieve and analyze pertinent organellar protein properties of interest. COPaKB provides an innovative and interactive resource that connects research interests with the new biological discoveries in protein sciences. With an array of intuitive tools in this unified Web server, nonproteomics investigators can conveniently collaborate with proteomics specialists to dissect the molecular signatures of cardiovascular phenotypes.

  7. An integrated native mass spectrometry and top-down proteomics method that connects sequence to structure and function of macromolecular complexes

    Science.gov (United States)

    Li, Huilin; Nguyen, Hong Hanh; Ogorzalek Loo, Rachel R.; Campuzano, Iain D. G.; Loo, Joseph A.

    2018-02-01

    Mass spectrometry (MS) has become a crucial technique for the analysis of protein complexes. Native MS has traditionally examined protein subunit arrangements, while proteomics MS has focused on sequence identification. These two techniques are usually performed separately without taking advantage of the synergies between them. Here we describe the development of an integrated native MS and top-down proteomics method using Fourier-transform ion cyclotron resonance (FTICR) to analyse macromolecular protein complexes in a single experiment. We address previous concerns of employing FTICR MS to measure large macromolecular complexes by demonstrating the detection of complexes up to 1.8 MDa, and we demonstrate the efficacy of this technique for direct acquirement of sequence to higher-order structural information with several large complexes. We then summarize the unique functionalities of different activation/dissociation techniques. The platform expands the ability of MS to integrate proteomics and structural biology to provide insights into protein structure, function and regulation.

  8. Comparative proteomic analysis in Miscanthus sinensis exposed to antimony stress

    International Nuclear Information System (INIS)

    Xue, Liang; Ren, Huadong; Li, Sheng; Gao, Ming; Shi, Shengqing; Chang, Ermei; Wei, Yuan; Yao, Xiaohua; Jiang, Zeping; Liu, Jianfeng

    2015-01-01

    To explore the molecular basis of Sb tolerance mechanism in plant, a comparative proteomic analysis of both roots and leaves in Miscanthus sinensis has been conducted in combination with physiological and biochemical analyses. M. sinensis seedlings were exposed to different doses of Sb, and both roots and leaves were collected after 3 days of treatment. Two-dimensional gel electrophoresis (2-DE) and image analyses found that 29 protein spots showed 1.5-fold change in abundance in leaves and 19 spots in roots, of which 31 were identified by MALDI-TOF-MS and MALDI-TOF-TOF-MS. Proteins involved in antioxidant defense and stress response generally increased their expression all over the Sb treatments. In addition, proteins relative to transcription, signal transduction, energy metabolism and cell division and cell structure showed a variable expression pattern over Sb concentrations. Overall these findings provide new insights into the probable survival mechanisms by which M. sinensis could be adapting to Sb phytotoxicity. - Highlights: • Proteomics in Miscanthus sinensis leaves and roots exposed to Sb stress were studied. • There were 31 spots that were identified by mass spectrometry. • Most of these proteins were involved in antioxidant defense and stress response. • Our findings provide new insights into the tolerant mechanisms to Sb stress. - Miscanthus sinensis proteomic analysis under Sb stress reveals probable molecular mechanisms on Sb detoxification

  9. Proteomic analysis of urine in rats chronically exposed to fluoride.

    Science.gov (United States)

    Kobayashi, Claudia Ayumi Nakai; Leite, Aline de Lima; da Silva, Thelma Lopes; dos Santos, Lucilene Delazari; Nogueira, Fábio César Sousa; Santos, Keity Souza; de Oliveira, Rodrigo Cardoso; Palma, Mario Sérgio; Domont, Gilberto Barbosa; Buzalaf, Marília Afonso Rabelo

    2011-01-01

    Urine is an ideal source of materials to search for potential disease-related biomarkers as it is produced by the affected tissues and can be easily obtained by noninvasive methods. 2-DE-based proteomic approach was used to better understand the molecular mechanisms of injury induced by fluoride (F(-)) and define potential biomarkers of dental fluorosis. Three groups of weanling male Wistar rats were treated with drinking water containing 0 (control), 5, or 50 ppm F(-) for 60 days (n = 15/group). During the experimental period, the animals were kept individually in metabolic cages, to analyze the water and food consumption, as well as fecal and urinary F(-) excretion. Urinary proteome profiles were examined using 2-DE and Colloidal Coomassie Brilliant Blue staining. A dose-response regarding F(-) intake and excretion was detected. Quantitative intensity analysis revealed 8, 11, and 8 significantly altered proteins between control vs. 5 ppm F(-), control vs. 50 ppm F(-) and 5 ppm F(-) vs. 50 ppm F(-) groups, respectively. Two proteins regulated by androgens (androgen-regulated 20-KDa protein and α-2μ-globulin) and one related to detoxification (aflatoxin-B1-aldehyde-reductase) were identified by MALDI-TOF-TOF MS/MS. Thus, proteomic analysis can help to better understand the mechanisms underlying F(-) toxicity, even in low doses. Copyright © 2010 Wiley Periodicals, Inc.

  10. EBprot: Statistical analysis of labeling-based quantitative proteomics data.

    Science.gov (United States)

    Koh, Hiromi W L; Swa, Hannah L F; Fermin, Damian; Ler, Siok Ghee; Gunaratne, Jayantha; Choi, Hyungwon

    2015-08-01

    Labeling-based proteomics is a powerful method for detection of differentially expressed proteins (DEPs). The current data analysis platform typically relies on protein-level ratios, which is obtained by summarizing peptide-level ratios for each protein. In shotgun proteomics, however, some proteins are quantified with more peptides than others, and this reproducibility information is not incorporated into the differential expression (DE) analysis. Here, we propose a novel probabilistic framework EBprot that directly models the peptide-protein hierarchy and rewards the proteins with reproducible evidence of DE over multiple peptides. To evaluate its performance with known DE states, we conducted a simulation study to show that the peptide-level analysis of EBprot provides better receiver-operating characteristic and more accurate estimation of the false discovery rates than the methods based on protein-level ratios. We also demonstrate superior classification performance of peptide-level EBprot analysis in a spike-in dataset. To illustrate the wide applicability of EBprot in different experimental designs, we applied EBprot to a dataset for lung cancer subtype analysis with biological replicates and another dataset for time course phosphoproteome analysis of EGF-stimulated HeLa cells with multiplexed labeling. Through these examples, we show that the peptide-level analysis of EBprot is a robust alternative to the existing statistical methods for the DE analysis of labeling-based quantitative datasets. The software suite is freely available on the Sourceforge website http://ebprot.sourceforge.net/. All MS data have been deposited in the ProteomeXchange with identifier PXD001426 (http://proteomecentral.proteomexchange.org/dataset/PXD001426/). © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  11. PIQMIe: a web server for semi-quantitative proteomics data management and analysis.

    Science.gov (United States)

    Kuzniar, Arnold; Kanaar, Roland

    2014-07-01

    We present the Proteomics Identifications and Quantitations Data Management and Integration Service or PIQMIe that aids in reliable and scalable data management, analysis and visualization of semi-quantitative mass spectrometry based proteomics experiments. PIQMIe readily integrates peptide and (non-redundant) protein identifications and quantitations from multiple experiments with additional biological information on the protein entries, and makes the linked data available in the form of a light-weight relational database, which enables dedicated data analyses (e.g. in R) and user-driven queries. Using the web interface, users are presented with a concise summary of their proteomics experiments in numerical and graphical forms, as well as with a searchable protein grid and interactive visualization tools to aid in the rapid assessment of the experiments and in the identification of proteins of interest. The web server not only provides data access through a web interface but also supports programmatic access through RESTful web service. The web server is available at http://piqmie.semiqprot-emc.cloudlet.sara.nl or http://www.bioinformatics.nl/piqmie. This website is free and open to all users and there is no login requirement. © The Author(s) 2014. Published by Oxford University Press on behalf of Nucleic Acids Research.

  12. A linear programming computational framework integrates phosphor-proteomics and prior knowledge to predict drug efficacy.

    Science.gov (United States)

    Ji, Zhiwei; Wang, Bing; Yan, Ke; Dong, Ligang; Meng, Guanmin; Shi, Lei

    2017-12-21

    In recent years, the integration of 'omics' technologies, high performance computation, and mathematical modeling of biological processes marks that the systems biology has started to fundamentally impact the way of approaching drug discovery. The LINCS public data warehouse provides detailed information about cell responses with various genetic and environmental stressors. It can be greatly helpful in developing new drugs and therapeutics, as well as improving the situations of lacking effective drugs, drug resistance and relapse in cancer therapies, etc. In this study, we developed a Ternary status based Integer Linear Programming (TILP) method to infer cell-specific signaling pathway network and predict compounds' treatment efficacy. The novelty of our study is that phosphor-proteomic data and prior knowledge are combined for modeling and optimizing the signaling network. To test the power of our approach, a generic pathway network was constructed for a human breast cancer cell line MCF7; and the TILP model was used to infer MCF7-specific pathways with a set of phosphor-proteomic data collected from ten representative small molecule chemical compounds (most of them were studied in breast cancer treatment). Cross-validation indicated that the MCF7-specific pathway network inferred by TILP were reliable predicting a compound's efficacy. Finally, we applied TILP to re-optimize the inferred cell-specific pathways and predict the outcomes of five small compounds (carmustine, doxorubicin, GW-8510, daunorubicin, and verapamil), which were rarely used in clinic for breast cancer. In the simulation, the proposed approach facilitates us to identify a compound's treatment efficacy qualitatively and quantitatively, and the cross validation analysis indicated good accuracy in predicting effects of five compounds. In summary, the TILP model is useful for discovering new drugs for clinic use, and also elucidating the potential mechanisms of a compound to targets.

  13. Comprehensive Proteomic Analysis of Human Milk-derived Extracellular Vesicles Unveils a Novel Functional Proteome Distinct from Other Milk Components*

    Science.gov (United States)

    van Herwijnen, Martijn J.C.; Zonneveld, Marijke I.; Goerdayal, Soenita; Nolte – 't Hoen, Esther N.M.; Garssen, Johan; Stahl, Bernd; Maarten Altelaar, A.F.; Redegeld, Frank A.; Wauben, Marca H.M.

    2016-01-01

    Breast milk contains several macromolecular components with distinctive functions, whereby milk fat globules and casein micelles mainly provide nutrition to the newborn, and whey contains molecules that can stimulate the newborn's developing immune system and gastrointestinal tract. Although extracellular vesicles (EV) have been identified in breast milk, their physiological function and composition has not been addressed in detail. EV are submicron sized vehicles released by cells for intercellular communication via selectively incorporated lipids, nucleic acids, and proteins. Because of the difficulty in separating EV from other milk components, an in-depth analysis of the proteome of human milk-derived EV is lacking. In this study, an extensive LC-MS/MS proteomic analysis was performed of EV that had been purified from breast milk of seven individual donors using a recently established, optimized density-gradient-based EV isolation protocol. A total of 1963 proteins were identified in milk-derived EV, including EV-associated proteins like CD9, Annexin A5, and Flotillin-1, with a remarkable overlap between the different donors. Interestingly, 198 of the identified proteins are not present in the human EV database Vesiclepedia, indicating that milk-derived EV harbor proteins not yet identified in EV of different origin. Similarly, the proteome of milk-derived EV was compared with that of other milk components. For this, data from 38 published milk proteomic studies were combined in order to construct the total milk proteome, which consists of 2698 unique proteins. Remarkably, 633 proteins identified in milk-derived EV have not yet been identified in human milk to date. Interestingly, these novel proteins include proteins involved in regulation of cell growth and controlling inflammatory signaling pathways, suggesting that milk-derived EVs could support the newborn's developing gastrointestinal tract and immune system. Overall, this study provides an expansion of

  14. A Systematic Analysis of a Deep Mouse Epididymal Sperm Proteome

    Energy Technology Data Exchange (ETDEWEB)

    Chauvin, Theodore; Xie, Fang; Liu, Tao; Nicora, Carrie D.; Yang, Feng; Camp, David G.; Smith, Richard D.; Roberts, Kenneth P.

    2012-12-21

    Spermatozoa are highly specialized cells that, when mature, are capable of navigating the female reproductive tract and fertilizing an oocyte. The sperm cell is thought to be largely quiescent in terms of transcriptional and translational activity. As a result, once it has left the male reproductive tract, the sperm cell is essentially operating with a static population of proteins. It is therefore theoretically possible to understand the protein networks contained in a sperm cell and to deduce its cellular function capabilities. To this end we have performed a proteomic analysis of mouse sperm isolated from the cauda epididymis and have confidently identified 2,850 proteins, which is the most comprehensive sperm proteome for any species reported to date. These proteins comprise many complete cellular pathways, including those for energy production via glycolysis, β-oxidation and oxidative phosphorylation, protein folding and transport, and cell signaling systems. This proteome should prove a useful tool for assembly and testing of protein networks important for sperm function.

  15. 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.

  16. Proteomic analysis of purified coronavirus infectious bronchitis virus particles

    Directory of Open Access Journals (Sweden)

    Shu Dingming

    2010-06-01

    Full Text Available Abstract Background Infectious bronchitis virus (IBV is the coronavirus of domestic chickens causing major economic losses to the poultry industry. Because of the complexity of the IBV life cycle and the small number of viral structural proteins, important virus-host relationships likely remain to be discovered. Toward this goal, we performed two-dimensional gel electrophoresis fractionation coupled to mass spectrometry identification approaches to perform a comprehensive proteomic analysis of purified IBV particles. Results Apart from the virus-encoded structural proteins, we detected 60 host proteins in the purified virions which can be grouped into several functional categories including intracellular trafficking proteins (20%, molecular chaperone (18%, macromolcular biosynthesis proteins (17%, cytoskeletal proteins (15%, signal transport proteins (15%, protein degradation (8%, chromosome associated proteins (2%, ribosomal proteins (2%, and other function proteins (3%. Interestingly, 21 of the total host proteins have not been reported to be present in virions of other virus families, such as major vault protein, TENP protein, ovalbumin, and scavenger receptor protein. Following identification of the host proteins by proteomic methods, the presence of 4 proteins in the purified IBV preparation was verified by western blotting and immunogold labeling detection. Conclusions The results present the first standard proteomic profile of IBV and may facilitate the understanding of the pathogenic mechanisms.

  17. Mechanism of cisplatin proximal tubule toxicity revealed by integrating transcriptomics, proteomics, metabolomics and biokinetics.

    Science.gov (United States)

    Wilmes, Anja; Bielow, Chris; Ranninger, Christina; Bellwon, Patricia; Aschauer, Lydia; Limonciel, Alice; Chassaigne, Hubert; Kristl, Theresa; Aiche, Stephan; Huber, Christian G; Guillou, Claude; Hewitt, Philipp; Leonard, Martin O; Dekant, Wolfgang; Bois, Frederic; Jennings, Paul

    2015-12-25

    Cisplatin is one of the most widely used chemotherapeutic agents for the treatment of solid tumours. The major dose-limiting factor is nephrotoxicity, in particular in the proximal tubule. Here, we use an integrated omics approach, including transcriptomics, proteomics and metabolomics coupled to biokinetics to identify cell stress response pathways induced by cisplatin. The human renal proximal tubular cell line RPTEC/TERT1 was treated with sub-cytotoxic concentrations of cisplatin (0.5 and 2 μM) in a daily repeat dose treating regime for up to 14 days. Biokinetic analysis showed that cisplatin was taken up from the basolateral compartment, transported to the apical compartment, and accumulated in cells over time. This is in line with basolateral uptake of cisplatin via organic cation transporter 2 and bioactivation via gamma-glutamyl transpeptidase located on the apical side of proximal tubular cells. Cisplatin affected several pathways including, p53 signalling, Nrf2 mediated oxidative stress response, mitochondrial processes, mTOR and AMPK signalling. In addition, we identified novel pathways changed by cisplatin, including eIF2 signalling, actin nucleation via the ARP/WASP complex and regulation of cell polarization. In conclusion, using an integrated omic approach together with biokinetics we have identified both novel and established mechanisms of cisplatin toxicity. Copyright © 2014 Elsevier Ltd. All rights reserved.

  18. Protein cleavage strategies for an improved analysis of the membrane proteome

    Directory of Open Access Journals (Sweden)

    Poetsch Ansgar

    2006-03-01

    Full Text Available Abstract Background Membrane proteins still remain elusive in proteomic studies. This is in part due to the distribution of the amino acids lysine and arginine, which are less frequent in integral membrane proteins and almost absent in transmembrane helices. As these amino acids are cleavage targets for the commonly used protease trypsin, alternative cleavage conditions, which should improve membrane protein analysis, were tested by in silico digestion for the three organisms Saccharomyces cerevisiae, Halobacterium sp. NRC-1, and Corynebacterium glutamicum as hallmarks for eukaryotes, archea and eubacteria. Results For the membrane proteomes from all three analyzed organisms, we identified cleavage conditions that achieve better sequence and proteome coverage than trypsin. Greater improvement was obtained for bacteria than for yeast, which was attributed to differences in protein size and GRAVY. It was demonstrated for bacteriorhodopsin that the in silico predictions agree well with the experimental observations. Conclusion For all three examined organisms, it was found that a combination of chymotrypsin and staphylococcal peptidase I gave significantly better results than trypsin. As some of the improved cleavage conditions are not more elaborate than trypsin digestion and have been proven useful in practice, we suppose that the cleavage at both hydrophilic and hydrophobic amino acids should facilitate in general the analysis of membrane proteins for all organisms.

  19. Integration of Proteomics, Bioinformatics, and Systems Biology in Traumatic Brain Injury Biomarker Discovery

    Science.gov (United States)

    Guingab-Cagmat, J.D.; Cagmat, E.B.; Hayes, R.L.; Anagli, J.

    2013-01-01

    Traumatic brain injury (TBI) is a major medical crisis without any FDA-approved pharmacological therapies that have been demonstrated to improve functional outcomes. It has been argued that discovery of disease-relevant biomarkers might help to guide successful clinical trials for TBI. Major advances in mass spectrometry (MS) have revolutionized the field of proteomic biomarker discovery and facilitated the identification of several candidate markers that are being further evaluated for their efficacy as TBI biomarkers. However, several hurdles have to be overcome even during the discovery phase which is only the first step in the long process of biomarker development. The high-throughput nature of MS-based proteomic experiments generates a massive amount of mass spectral data presenting great challenges in downstream interpretation. Currently, different bioinformatics platforms are available for functional analysis and data mining of MS-generated proteomic data. These tools provide a way to convert data sets to biologically interpretable results and functional outcomes. A strategy that has promise in advancing biomarker development involves the triad of proteomics, bioinformatics, and systems biology. In this review, a brief overview of how bioinformatics and systems biology tools analyze, transform, and interpret complex MS datasets into biologically relevant results is discussed. In addition, challenges and limitations of proteomics, bioinformatics, and systems biology in TBI biomarker discovery are presented. A brief survey of researches that utilized these three overlapping disciplines in TBI biomarker discovery is also presented. Finally, examples of TBI biomarkers and their applications are discussed. PMID:23750150

  20. Low cost, scalable proteomics data analysis using Amazon's cloud computing services and open source search algorithms.

    Science.gov (United States)

    Halligan, Brian D; Geiger, Joey F; Vallejos, Andrew K; Greene, Andrew S; Twigger, Simon N

    2009-06-01

    One of the major difficulties for many laboratories setting up proteomics programs has been obtaining and maintaining the computational infrastructure required for the analysis of the large flow of proteomics data. We describe a system that combines distributed cloud computing and open source software to allow laboratories to set up scalable virtual proteomics analysis clusters without the investment in computational hardware or software licensing fees. Additionally, the pricing structure of distributed computing providers, such as Amazon Web Services, allows laboratories or even individuals to have large-scale computational resources at their disposal at a very low cost per run. We provide detailed step-by-step instructions on how to implement the virtual proteomics analysis clusters as well as a list of current available preconfigured Amazon machine images containing the OMSSA and X!Tandem search algorithms and sequence databases on the Medical College of Wisconsin Proteomics Center Web site ( http://proteomics.mcw.edu/vipdac ).

  1. Proteomic analysis of chromate response in Staphylococcus ...

    African Journals Online (AJOL)

    user

    2012-04-18

    Apr 18, 2012 ... analysis was performed to identify proteins involved in chromate stress response of Staphylococcus saprophyticus .... Proteins were visualized by PharosFXTM molecular imager and scanner ..... Molecular dynamics of the.

  2. metabolicMine: an integrated genomics, genetics and proteomics data warehouse for common metabolic disease research.

    Science.gov (United States)

    Lyne, Mike; Smith, Richard N; Lyne, Rachel; Aleksic, Jelena; Hu, Fengyuan; Kalderimis, Alex; Stepan, Radek; Micklem, Gos

    2013-01-01

    Common metabolic and endocrine diseases such as diabetes affect millions of people worldwide and have a major health impact, frequently leading to complications and mortality. In a search for better prevention and treatment, there is ongoing research into the underlying molecular and genetic bases of these complex human diseases, as well as into the links with risk factors such as obesity. Although an increasing number of relevant genomic and proteomic data sets have become available, the quantity and diversity of the data make their efficient exploitation challenging. Here, we present metabolicMine, a data warehouse with a specific focus on the genomics, genetics and proteomics of common metabolic diseases. Developed in collaboration with leading UK metabolic disease groups, metabolicMine integrates data sets from a range of experiments and model organisms alongside tools for exploring them. The current version brings together information covering genes, proteins, orthologues, interactions, gene expression, pathways, ontologies, diseases, genome-wide association studies and single nucleotide polymorphisms. Although the emphasis is on human data, key data sets from mouse and rat are included. These are complemented by interoperation with the RatMine rat genomics database, with a corresponding mouse version under development by the Mouse Genome Informatics (MGI) group. The web interface contains a number of features including keyword search, a library of Search Forms, the QueryBuilder and list analysis tools. This provides researchers with many different ways to analyse, view and flexibly export data. Programming interfaces and automatic code generation in several languages are supported, and many of the features of the web interface are available through web services. The combination of diverse data sets integrated with analysis tools and a powerful query system makes metabolicMine a valuable research resource. The web interface makes it accessible to first

  3. Integrating proteomic and functional genomic technologies in discovery-driven translational breast cancer research

    DEFF Research Database (Denmark)

    Celis, Julio E; Gromov, Pavel; Gromova, Irina

    2003-01-01

    The application of state-of-the-art proteomics and functional genomics technologies to the study of cancer is rapidly shifting toward the analysis of clinically relevant samples derived from patients, as the ultimate aim of translational research is to bring basic discoveries closer to the bedside...

  4. Analysis of the functional aspects and seminal plasma proteomic profile of sperm from smokers.

    Science.gov (United States)

    Antoniassi, Mariana Pereira; Intasqui, Paula; Camargo, Mariana; Zylbersztejn, Daniel Suslik; Carvalho, Valdemir Melechco; Cardozo, Karina H M; Bertolla, Ricardo Pimenta

    2016-11-01

    To evaluate the effect of smoking on sperm functional quality and seminal plasma proteomic profile. Sperm functional tests were performed in 20 non-smoking men with normal semen quality, according to the World Health Organization (2010) and in 20 smoking patients. These included: evaluation of DNA fragmentation by alkaline Comet assay; analysis of mitochondrial activity using DAB staining; and acrosomal integrity evaluation by PNA binding. The remaining semen was centrifuged and seminal plasma was used for proteomic analysis (liquid chromatography-tandem mass spectrometry). The quantified proteins were used for Venn diagram construction in Cytoscape 3.2.1 software, using the PINA4MS plug-in. Then, differentially expressed proteins were used for functional enrichment analysis of Gene Ontology categories, Kyoto Encyclopedia of Genes and Genomes and Reactome, using Cytoscape software and the ClueGO 2.2.0 plug-in. Smokers had a higher percentage of sperm DNA damage (Comet classes III and IV; P analysis, 422 proteins were identified and quantified, of which one protein was absent, 27 proteins were under-represented and six proteins were over-represented in smokers. Functional enrichment analysis showed the enrichment of antigen processing and presentation, positive regulation of prostaglandin secretion involved in immune response, protein kinase A signalling and arachidonic acid secretion, complement activation, regulation of the cytokine-mediated signalling pathway and regulation of acute inflammatory response in the study group (smokers). In conclusion, cigarette smoking was associated with an inflammatory state in the accessory glands and in the testis, as shown by enriched proteomic pathways. This state causes an alteration in sperm functional quality, which is characterized by decreased acrosome integrity and mitochondrial activity, as well as by increased nuclear DNA fragmentation. © 2016 The Authors BJU International © 2016 BJU International Published by John

  5. Analysis of the pumpkin phloem proteome provides insights into angiosperm sieve tube function.

    Science.gov (United States)

    Lin, Ming-Kuem; Lee, Young-Jin; Lough, Tony J; Phinney, Brett S; Lucas, William J

    2009-02-01

    Increasing evidence suggests that proteins present in the angiosperm sieve tube system play an important role in the long distance signaling system of plants. To identify the nature of these putatively non-cell-autonomous proteins, we adopted a large scale proteomics approach to analyze pumpkin phloem exudates. Phloem proteins were fractionated by fast protein liquid chromatography using both anion and cation exchange columns and then either in-solution or in-gel digested following further separation by SDS-PAGE. A total of 345 LC-MS/MS data sets were analyzed using a combination of Mascot and X!Tandem against the NCBI non-redundant green plant database and an extensive Cucurbit maxima expressed sequence tag database. In this analysis, 1,209 different consensi were obtained of which 1,121 could be annotated from GenBank and BLAST search analyses against three plant species, Arabidopsis thaliana, rice (Oryza sativa), and poplar (Populus trichocarpa). Gene ontology (GO) enrichment analyses identified sets of phloem proteins that function in RNA binding, mRNA translation, ubiquitin-mediated proteolysis, and macromolecular and vesicle trafficking. Our findings indicate that protein synthesis and turnover, processes that were thought to be absent in enucleate sieve elements, likely occur within the angiosperm phloem translocation stream. In addition, our GO analysis identified a set of phloem proteins that are associated with the GO term "embryonic development ending in seed dormancy"; this finding raises the intriguing question as to whether the phloem may exert some level of control over seed development. The universal significance of the phloem proteome was highlighted by conservation of the phloem proteome in species as diverse as monocots (rice), eudicots (Arabidopsis and pumpkin), and trees (poplar). These results are discussed from the perspective of the role played by the phloem proteome as an integral component of the whole plant communication system.

  6. Proteomic analysis of GPI-anchored membrane proteins

    DEFF Research Database (Denmark)

    Jung, Hye Ryung; Jensen, Ole Nørregaard

    2006-01-01

    Glycosyl-phosphatidyl-inositol-anchored proteins (GPI-APs) represent a subset of post-translationally modified proteins that are tethered to the outer leaflet of the plasma membrane via a C-terminal GPI anchor. GPI-APs are found in a variety of eukaryote species, from pathogenic microorganisms...... to humans. GPI-APs confer important cellular functions as receptors, enzymes and scaffolding molecules. Specific enzymes and detergent extraction methods combined with separation technologies and mass spectrometry permit proteomic analysis of GPI-APs from plasma membrane preparations to reveal cell...

  7. Assesment of sperm quality using monoclonal antibodies and proteomic analysis

    Czech Academy of Sciences Publication Activity Database

    Čapková, Jana; Kubátová, Alena; Margaryan, Hasmik; Pěknicová, Jana

    2012-01-01

    Roč. 67, Issue Supplement s1 (2012), s. 28-28 ISSN 1046-7408. [13th International Symposium for Immunology of reproduction "From the roots to the tops of Reproductive Immunology". 22.06.2012-24.06.2012, Varna] R&D Projects: GA ČR(CZ) GA523/09/1793; GA ČR(CZ) GAP503/12/1834 Institutional research plan: CEZ:AV0Z50520701 Keywords : human sperm * immunofluorescence test * human seminal plasma proteins * flow cytometry * proteomic analysis Subject RIV: EC - Immunology

  8. Accelerating the design of biomimetic materials by integrating RNA-seq with proteomics and materials science.

    Science.gov (United States)

    Guerette, Paul A; Hoon, Shawn; Seow, Yiqi; Raida, Manfred; Masic, Admir; Wong, Fong T; Ho, Vincent H B; Kong, Kiat Whye; Demirel, Melik C; Pena-Francesch, Abdon; Amini, Shahrouz; Tay, Gavin Z; Ding, Dawei; Miserez, Ali

    2013-10-01

    Efforts to engineer new materials inspired by biological structures are hampered by the lack of genomic data from many model organisms studied in biomimetic research. Here we show that biomimetic engineering can be accelerated by integrating high-throughput RNA-seq with proteomics and advanced materials characterization. This approach can be applied to a broad range of systems, as we illustrate by investigating diverse high-performance biological materials involved in embryo protection, adhesion and predation. In one example, we rapidly engineer recombinant squid sucker ring teeth proteins into a range of structural and functional materials, including nanopatterned surfaces and photo-cross-linked films that exceed the mechanical properties of most natural and synthetic polymers. Integrating RNA-seq with proteomics and materials science facilitates the molecular characterization of natural materials and the effective translation of their molecular designs into a wide range of bio-inspired materials.

  9. 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...

  10. Quantitative proteomic analysis of intact plastids.

    Science.gov (United States)

    Shiraya, Takeshi; Kaneko, Kentaro; Mitsui, Toshiaki

    2014-01-01

    Plastids are specialized cell organelles in plant cells that are differentiated into various forms including chloroplasts, chromoplasts, and amyloplasts, and fulfill important functions in maintaining the overall cell metabolism and sensing environmental factors such as sunlight. It is therefore important to grasp the mechanisms of differentiation and functional changes of plastids in order to enhance the understanding of vegetality. In this chapter, details of a method for the extraction of intact plastids that makes analysis possible while maintaining the plastid functions are provided; in addition, a quantitative shotgun method for analyzing the composition and changes in the content of proteins in plastids as a result of environmental impacts is described.

  11. Alanine Enhances Aminoglycosides-Induced ROS Production as Revealed by Proteomic Analysis

    Directory of Open Access Journals (Sweden)

    Jin-zhou Ye

    2018-01-01

    Full Text Available Metabolite-enabled killing of antibiotic-resistant pathogens by antibiotics is an attractive strategy to manage antibiotic resistance. Our previous study demonstrated that alanine or/and glucose increased the killing efficacy of kanamycin on antibiotic-resistant bacteria, whose action is through up-regulating TCA cycle, increasing proton motive force and enhancing antibiotic uptake. Despite the fact that alanine altered several metabolic pathways, other mechanisms could be potentially involved in alanine-mediated kanamycin killing of bacteria which remains to be explored. In the present study, we adopted proteomic approach to analyze the proteome changes induced by exogenous alanine. Our results revealed that the expression of three outer membrane proteins was altered and the deletion of nagE and fadL decreased the intracellular kanamycin concentration, implying their possible roles in mediating kanamycin transport. More importantly, the integrated analysis of proteomic and metabolomic data pointed out that alanine metabolism could connect to riboflavin metabolism that provides the source for reactive oxygen species (ROS production. Functional studies confirmed that alanine treatment together with kanamycin could promote ROS production that in turn potentiates the killing of antibiotic-resistant bacteria. Further investigation showed that alanine repressed the transcription of antioxidant-encoding genes, and alanine metabolism to riboflavin metabolism connected with riboflavin metabolism through TCA cycle, glucogenesis pathway and pentose phosphate pathway. Our results suggest a novel mechanism by which alanine facilitates kanamycin killing of antibiotic-resistant bacteria via promoting ROS production.

  12. Transcriptome and proteome analysis of Eucalyptus infected with Calonectria pseudoreteaudii.

    Science.gov (United States)

    Chen, Quanzhu; Guo, Wenshuo; Feng, Lizhen; Ye, Xiaozhen; Xie, Wanfeng; Huang, Xiuping; Liu, Jinyan

    2015-02-06

    Cylindrocladium leaf blight is one of the most severe diseases in Eucalyptus plantations and nurseries. There are Eucalyptus cultivars with resistance to the disease. However, little is known about the defense mechanism of resistant cultivars. Here, we investigated the transcriptome and proteome of Eucalyptus leaves (E. urophylla×E. tereticornis M1), infected or not with Calonectria pseudoreteaudii. A total of 8585 differentially expressed genes (|log2 ratio| ≥1, FDR ≤0.001) at 12 and 24hours post-inoculation were detected using RNA-seq. Transcriptional changes for five genes were further confirmed by qRT-PCR. A total of 3680 proteins at the two time points were identified using iTRAQ technique.The combined transcriptome and proteome analysis revealed that the shikimate/phenylpropanoid pathway, terpenoid biosynthesis, signalling pathway (jasmonic acid and sugar) were activated. The data also showed that some proteins (WRKY33 and PR proteins) which have been reported to involve in plant defense response were up-regulated. However, photosynthesis, nucleic acid metabolism and protein metabolism were impaired by the infection of C. pseudoreteaudii. This work will facilitate the identification of defense related genes and provide insights into Eucalyptus defense responses to Cylindrocladium leaf blight. In this study, a total of 130 proteins and genes involved in the shikimate/phenylpropanoid pathway, terpenoid biosynthesis, signalling pathway, cell transport, carbohydrate and energy metabolism, nucleic acid metabolism and protein metabolism in Eucalyptus leaves after infected with C. pseudoreteaudii were identified. This is the first report of a comprehensive transcriptomic and proteomic analysis of Eucalyptus in response to Calonectria sp. Copyright © 2014 Elsevier B.V. All rights reserved.

  13. Proteomics in medical microbiology.

    Science.gov (United States)

    Cash, P

    2000-04-01

    The techniques of proteomics (high resolution two-dimensional electrophoresis and protein characterisation) are widely used for microbiological research to analyse global protein synthesis as an indicator of gene expression. The rapid progress in microbial proteomics has been achieved through the wide availability of whole genome sequences for a number of bacterial groups. Beyond providing a basic understanding of microbial gene expression, proteomics has also played a role in medical areas of microbiology. Progress has been made in the use of the techniques for investigating the epidemiology and taxonomy of human microbial pathogens, the identification of novel pathogenic mechanisms and the analysis of drug resistance. In each of these areas, proteomics has provided new insights that complement genomic-based investigations. This review describes the current progress in these research fields and highlights some of the technical challenges existing for the application of proteomics in medical microbiology. The latter concern the analysis of genetically heterogeneous bacterial populations and the integration of the proteomic and genomic data for these bacteria. The characterisation of the proteomes of bacterial pathogens growing in their natural hosts remains a future challenge.

  14. Proteome analysis of muscadine grape leaves

    Directory of Open Access Journals (Sweden)

    Sheikh M Basha

    2009-04-01

    Full Text Available Sheikh M Basha1, Ramesh Katam1, Hemanth Vasanthaiah1, Frank Matta21Center for Viticulture and Small Fruit Research, Florida A and M University, Tallahassee, FL, USA; 2Plant and Soil Science Department, Mississippi State University, Mississippi State, MS, USAAbstract: Muscadine grapes are native to the southeastern United States and are used for making wine and consumed as fresh fruit. Grape berries, as ‘sink organs,’ rely on the use of available carbohydrate resources produced by photosynthesis to support their development and composition. A high throughput two-dimensional gel electrophoresis (2-DE was conducted on muscadine (Vitis rotundifolia grape leaf proteins to document complexity in their composition and to determine protein identity and function for enhancing photosynthetic efficiency of muscadine grape. 2-DE resolved muscadine leaf proteins into >258 polypeptides with pIs between 3.5 and 8.0 and molecular weight between 12,000 to 15,0000 Daltons. The consistently expressed proteins were excised and subjected to sequencing. Homology search of protein sequences showed 84% identity with Viridi plantae database. Identity of some of these proteins included RuBisCO, glutamine synthetase, pathogenesis-related protein, glyoxisomal malate dehydrogenase, ribonucleoprotein, chloroplast precursor, oxygen evolving enhancer protein. Comparative analysis of 10 muscadine cultivars showed quantitative differences in expression of 39 polypeptides among these genotypes. The results suggested that the polypeptide composition of muscadine grape leaf is complex, and polypeptide number and amount vary widely among muscadine genotypes, and these variations may be responsible for differences in their physiology, berry and stress tolerance characteristics.Keywords: grapevine, leaves, muscadine, proteins, sequencing, 2-DE

  15. 1001 Proteomes: a functional proteomics portal for the analysis of Arabidopsis thaliana accessions.

    Science.gov (United States)

    Joshi, Hiren J; Christiansen, Katy M; Fitz, Joffrey; Cao, Jun; Lipzen, Anna; Martin, Joel; Smith-Moritz, A Michelle; Pennacchio, Len A; Schackwitz, Wendy S; Weigel, Detlef; Heazlewood, Joshua L

    2012-05-15

    The sequencing of over a thousand natural strains of the model plant Arabidopsis thaliana is producing unparalleled information at the genetic level for plant researchers. To enable the rapid exploitation of these data for functional proteomics studies, we have created a resource for the visualization of protein information and proteomic datasets for sequenced natural strains of A. thaliana. The 1001 Proteomes portal can be used to visualize amino acid substitutions or non-synonymous single-nucleotide polymorphisms in individual proteins of A. thaliana based on the reference genome Col-0. We have used the available processed sequence information to analyze the conservation of known residues subject to protein phosphorylation among these natural strains. The substitution of amino acids in A. thaliana natural strains is heavily constrained and is likely a result of the conservation of functional attributes within proteins. At a practical level, we demonstrate that this information can be used to clarify ambiguously defined phosphorylation sites from phosphoproteomic studies. Protein sets of available natural variants are available for download to enable proteomic studies on these accessions. Together this information can be used to uncover the possible roles of specific amino acids in determining the structure and function of proteins in the model plant A. thaliana. An online portal to enable the community to exploit these data can be accessed at http://1001proteomes.masc-proteomics.org/

  16. Fish gut-liver immunity during homeostasis or inflammation revealed by integrative transcriptome and proteome studies

    Science.gov (United States)

    Wu, Nan; Song, Yu-Long; Wang, Bei; Zhang, Xiang-Yang; Zhang, Xu-Jie; Wang, Ya-Li; Cheng, Ying-Yin; Chen, Dan-Dan; Xia, Xiao-Qin; Lu, Yi-Shan; Zhang, Yong-An

    2016-11-01

    The gut-associated lymphoid tissue, connected with liver via bile and blood, constructs a local immune environment of both defense and tolerance. The gut-liver immunity has been well-studied in mammals, yet in fish remains largely unknown, even though enteritis as well as liver and gallbladder syndrome emerged as a limitation in aquaculture. In this study, we performed integrative bioinformatic analysis for both transcriptomic (gut and liver) and proteomic (intestinal mucus and bile) data, in both healthy and infected tilapias. We found more categories of immune transcripts in gut than liver, as well as more adaptive immune in gut meanwhile more innate in liver. Interestingly reduced differential immune transcripts between gut and liver upon inflammation were also revealed. In addition, more immune proteins in bile than intestinal mucus were identified. And bile probably providing immune effectors to intestinal mucus upon inflammation was deduced. Specifically, many key immune transcripts in gut or liver as well as key immune proteins in mucus or bile were demonstrated. Accordingly, we proposed a hypothesized profile of fish gut-liver immunity, during either homeostasis or inflammation. Current data suggested that fish gut and liver may collaborate immunologically while keep homeostasis using own strategies, including potential unique mechanisms.

  17. Proteomic analysis of fetal programming-related obesity markers.

    Science.gov (United States)

    Lee, Ji Hye; Yoo, Jae Young; You, Young-Ah; Kwon, Woo-Sung; Lee, Sang Mi; Pang, Myung-Geol; Kim, Young Ju

    2015-08-01

    The objectives of this study were to analyze fetal programming in rat brain using proteomic analysis and to identify fetal programming-related obesity markers. Sprague-Dawley rats were divided into four feeding groups: (i) the Ad Libitum (AdLib)/AdLib group was given a normal diet during pregnancy and the lactation period; (ii) the AdLib/maternal food restriction group (FR) was subjected to 50% FR during the lactation period; (iii) the FR/AdLib group was subjected to 50% FR during pregnancy; and (iv) the FR/FR group was subjected to 50% FR during pregnancy and the lactation period. Offspring from each group were sacrificed at 3 weeks of age and whole brains were dissected. To obtain a maximum number of protein markers related to obesity, 2DE and Pathway Studio bioinformatics analysis were performed. The identities of the markers among the selected and candidate proteins were confirmed by Western blotting and immunohistochemistry. Proteomic and bioinformatics analyses revealed that expression of ubiquitin carboxy-terminal hydrolase L1 (UCHL1) and Secernin 1 (SCRN1) were significantly different in the FR/AdLib group compared with the AdLib/AdLib group for both male and female offspring. These findings suggest that UCHL1 and SCRN1 may be used as fetal programming-related obesity markers. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  18. A Method for Microalgae Proteomics Analysis Based on Modified Filter-Aided Sample Preparation.

    Science.gov (United States)

    Li, Song; Cao, Xupeng; Wang, Yan; Zhu, Zhen; Zhang, Haowei; Xue, Song; Tian, Jing

    2017-11-01

    With the fast development of microalgal biofuel researches, the proteomics studies of microalgae increased quickly. A filter-aided sample preparation (FASP) method is widely used proteomics sample preparation method since 2009. Here, a method of microalgae proteomics analysis based on modified filter-aided sample preparation (mFASP) was described to meet the characteristics of microalgae cells and eliminate the error caused by over-alkylation. Using Chlamydomonas reinhardtii as the model, the prepared sample was tested by standard LC-MS/MS and compared with the previous reports. The results showed mFASP is suitable for most of occasions of microalgae proteomics studies.

  19. Proteomic analysis of Magnolia sieboldii K. Koch seed germination.

    Science.gov (United States)

    Lu, Xiu-Jun; Zhang, Xiao-Lin; Mei, Mei; Liu, Guang-Lin; Ma, Bei-Bei

    2016-02-05

    Magnolia sieboldii is a deciduous tree native to China. This species has a deep dormancy characteristic. To better understand seed germination, we used protein analysis of changes in seed protein at 0, 65, 110 and 150 d of stratification. Comparative 2DE analysis of M. sieboldii seed protein profiles at 0, 65, 110 and 150 d of stratification revealed 80 differentially abundance protein species. Comparative analysis showed that ADP-glucose pyrophosphorylase small subunit was degraded during germination. In particular, it was degraded almost completely at 110 d of germination. Starch granules in the microstructure decreased after 65 d of stratification. Starch granules provided a sufficient amount of substrates and ATPs for subsequent germination. Four storage protein species were identified, of which all were down accumulated. Spots 44 and 46 had different MW and pI values, spots 36 and 46 had nearly the same MW with pI shift in the 2-DE gels, suggesting that they might be present as different isoforms of the same protein family and the post translational modification. Our results suggested that degradation of starch granules and storage protein species prepared the seed embryo for growth, as well as regulated seed germination. The present proteomics analysis provides novel insights into the mobilisation of nutrient reserves during the germination of M. sieboldii seeds. To better understand seed germination, a complex developmental process, we developed a proteome analysis of M. sieboldii seed. We performed the first comprehensive proteomic and microstructure analysis during different seed stratification stages of M. sieboldii. Among the 80 protein species, 26 were identified, 7 and 14 protein species were up or down accumulated significantly. Many of the identified key proteins were involved in embryo development, starch biosynthesis and energy metabolism, Microstructure of stratification seed analysis revealed degradation of starch was used for preparing the seed

  20. ProteomicsDB.

    Science.gov (United States)

    Schmidt, Tobias; Samaras, Patroklos; Frejno, Martin; Gessulat, Siegfried; Barnert, Maximilian; Kienegger, Harald; Krcmar, Helmut; Schlegl, Judith; Ehrlich, Hans-Christian; Aiche, Stephan; Kuster, Bernhard; Wilhelm, Mathias

    2018-01-04

    ProteomicsDB (https://www.ProteomicsDB.org) is a protein-centric in-memory database for the exploration of large collections of quantitative mass spectrometry-based proteomics data. ProteomicsDB was first released in 2014 to enable the interactive exploration of the first draft of the human proteome. To date, it contains quantitative data from 78 projects totalling over 19k LC-MS/MS experiments. A standardized analysis pipeline enables comparisons between multiple datasets to facilitate the exploration of protein expression across hundreds of tissues, body fluids and cell lines. We recently extended the data model to enable the storage and integrated visualization of other quantitative omics data. This includes transcriptomics data from e.g. NCBI GEO, protein-protein interaction information from STRING, functional annotations from KEGG, drug-sensitivity/selectivity data from several public sources and reference mass spectra from the ProteomeTools project. The extended functionality transforms ProteomicsDB into a multi-purpose resource connecting quantification and meta-data for each protein. The rich user interface helps researchers to navigate all data sources in either a protein-centric or multi-protein-centric manner. Several options are available to download data manually, while our application programming interface enables accessing quantitative data systematically. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

  1. Proteomics dataset

    DEFF Research Database (Denmark)

    Bennike, Tue Bjerg; Carlsen, Thomas Gelsing; Ellingsen, Torkell

    2017-01-01

    The datasets presented in this article are related to the research articles entitled “Neutrophil Extracellular Traps in Ulcerative Colitis: A Proteome Analysis of Intestinal Biopsies” (Bennike et al., 2015 [1]), and “Proteome Analysis of Rheumatoid Arthritis Gut Mucosa” (Bennike et al., 2017 [2])...... been deposited to the ProteomeXchange Consortium via the PRIDE partner repository with the dataset identifiers PXD001608 for ulcerative colitis and control samples, and PXD003082 for rheumatoid arthritis samples....

  2. [Effect of the lysine guanidination on proteomic analysis].

    Science.gov (United States)

    Zheng, Hao; Mao, Jiawei; Pan, Yanbo; Liu, Zhongshan; Liu, Zheyi; Ye, Mingliang; Zou, Hanfa

    2014-04-01

    The guanidination of lysine side chain was paid great attention in recent years. It plays an important role in qualitative and quantitative proteomics. In this study, based on the results of separated peptides extracted from HeLa cells before and after the guanidination by liquid chromatography-tandem mass spectrometry (LC-MS/MS), the effect of the guanidination of three different kinds of peptides was systematically analyzed. It was found that the selectivity of the guanidination of the lysine side chain was as high as 96.8%. The ratio of identified peptides with lysine at C-term to all peptides increased from 51.7% to 57.3% and more new peptides were identified, while the ratio of peptides with lysine in the middle or without lysine changed little. Further study on the ratio of b and y ions indicated that there were more y ions of peptides with lysine at C-term after the guanidination. The results proved that the selective conversion of lysine to homoarginine by the guanidination could increase the sensitivity and selectivity of mass spectrum. The increased basicity and ability to sequester proton of lysine produced more y ions fragmentation information, which contributed to more identified peptides. It concluded that the lysine guanidination can improve the coverage of proteomic analysis.

  3. Proteomic analysis of proton beam irradiated human melanoma cells.

    Directory of Open Access Journals (Sweden)

    Sylwia Kedracka-Krok

    Full Text Available Proton beam irradiation is a form of advanced radiotherapy providing superior distributions of a low LET radiation dose relative to that of photon therapy for the treatment of cancer. Even though this clinical treatment has been developing for several decades, the proton radiobiology critical to the optimization of proton radiotherapy is far from being understood. Proteomic changes were analyzed in human melanoma cells treated with a sublethal dose (3 Gy of proton beam irradiation. The results were compared with untreated cells. Two-dimensional electrophoresis was performed with mass spectrometry to identify the proteins. At the dose of 3 Gy a minimal slowdown in proliferation rate was seen, as well as some DNA damage. After allowing time for damage repair, the proteomic analysis was performed. In total 17 protein levels were found to significantly (more than 1.5 times change: 4 downregulated and 13 upregulated. Functionally, they represent four categories: (i DNA repair and RNA regulation (VCP, MVP, STRAP, FAB-2, Lamine A/C, GAPDH, (ii cell survival and stress response (STRAP, MCM7, Annexin 7, MVP, Caprin-1, PDCD6, VCP, HSP70, (iii cell metabolism (TIM, GAPDH, VCP, and (iv cytoskeleton and motility (Moesin, Actinin 4, FAB-2, Vimentin, Annexin 7, Lamine A/C, Lamine B. A substantial decrease (2.3 x was seen in the level of vimentin, a marker of epithelial to mesenchymal transition and the metastatic properties of melanoma.

  4. Proteome analysis of bell pepper (Capsicum annuum L.) chromoplasts.

    Science.gov (United States)

    Siddique, Muhammad Asim; Grossmann, Jonas; Gruissem, Wilhelm; Baginsky, Sacha

    2006-12-01

    We report a comprehensive proteome analysis of chromoplasts from bell pepper (Capsicum annuum L.). The combination of a novel strategy for database-independent detection of proteins from tandem mass spectrometry (MS/MS) data with standard database searches allowed us to identify 151 proteins with a high level of confidence. These include several well-known plastid proteins but also novel proteins that were not previously reported from other plastid proteome studies. The majority of the identified proteins are active in plastid carbohydrate and amino acid metabolism. Among the most abundant individual proteins are capsanthin/capsorubin synthase and fibrillin, which are involved in the synthesis and storage of carotenoids that accumulate to high levels in chromoplasts. The relative abundances of the identified chromoplast proteins differ remarkably compared with their abundances in other plastid types, suggesting a chromoplast-specific metabolic network. Our results provide an overview of the major metabolic pathways active in chromoplasts and extend existing knowledge about prevalent metabolic activities of different plastid types.

  5. Proteomic analysis of barley response during early spot blotch infection

    International Nuclear Information System (INIS)

    Al-Daoude, A.; Jawhar, M.; Shoaib, A.; Arabi, M.I.E.

    2015-01-01

    Spot blotch (SB), caused by the fungus Cochliobolus sativus, is a common foliar disease of barley worldwide, but little is known about the host response to infection at the protein level. In this study, a systematic shotgun proteomics approach was chosen to document the early barley response to C. sativus infection. Overall, 28 protein spots were consistently observed as differential in the proteome profiles of the challenged and unchallenged plants. After tryptic digestion, MALDI-TOF/MS analysis and MASCOT database searching identified proteins associated with the defense response including resistance proteins, putative hydrolase, proteinase, kinase and general metabolism and transport proteins. These afford important functions in host resistance and pathogen's inhibition in plants. One of the identified products is a putative NBS-LRR protein which is considered one of the major plant disease resistance proteins identified to date. This work indicates that, in combination with functional genomics, response of barley to challenge by C. sativus involved the recruitment of proteins from various defense pathways.(author)

  6. Comparative proteome analysis of human epithelial ovarian cancer

    Directory of Open Access Journals (Sweden)

    Gagné Jean-Philippe

    2007-09-01

    Full Text Available Abstract Background Epithelial ovarian cancer is a devastating disease associated with low survival prognosis mainly because of the lack of early detection markers and the asymptomatic nature of the cancer until late stage. Using two complementary proteomics approaches, a differential protein expression profile was carried out between low and highly transformed epithelial ovarian cancer cell lines which realistically mimic the phenotypic changes observed during evolution of a tumour metastasis. This investigation was aimed at a better understanding of the molecular mechanisms underlying differentiation, proliferation and neoplastic progression of ovarian cancer. Results The quantitative profiling of epithelial ovarian cancer model cell lines TOV-81D and TOV-112D generated using iTRAQ analysis and two-dimensional electrophoresis coupled to liquid chromatography tandem mass spectrometry revealed some proteins with altered expression levels. Several of these proteins have been the object of interest in cancer research but others were unrecognized as differentially expressed in a context of ovarian cancer. Among these, series of proteins involved in transcriptional activity, cellular metabolism, cell adhesion or motility and cytoskeleton organization were identified, suggesting their possible role in the emergence of oncogenic pathways leading to aggressive cellular behavior. Conclusion The differential protein expression profile generated by the two proteomics approaches combined to complementary characterizations studies will open the way to more exhaustive and systematic representation of the disease and will provide valuable information that may be helpful to uncover the molecular mechanisms related to epithelial ovarian cancer.

  7. Proteomic Analysis of Pachytene Spermatocytes of Sterile Hybrid Male Mice.

    Science.gov (United States)

    Wang, Lu; Guo, Yueshuai; Liu, Wenjing; Zhao, Weidong; Song, Gendi; Zhou, Tao; Huang, Hefeng; Guo, Xuejiang; Sun, Fei

    2016-09-01

    Incompatibilities in interspecific hybrids, such as reduced hybrid fertility and lethality, are common features resulting from reproductive isolation that lead to speciation. Subspecies crosses of house mice produce offspring in which one sex is infertile or absent, yet the molecular mechanisms of hybrid sterility are poorly understood. In this study, we observed extensive asynapsis of chromosomes and disturbance of the sex body in pachytene spermatocytes of sterile F1 males (PWK/Ph female × C57BL/6J male). We report the high-confidence identification of 4005 proteins in the pachytene spermatocytes of fertile F1 males (PWK/Ph male × C57BL/6J female) and sterile F1 males (PWK/Ph female × C57BL/6J male), of which 215 were upregulated and 381 were downregulated. Bioinformatics analysis of the proteome led to the identification of 43 and 59 proteins known to be essential for male meiosis and spermatogenesis in mice, respectively. Characterization of the proteome of pachytene spermatocytes associated with hybrid male sterility provides an inventory of proteins that is useful for understanding meiosis and the mechanisms of hybrid male infertility. © 2016 by the Society for the Study of Reproduction, Inc.

  8. In-depth analysis of the adipocyte proteome by mass spectrometry and bioinformatics

    DEFF Research Database (Denmark)

    Adachi, Jun; Kumar, Chanchal; Zhang, Yanling

    2007-01-01

    , mitochondria, membrane, and cytosol of 3T3-L1 adipocytes. We identified 3,287 proteins while essentially eliminating false positives, making this one of the largest high confidence proteomes reported to date. Comprehensive bioinformatics analysis revealed that the adipocyte proteome, despite its specialized...

  9. Subnanogram proteomics: Impact of LC column selection, MS instrumentation and data analysis strategy on proteome coverage for trace samples

    Energy Technology Data Exchange (ETDEWEB)

    Zhu, Ying; Zhao, Rui; Piehowski, Paul D.; Moore, Ronald J.; Lim, Sujung; Orphan, Victoria J.; Paša-Tolić, Ljiljana; Qian, Wei-Jun; Smith, Richard D.; Kelly, Ryan T.

    2018-04-01

    One of the greatest challenges for mass spectrometry (MS)-based proteomics is the limited ability to analyze small samples. Here we investigate the relative contributions of liquid chromatography (LC), MS instrumentation and data analysis methods with the aim of improving proteome coverage for sample sizes ranging from 0.5 ng to 50 ng. We show that the LC separations utilizing 30-µm-i.d. columns increase signal intensity by >3-fold relative to those using 75-µm-i.d. columns, leading to 32% increase in peptide identifications. The Orbitrap Fusion Lumos mass spectrometer significantly boosted both sensitivity and sequencing speed relative to earlier generation Orbitraps (e.g., LTQ-Orbitrap), leading to a ~3× increase in peptide identifications and 1.7× increase in identified protein groups for 2 ng tryptic digests of bacterial lysate. The Match Between Runs algorithm of open-source MaxQuant software further increased proteome coverage by ~ 95% for 0.5 ng samples and by ~42% for 2 ng samples. The present platform is capable of identifying >3000 protein groups from tryptic digestion of cell lysates equivalent to 50 HeLa cells and 100 THP-1 cells (~10 ng total proteins), respectively, and >950 proteins from subnanogram bacterial and archaeal cell lysates. The present ultrasensitive LC-MS platform is expected to enable deep proteome coverage for subnanogram samples, including single mammalian cells.

  10. Comprehensive Proteomic Analysis of Human Milk-derived Extracellular Vesicles Unveils a Novel Functional Proteome Distinct from Other Milk Components.

    Science.gov (United States)

    van Herwijnen, Martijn J C; Zonneveld, Marijke I; Goerdayal, Soenita; Nolte-'t Hoen, Esther N M; Garssen, Johan; Stahl, Bernd; Maarten Altelaar, A F; Redegeld, Frank A; Wauben, Marca H M

    2016-11-01

    Breast milk contains several macromolecular components with distinctive functions, whereby milk fat globules and casein micelles mainly provide nutrition to the newborn, and whey contains molecules that can stimulate the newborn's developing immune system and gastrointestinal tract. Although extracellular vesicles (EV) have been identified in breast milk, their physiological function and composition has not been addressed in detail. EV are submicron sized vehicles released by cells for intercellular communication via selectively incorporated lipids, nucleic acids, and proteins. Because of the difficulty in separating EV from other milk components, an in-depth analysis of the proteome of human milk-derived EV is lacking. In this study, an extensive LC-MS/MS proteomic analysis was performed of EV that had been purified from breast milk of seven individual donors using a recently established, optimized density-gradient-based EV isolation protocol. A total of 1963 proteins were identified in milk-derived EV, including EV-associated proteins like CD9, Annexin A5, and Flotillin-1, with a remarkable overlap between the different donors. Interestingly, 198 of the identified proteins are not present in the human EV database Vesiclepedia, indicating that milk-derived EV harbor proteins not yet identified in EV of different origin. Similarly, the proteome of milk-derived EV was compared with that of other milk components. For this, data from 38 published milk proteomic studies were combined in order to construct the total milk proteome, which consists of 2698 unique proteins. Remarkably, 633 proteins identified in milk-derived EV have not yet been identified in human milk to date. Interestingly, these novel proteins include proteins involved in regulation of cell growth and controlling inflammatory signaling pathways, suggesting that milk-derived EVs could support the newborn's developing gastrointestinal tract and immune system. Overall, this study provides an expansion of

  11. Pathway analysis of kidney cancer using proteomics and metabolic profiling

    Directory of Open Access Journals (Sweden)

    Fiehn Oliver

    2006-11-01

    Full Text Available Abstract Background Renal cell carcinoma (RCC is the sixth leading cause of cancer death and is responsible for 11,000 deaths per year in the US. Approximately one-third of patients present with disease which is already metastatic and for which there is currently no adequate treatment, and no biofluid screening tests exist for RCC. In this study, we have undertaken a comprehensive proteomic analysis and subsequently a pathway and network approach to identify biological processes involved in clear cell RCC (ccRCC. We have used these data to investigate urinary markers of RCC which could be applied to high-risk patients, or to those being followed for recurrence, for early diagnosis and treatment, thereby substantially reducing mortality of this disease. Results Using 2-dimensional electrophoresis and mass spectrometric analysis, we identified 31 proteins which were differentially expressed with a high degree of significance in ccRCC as compared to adjacent non-malignant tissue, and we confirmed some of these by immunoblotting, immunohistochemistry, and comparison to published transcriptomic data. When evaluated by several pathway and biological process analysis programs, these proteins are demonstrated to be involved with a high degree of confidence (p values Conclusion Extensive pathway and network analysis allowed for the discovery of highly significant pathways from a set of clear cell RCC samples. Knowledge of activation of these processes will lead to novel assays identifying their proteomic and/or metabolomic signatures in biofluids of patient at high risk for this disease; we provide pilot data for such a urinary bioassay. Furthermore, we demonstrate how the knowledge of networks, processes, and pathways altered in kidney cancer may be used to influence the choice of optimal therapy.

  12. Detachable strong cation exchange monolith, integrated with capillary zone electrophoresis and coupled with pH gradient elution, produces improved sensitivity and numbers of peptide identifications during bottom-up analysis of complex proteomes.

    Science.gov (United States)

    Zhang, Zhenbin; Yan, Xiaojing; Sun, Liangliang; Zhu, Guijie; Dovichi, Norman J

    2015-04-21

    A detachable sulfonate-silica hybrid strong cation-exchange monolith was synthesized in a fused silica capillary, and used for solid phase extraction with online pH gradient elution during capillary zone electrophoresis-tandem mass spectrometry (CZE-MS/MS) proteomic analysis. Tryptic digests were prepared in 50 mM formic acid and loaded onto the strong cation-exchange monolith. Fractions were eluted using a series of buffers with lower concentration but higher pH values than the 50 mM formic acid background electrolyte. This combination of elution and background electrolytes results in both sample stacking and formation of a dynamic pH junction and allows use of relatively large elution buffer volumes while maintaining reasonable peak efficiency and resolution. A series of five pH bumps were applied to elute E. coli tryptic peptides from the monolith, followed by analysis using CZE coupled to an LTQ-Orbitrap Velos mass spectrometer; 799 protein groups and 3381 peptides were identified from 50 ng of the digest in a 2.5 h analysis, which approaches the identification rate for this organism that was obtained with an Orbitrap Fusion. We attribute the improved numbers of peptide and protein identifications to the efficient fractionation by the online pH gradient elution, which decreased the complexity of the sample in each elution step and improved the signal intensity of low abundance peptides. We also performed a comparative analysis using a nanoACQUITY UltraPerformance LCH system. Similar numbers of protein and peptide identifications were produced by the two methods. Protein identifications showed significant overlap between the two methods, whereas peptide identifications were complementary.

  13. Proteomic analysis of liver in rats chronically exposed to fluoride.

    Directory of Open Access Journals (Sweden)

    Heloísa Aparecida Barbosa da Silva Pereira

    Full Text Available Fluoride (F is a potent anti-cariogenic element, but when ingestion is excessive, systemic toxicity may be observed. This can occur as acute or chronic responses, depending on both the amount of F and the time of exposure. The present study identified the profile of protein expression possibly associated with F-induced chronic hepatotoxicity. Weanling male Wistar rats (three-weeks old were divided into three groups and treated with drinking water containing 0, 5 or 50 mg/L F for 60 days (n=6/group. At this time point, serum and livers were collected for F analysis, which was done using the ion-sensitive electrode, after hexamethyldisiloxane-facilitated diffusion. Livers were also submitted to histological and proteomic analyses (2D-PAGE followed by LC-MS/MS. Western blotting was done for confirmation of the proteomic data A dose-response was observed in serum F levels. In the livers, F levels were significantly increased in the 50 mg/L F group compared to groups treated with 0 and 5 mg/L F. Liver morphometric analysis did not reveal alterations in the cellular structures and lipid droplets were present in all groups. Proteomic quantitative intensity analysis detected 33, 44, and 29 spots differentially expressed in the comparisons between control vs. 5 mg/L F, control vs. 50 mg/L F, and 5 mg/L vs. 50 mg/L F, respectively. From these, 92 proteins were successfully identified. In addition, 18, 1, and 5 protein spots were shown to be exclusive in control, 5, and 50 mg/L F, respectively. Most of proteins were related to metabolic process and pronounced alterations were seen for the high-F level group. In F-treated rats, changes in the apolipoprotein E (ApoE and GRP-78 expression may account for the F-induced toxicity in the liver. This can contribute to understanding the molecular mechanisms underlying hepatoxicity induced by F, by indicating key-proteins that should be better addressed in future studies.

  14. Analysis of Pacific oyster larval proteome and its response to high-CO2

    KAUST Repository

    Dineshram, R.; Wong, Kelvin K.W.; Xiao, Shu; Yu, Ziniu; Qian, Pei Yuan; Thiyagarajan, Vengatesen

    2012-01-01

    Most calcifying organisms show depressed metabolic, growth and calcification rates as symptoms to high-CO2 due to ocean acidification (OA) process. Analysis of the global expression pattern of proteins (proteome analysis) represents a powerful tool

  15. Identification and proteomic analysis of osteoblast-derived exosomes

    Energy Technology Data Exchange (ETDEWEB)

    Ge, Min; Ke, Ronghu; Cai, Tianyi; Yang, Junyi; Mu, Xiongzheng, E-mail: cranio@vip.163.com

    2015-11-06

    Exosomes are nanometer-sized vesicles with the function of intercellular communication, and they are released by various cell types. To reveal the knowledge about the exosomes from osteoblast, and explore the potential functions of osteogenesis, we isolated microvesicles from supernatants of mouse Mc3t3 by ultracentrifugation, characterized exosomes by electron microscopy and immunoblotting and presented the protein profile by proteomic analysis. The result demonstrated that microvesicles were between 30 and 100 nm in diameter, round shape with cup-like concavity and expressed exosomal marker tumor susceptibility gene (TSG) 101 and flotillin (Flot) 1. We identified a total number of 1069 proteins among which 786 proteins overlap with ExoCarta database. Gene Oncology analysis indicated that exosomes mostly derived from plasma membrane and mainly involved in protein localization and intracellular signaling. The Ingenuity Pathway Analysis showed pathways are mostly involved in exosome biogenesis, formation, uptake and osteogenesis. Among the pathways, eukaryotic initiation factor 2 pathways played an important role in osteogenesis. Our study identified osteoblast-derived exosomes, unveiled the content of them, presented potential osteogenesis-related proteins and pathways and provided a rich proteomics data resource that will be valuable for further studies of the functions of individual proteins in bone diseases. - Highlights: • We for the first time identified exosomes from mouse osteoblast. • Osteoblasts-derived exosomes contain osteoblast peculiar proteins. • Proteins from osteoblasts-derived exosomes are intently involved in EIF2 pathway. • EIF2α from the EIF2 pathway plays an important role in osteogenesis.

  16. LFQProfiler and RNP(xl): Open-Source Tools for Label-Free Quantification and Protein-RNA Cross-Linking Integrated into Proteome Discoverer.

    Science.gov (United States)

    Veit, Johannes; Sachsenberg, Timo; Chernev, Aleksandar; Aicheler, Fabian; Urlaub, Henning; Kohlbacher, Oliver

    2016-09-02

    Modern mass spectrometry setups used in today's proteomics studies generate vast amounts of raw data, calling for highly efficient data processing and analysis tools. Software for analyzing these data is either monolithic (easy to use, but sometimes too rigid) or workflow-driven (easy to customize, but sometimes complex). Thermo Proteome Discoverer (PD) is a powerful software for workflow-driven data analysis in proteomics which, in our eyes, achieves a good trade-off between flexibility and usability. Here, we present two open-source plugins for PD providing additional functionality: LFQProfiler for label-free quantification of peptides and proteins, and RNP(xl) for UV-induced peptide-RNA cross-linking data analysis. LFQProfiler interacts with existing PD nodes for peptide identification and validation and takes care of the entire quantitative part of the workflow. We show that it performs at least on par with other state-of-the-art software solutions for label-free quantification in a recently published benchmark ( Ramus, C.; J. Proteomics 2016 , 132 , 51 - 62 ). The second workflow, RNP(xl), represents the first software solution to date for identification of peptide-RNA cross-links including automatic localization of the cross-links at amino acid resolution and localization scoring. It comes with a customized integrated cross-link fragment spectrum viewer for convenient manual inspection and validation of the results.

  17. Comparative proteomics and codon substitution analysis reveal mechanisms of differential resistance to hypoxia in congeneric snails

    KAUST Repository

    Mu, Huawei; Sun, Jin; Cheung, Siu Gin; Fang, Ling; Zhou, Haiyun; Luan, Tiangang; Zhang, Huoming; Wong, Chris K.C.; Qiu, Jian-Wen

    2017-01-01

    Although high-throughput proteomics has been widely applied to study mechanisms of environmental adaptation, the conclusions from studies that are based on one species can be confounded by phylogeny. We compare the freshwater snail Pomacea canaliculata (a notorious invasive species) and its congener Pomacea diffusa (a non-invasive species) to understand the molecular mechanisms of their differential resistance to hypoxia. A 72-h acute exposure experiment showed that P. canaliculata is more tolerant to hypoxia than P. diffusa. The two species were then exposed to three levels of dissolved oxygen (6.7, 2.0 and 1.0mgL−1) for 8h, and their gill proteins were analyzed using iTRAQ-coupled LC-MS/MS. The two species showed striking differences in protein expression profiles, with the more hypoxia tolerant P. canaliculata having more up-regulated proteins in signal transduction and down-regulated proteins in glycolysis and the tricarboxylic acid cycle. Evolutionary analysis revealed five orthologous genes encoding differentially expressed proteins having clear signal of positive selection, indicating selection has acted on some of the hypoxia responsive genes. Our case study has highlighted the potential of integrated proteomics and comparative evolutionary analysis for understanding the genetic basis of adaptation to global environmental change in non-model species. SignificanceRapid globalization in recent decades has greatly facilitated species introduction around the world. Successfully established introduced species, so-called invasive species, have threatened the invaded ecosystems. There has been substantial interest in studying how invasive species respond to extreme environmental conditions because the results can help not only predict their range of expansion and manage their impact, but also may reveal the adaptive mechanisms underlying their invasiveness. Our study has adopted a comparative approach to study the differential physiological and proteomic

  18. Comparative proteomics and codon substitution analysis reveal mechanisms of differential resistance to hypoxia in congeneric snails

    KAUST Repository

    Mu, Huawei

    2017-11-06

    Although high-throughput proteomics has been widely applied to study mechanisms of environmental adaptation, the conclusions from studies that are based on one species can be confounded by phylogeny. We compare the freshwater snail Pomacea canaliculata (a notorious invasive species) and its congener Pomacea diffusa (a non-invasive species) to understand the molecular mechanisms of their differential resistance to hypoxia. A 72-h acute exposure experiment showed that P. canaliculata is more tolerant to hypoxia than P. diffusa. The two species were then exposed to three levels of dissolved oxygen (6.7, 2.0 and 1.0mgL−1) for 8h, and their gill proteins were analyzed using iTRAQ-coupled LC-MS/MS. The two species showed striking differences in protein expression profiles, with the more hypoxia tolerant P. canaliculata having more up-regulated proteins in signal transduction and down-regulated proteins in glycolysis and the tricarboxylic acid cycle. Evolutionary analysis revealed five orthologous genes encoding differentially expressed proteins having clear signal of positive selection, indicating selection has acted on some of the hypoxia responsive genes. Our case study has highlighted the potential of integrated proteomics and comparative evolutionary analysis for understanding the genetic basis of adaptation to global environmental change in non-model species. SignificanceRapid globalization in recent decades has greatly facilitated species introduction around the world. Successfully established introduced species, so-called invasive species, have threatened the invaded ecosystems. There has been substantial interest in studying how invasive species respond to extreme environmental conditions because the results can help not only predict their range of expansion and manage their impact, but also may reveal the adaptive mechanisms underlying their invasiveness. Our study has adopted a comparative approach to study the differential physiological and proteomic

  19. Functional proteomic analysis of Ankaferd® Blood Stopper

    Directory of Open Access Journals (Sweden)

    Duygu Özel Demiralp

    2010-06-01

    Full Text Available Objective: Ankaferd® Blood Stopper (ABS comprises a standardized mixture of the plants Thymus vulgaris, Glycyrrhiza glabra, Vitis vinifera, Alpinia officinarum, and Urtica dioica. The basic mechanism of action for ABS is the formation of an encapsulated protein network that provides focal points for vital erythrocyte aggregation. ABS–induced protein network formation with blood cells, particularly erythrocytes, covers the primary and secondary hemostatic system without disturbing individual coagulation factors. Materials and Methods: To understand the effect mechanisms of ABS on hemostasis, a proteomic analysis using 2D gel electrophoresis and mass spectrometer was performed. Results: Proteins of plant origin in Ankaferd® were NADP-dependent-malic enzyme, ribulose bisphosphate-carboxylase-large chain, maturase K, ATP synthase subunit-beta, ATP synthase subunit-alpha, chalcone-flavanone isomerase-1, chalcone-flavanone isomerase-2, and actin-depolymerizing factor. Furthermore, functional proteomic studies revealed that proteins resembling human peptides have been detected within Ankaferd®, including ATP synthase, mucin-16 (CD164 sialomucin-like 2 protein, coiled-coil domain containing 141 hypothetical protein LOC283638 isoform 1, hypothetical protein LOC283638 isoform 2, dynactin 5, complex I intermediate-associated protein 30, mitochondrial, NADH dehydrogenase (ubiquinone 1 alpha subcomplex, TP synthase, H+ transporting, mitochondrial actin binding 1 isoform, LIM domain and actin binding 1 isoform a, LIM domain and actin binding 1 isoform b, spectrin alpha non erythrocytic 1, prolactin releasing hormone receptor, utrophin, tet oncogene family member 2 isoform b, protein phosphatase 1 regulatory subunit 12A, NIMA (never in mitosis gene a-related kinase, ATP-binding cassette protein C12, Homo sapiens malic enzyme 1, mitochondrial NADP(+-dependent malic enzyme 3, ME2 protein, nuclear factor 1 B-type, abhydrolase domain-containing protein 12B, E

  20. Proteomic analysis of soybean hypocotyl during recovery after flooding stress.

    Science.gov (United States)

    Khan, Mudassar Nawaz; Sakata, Katsumi; Komatsu, Setsuko

    2015-05-21

    Soybean is a nutritionally important crop, but exhibits reduced growth and yields under flooding stress. To investigate soybean responses during post-flooding recovery, a gel-free proteomic technique was used to examine the protein profile in the hypocotyl. Two-day-old soybeans were flooded for 2 days and hypocotyl was collected under flooding and during the post-flooding recovery period. A total of 498 and 70 proteins were significantly changed in control and post-flooding recovering soybeans, respectively. Based on proteomic and clustering analyses, three proteins were selected for mRNA expression and enzyme activity assays. Pyruvate kinase was increased under flooding, but gradually decreased during post-flooding recovery period at protein abundance, mRNA, and enzyme activity levels. Nucleotidylyl transferase was decreased under flooding and increased during post-flooding recovery at both mRNA expression and enzyme activity levels. Beta-ketoacyl reductase 1 was increased under flooding and decreased during recovery at protein abundance and mRNA expression levels, but its enzyme activity gradually increased during the post-flooding recovery period. These results suggest that pyruvate kinase, nucleotidylyl transferase, and beta-ketoacyl reductase play key roles in post-flooding recovery in soybean hypocotyl by promoting glycolysis for the generation of ATP and regulation of secondary metabolic pathways. This study analyzed post-flooding recovery response mechanisms in soybean hypocotyl, which is a model organ for studying secondary growth, using a gel-free proteomic technique. Mass spectrometry analysis of proteins extracted from soybean hypocotyls identified 20 common proteins between control and flooding-stressed soybeans that changed significantly in abundance over time. The hypocotyl proteins that changed during post-flooding recovery were assigned to protein, development, secondary metabolism, and glycolysis categories. The analysis revealed that three

  1. Biochemical Markers of Brain Injury: An Integrated Proteomics-Based Approach

    Science.gov (United States)

    2006-02-01

    some variability between protein assay measurements. Using densitometric analysis after 1D-PAGE, microfiltration alone showed an 11 ( 5% sample reduction...KOPETSCH, O., WOSZCZYK, A., et al. (2003). Serum S-100B protein as a molecular marker in severe trau- matic brain injury. Restor . Neurol. Neurosci. 21...proteomics. Implications in the search for preventive initiatives to slow the clinical progression of Alzheimer’s disease dementia. Restor . Neurol. Neurosci

  2. Proteome analysis of Radiation-induced pulmonary fibrosis

    International Nuclear Information System (INIS)

    Song, Jie Young; Lim, Hee Soon; Kim, Hyung Doo; Shim, Ji Young; Han, Young Soo; Son, Hyeog Jin Son; Yun, Yeon Sook

    2005-01-01

    Pulmonary fibrosis is perhaps the most universal late effect of organ damage after both chemical insult and irradiation in the treatment of lung cancer. The use chemotherapy and radiation therapy, alone or combined, can be associated with clinically significant pulmonary toxicity, which leads to pneumonia and pulmonary fibrosis. It is also reported that about 100,000 people in the United States are suffered from pulmonary fibrosis. Therefore, pulmonary fibrosis will be more focused by medicinal researchers. Because current therapies, aimed at inhibiting pulmonary inflammation that often precedes fibrosis, are effective only in a minority of suffered patients, novel therapeutic methods are highly needed. Some researchers have used bleomycininduced pulmonary fibrosis as a basis for looking at the molecular mechanisms of fibrosis, and total gene expression was monitored using genomics method. However, radiation-induced pulmonary fibrosis has not been fully focused and investigated. Here, we have analyzed changes in gene expression in response to γ- irradiation by using proteomic analysis

  3. Proteomic analysis of pancreas derived from adult cloned pig

    International Nuclear Information System (INIS)

    Chae, Jung-Il; Cho, Young Keun; Cho, Seong-Keun; Kim, Jin-Hoi; Han, Yong-Mahn; Koo, Deog-Bon; Lee, Kyung-Kwang

    2008-01-01

    The potential medical applications of animal cloning include xenotransplantation, but the complex molecular cascades that control porcine organ development are not fully understood. Still, it has become apparent that organs derived from cloned pigs may be suitable for transplantation into humans. In this study, we examined the pancreas of an adult cloned pig developed through somatic cell nuclear transfer (SCNT) using two-dimensional electrophoresis (2-DE) and Western blotting. Proteomic analysis revealed 69 differentially regulated proteins, including such apoptosis-related species as annexins, lamins, and heat shock proteins, which were unanimously upregulated in the SCNT sample. Among the downregulated proteins in SCNT pancreas were peroxiredoxins and catalase. Western blot results indicate that several antioxidant enzymes and the anti-apoptotic protein were downregulated in SCNT pancreas, whereas several caspases were upregulated. Together, these data suggest that the accumulation of reactive oxygen species (ROS) in the pancreas of an adult cloned pig leads to apoptosis

  4. Analysis of the SUMO2 Proteome during HSV-1 Infection.

    Directory of Open Access Journals (Sweden)

    Elizabeth Sloan

    2015-07-01

    Full Text Available Covalent linkage to members of the small ubiquitin-like (SUMO family of proteins is an important mechanism by which the functions of many cellular proteins are regulated. Sumoylation has roles in the control of protein stability, activity and localization, and is involved in the regulation of transcription, gene expression, chromatin structure, nuclear transport and RNA metabolism. Sumoylation is also linked, both positively and negatively, with the replication of many different viruses both in terms of modification of viral proteins and modulation of sumoylated cellular proteins that influence the efficiency of infection. One prominent example of the latter is the widespread reduction in the levels of cellular sumoylated species induced by herpes simplex virus type 1 (HSV-1 ubiquitin ligase ICP0. This activity correlates with relief from intrinsic immunity antiviral defence mechanisms. Previous work has shown that ICP0 is selective in substrate choice, with some sumoylated proteins such the promyelocytic leukemia protein PML being extremely sensitive, while RanGAP is completely resistant. Here we present a comprehensive proteomic analysis of changes in the cellular SUMO2 proteome during HSV-1 infection. Amongst the 877 potentially sumoylated species detected, we identified 124 whose abundance was decreased by a factor of 3 or more by the virus, several of which were validated by western blot and expression analysis. We found many previously undescribed substrates of ICP0 whose degradation occurs by a range of mechanisms, influenced or not by sumoylation and/or the SUMO2 interaction motif within ICP0. Many of these proteins are known or are predicted to be involved in the regulation of transcription, chromatin assembly or modification. These results present novel insights into mechanisms and host cell proteins that might influence the efficiency of HSV-1 infection.

  5. Proteomic analysis of honeybee (Apis mellifera L. pupae head development.

    Directory of Open Access Journals (Sweden)

    Aijuan Zheng

    Full Text Available The honeybee pupae development influences its future adult condition as well as honey and royal jelly productions. However, the molecular mechanism that regulates honeybee pupae head metamorphosis is still poorly understood. To further our understand of the associated molecular mechanism, we investigated the protein change of the honeybee pupae head at 5 time-points using 2-D electrophoresis, mass spectrometry, bioinformatics, quantitative real-time polymerase chain reaction and Western blot analysis. Accordingly, 58 protein spots altered their expression across the 5 time points (13-20 days, of which 36 proteins involved in the head organogenesis were upregulated during early stages (13-17 days. However, 22 proteins involved in regulating the pupae head neuron and gland development were upregulated at later developmental stages (19-20 days. Also, the functional enrichment analysis further suggests that proteins related to carbohydrate metabolism and energy production, development, cytoskeleton and protein folding were highly involved in the generation of organs and development of honeybee pupal head. Furthermore, the constructed protein interaction network predicted 33 proteins acting as key nodes of honeybee pupae head growth of which 9 and 4 proteins were validated at gene and protein levels, respectively. In this study, we uncovered potential protein species involved in the formation of honeybee pupae head development along with their specific temporal requirements. This first proteomic result allows deeper understanding of the proteome profile changes during honeybee pupae head development and provides important potential candidate proteins for future reverse genetic research on honeybee pupae head development to improve the performance of related organs.

  6. An individual urinary proteome analysis in normal human beings to define the minimal sample number to represent the normal urinary proteome

    Directory of Open Access Journals (Sweden)

    Liu Xuejiao

    2012-11-01

    Full Text Available Abstract Background The urinary proteome has been widely used for biomarker discovery. A urinary proteome database from normal humans can provide a background for discovery proteomics and candidate proteins/peptides for targeted proteomics. Therefore, it is necessary to define the minimum number of individuals required for sampling to represent the normal urinary proteome. Methods In this study, inter-individual and inter-gender variations of urinary proteome were taken into consideration to achieve a representative database. An individual analysis was performed on overnight urine samples from 20 normal volunteers (10 males and 10 females by 1DLC/MS/MS. To obtain a representative result of each sample, a replicate 1DLCMS/MS analysis was performed. The minimal sample number was estimated by statistical analysis. Results For qualitative analysis, less than 5% of new proteins/peptides were identified in a male/female normal group by adding a new sample when the sample number exceeded nine. In addition, in a normal group, the percentage of newly identified proteins/peptides was less than 5% upon adding a new sample when the sample number reached 10. Furthermore, a statistical analysis indicated that urinary proteomes from normal males and females showed different patterns. For quantitative analysis, the variation of protein abundance was defined by spectrum count and western blotting methods. And then the minimal sample number for quantitative proteomic analysis was identified. Conclusions For qualitative analysis, when considering the inter-individual and inter-gender variations, the minimum sample number is 10 and requires a balanced number of males and females in order to obtain a representative normal human urinary proteome. For quantitative analysis, the minimal sample number is much greater than that for qualitative analysis and depends on the experimental methods used for quantification.

  7. Proteomic analysis of the Arabidopsis thaliana-Botrytis cinerea ...

    African Journals Online (AJOL)

    A two-dimensional liquid chromatography (2D LC) system, ProteomeLab PF 2D, was employed to study the defence proteome of Arabidopsis following infection with the necrotrophic fungal pathogen, Botrytis cinerea. This system demonstrated differential protein expression in control and treated samples in some fractions.

  8. Dioscorea alata tuber proteome analysis shows over thirty dioscorin isoforms and novel tuber proteins.

    Science.gov (United States)

    Sharma, Shruti; Gupta, Ravi; Deswal, Renu

    2017-05-01

    In Dioscorea, dioscorin (31 kDa) is the major storage protein constituting 85% of the total tuber proteins. An integrated proteomic and biochemical approach was used to understand the physiological role of dioscorin in the two contrasting growth stages (germinating and mature tuber). HPLC analysis showed 3 fold reduction in mannitol and 12.88 and 1.24 fold increase in sucrose and maltose in the germinating tuber. A 1.8 and 3 fold increase in sucrose phosphate synthase and mannitol dehydrogenase activity respectively was observed in the germinating tuber while a 2 fold higher invertase probably lowers the sucrose accumulation in the mature tuber. SDS-PAGE and 2-D maps of the mature and germinating tubers confirmed depletion (more than 50%) of dioscorin on germination. Dioscorin was purified using ion exchange and gel filtration chromatography with 43.32 fold purification and 38.16 yield. Out of a trail of 35 spots at 31 kDa only 12 spots (identified as dioscorin isoforms) were present in the 2D gel of the purified fraction. To search for other tuber proteins besides dioscorin, the unbound fractions of DEAE column were analysed by 2DGE. DREB 1A, caffeic acid 3-O-methyltransferase and Rab-1 small GTP binding protein were identified perhaps for the first time in the Dioscorea proteome. The interactome analysis revealed these to be involved in oxidative stress, carotenoid synthesis and vesicular transport. This is perhaps the first attempt to identify tuber proteome (although limited) and to understand the physiological significance of these proteins. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  9. A knowledge-based T2-statistic to perform pathway analysis for quantitative proteomic data.

    Science.gov (United States)

    Lai, En-Yu; Chen, Yi-Hau; Wu, Kun-Pin

    2017-06-01

    Approaches to identify significant pathways from high-throughput quantitative data have been developed in recent years. Still, the analysis of proteomic data stays difficult because of limited sample size. This limitation also leads to the practice of using a competitive null as common approach; which fundamentally implies genes or proteins as independent units. The independent assumption ignores the associations among biomolecules with similar functions or cellular localization, as well as the interactions among them manifested as changes in expression ratios. Consequently, these methods often underestimate the associations among biomolecules and cause false positives in practice. Some studies incorporate the sample covariance matrix into the calculation to address this issue. However, sample covariance may not be a precise estimation if the sample size is very limited, which is usually the case for the data produced by mass spectrometry. In this study, we introduce a multivariate test under a self-contained null to perform pathway analysis for quantitative proteomic data. The covariance matrix used in the test statistic is constructed by the confidence scores retrieved from the STRING database or the HitPredict database. We also design an integrating procedure to retain pathways of sufficient evidence as a pathway group. The performance of the proposed T2-statistic is demonstrated using five published experimental datasets: the T-cell activation, the cAMP/PKA signaling, the myoblast differentiation, and the effect of dasatinib on the BCR-ABL pathway are proteomic datasets produced by mass spectrometry; and the protective effect of myocilin via the MAPK signaling pathway is a gene expression dataset of limited sample size. Compared with other popular statistics, the proposed T2-statistic yields more accurate descriptions in agreement with the discussion of the original publication. We implemented the T2-statistic into an R package T2GA, which is available at https

  10. Comparative Proteomic Analysis of Hymenolepis diminuta Cysticercoid and Adult Stages

    Directory of Open Access Journals (Sweden)

    Anna Sulima

    2018-01-01

    Full Text Available Cestodiases are common parasitic diseases of animals and humans. As cestodes have complex lifecycles, hexacanth larvae, metacestodes (including cysticercoids, and adults produce proteins allowing them to establish invasion and to survive in the hostile environment of the host. Hymenolepis diminuta is the most commonly used model cestode in experimental parasitology. The aims of the present study were to perform a comparative proteomic analysis of two consecutive developmental stages of H. diminuta (cysticercoid and adult and to distinguish proteins which might be characteristic for each of the stages from those shared by both stages. Somatic proteins of H. diminuta were isolated from 6-week-old cysticercoids and adult tapeworms. Cysticercoids were obtained from experimentally infected beetles, Tenebrio molitor, whereas adult worms were collected from experimentally infected rats. Proteins were separated by GeLC-MS/MS (one dimensional gel electrophoresis coupled with liquid chromatography and tandem mass spectrometry. Additionally protein samples were digested in-liquid and identified by LC-MS/MS. The identified proteins were classified according to molecular function, cellular components and biological processes. Our study showed a number of differences and similarities in the protein profiles of cysticercoids and adults; 233 cysticercoid and 182 adult proteins were identified. From these proteins, 131 were present only in the cysticercoid and 80 only in the adult stage samples. Both developmental stages shared 102 proteins; among which six represented immunomodulators and one is a potential drug target. In-liquid digestion and LC-MS/MS complemented and confirmed some of the GeLC-MS/MS identifications. Possible roles and functions of proteins identified with both proteomic approaches are discussed.

  11. Primary metabolism in Lactobacillus sakei food isolates by proteomic analysis

    Directory of Open Access Journals (Sweden)

    Champomier-Vergès Marie-Christine

    2010-04-01

    Full Text Available Abstract Background Lactobacillus sakei is an important food-associated lactic acid bacterium commonly used as starter culture for industrial meat fermentation, and with great potential as a biopreservative in meat and fish products. Understanding the metabolic mechanisms underlying the growth performance of a strain to be used for food fermentations is important for obtaining high-quality and safe products. Proteomic analysis was used to study the primary metabolism in ten food isolates after growth on glucose and ribose, the main sugars available for L. sakei in meat and fish. Results Proteins, the expression of which varied depending on the carbon source were identified, such as a ribokinase and a D-ribose pyranase directly involved in ribose catabolism, and enzymes involved in the phosphoketolase and glycolytic pathways. Expression of enzymes involved in pyruvate and glycerol/glycerolipid metabolism were also affected by the change of carbon source. Interestingly, a commercial starter culture and a protective culture strain down-regulated the glycolytic pathway more efficiently than the rest of the strains when grown on ribose. The overall two-dimensional gel electrophoresis (2-DE protein expression pattern was similar for the different strains, though distinct differences were seen between the two subspecies (sakei and carnosus, and a variation of about 20% in the number of spots in the 2-DE gels was observed between strains. A strain isolated from fermented fish showed a higher expression of stress related proteins growing on both carbon sources. Conclusions It is obvious from the data obtained in this study that the proteomic approach efficiently identifies differentially expressed proteins caused by the change of carbon source. Despite the basic similarity in the strains metabolic routes when they ferment glucose and ribose, there were also interesting differences. From the application point of view, an understanding of regulatory

  12. Experimental design and data-analysis in label-free quantitative LC/MS proteomics: A tutorial with MSqRob.

    Science.gov (United States)

    Goeminne, Ludger J E; Gevaert, Kris; Clement, Lieven

    2018-01-16

    Label-free shotgun proteomics is routinely used to assess proteomes. However, extracting relevant information from the massive amounts of generated data remains difficult. This tutorial provides a strong foundation on analysis of quantitative proteomics data. We provide key statistical concepts that help researchers to design proteomics experiments and we showcase how to analyze quantitative proteomics data using our recent free and open-source R package MSqRob, which was developed to implement the peptide-level robust ridge regression method for relative protein quantification described by Goeminne et al. MSqRob can handle virtually any experimental proteomics design and outputs proteins ordered by statistical significance. Moreover, its graphical user interface and interactive diagnostic plots provide easy inspection and also detection of anomalies in the data and flaws in the data analysis, allowing deeper assessment of the validity of results and a critical review of the experimental design. Our tutorial discusses interactive preprocessing, data analysis and visualization of label-free MS-based quantitative proteomics experiments with simple and more complex designs. We provide well-documented scripts to run analyses in bash mode on GitHub, enabling the integration of MSqRob in automated pipelines on cluster environments (https://github.com/statOmics/MSqRob). The concepts outlined in this tutorial aid in designing better experiments and analyzing the resulting data more appropriately. The two case studies using the MSqRob graphical user interface will contribute to a wider adaptation of advanced peptide-based models, resulting in higher quality data analysis workflows and more reproducible results in the proteomics community. We also provide well-documented scripts for experienced users that aim at automating MSqRob on cluster environments. Copyright © 2017 Elsevier B.V. All rights reserved.

  13. Proteomics-based network analysis characterizes biological processes and pathways activated by preconditioned mesenchymal stem cells in cardiac repair mechanisms.

    Science.gov (United States)

    Di Silvestre, Dario; Brambilla, Francesca; Scardoni, Giovanni; Brunetti, Pietro; Motta, Sara; Matteucci, Marco; Laudanna, Carlo; Recchia, Fabio A; Lionetti, Vincenzo; Mauri, Pierluigi

    2017-05-01

    We have demonstrated that intramyocardial delivery of human mesenchymal stem cells preconditioned with a hyaluronan mixed ester of butyric and retinoic acid (MSCp + ) is more effective in preventing the decay of regional myocardial contractility in a swine model of myocardial infarction (MI). However, the understanding of the role of MSCp + in proteomic remodeling of cardiac infarcted tissue is not complete. We therefore sought to perform a comprehensive analysis of the proteome of infarct remote (RZ) and border zone (BZ) of pigs treated with MSCp + or unconditioned stem cells. Heart tissues were analyzed by MudPIT and differentially expressed proteins were selected by a label-free approach based on spectral counting. Protein profiles were evaluated by using PPI networks and their topological analysis. The proteomic remodeling was largely prevented in MSCp + group. Extracellular proteins involved in fibrosis were down-regulated, while energetic pathways were globally up-regulated. Cardioprotectant pathways involved in the production of keto acid metabolites were also activated. Additionally, we found that new hub proteins support the cardioprotective phenotype characterizing the left ventricular BZ treated with MSCp + . In fact, the up-regulation of angiogenic proteins NCL and RAC1 can be explained by the increase of capillary density induced by MSCp + . Our results show that angiogenic pathways appear to be uniquely positioned to integrate signaling with energetic pathways involving cardiac repair. Our findings prompt the use of proteomics-based network analysis to optimize new approaches preventing the post-ischemic proteomic remodeling that may underlie the limited self-repair ability of adult heart. Copyright © 2017 Elsevier B.V. All rights reserved.

  14. Stressor-induced proteome alterations in zebrafish: A meta-analysis of response patterns

    Energy Technology Data Exchange (ETDEWEB)

    Groh, Ksenia J., E-mail: ksenia.groh@eawag.ch [Eawag, Swiss Federal Institute of Aquatic Science and Technology, 8600 Dübendorf (Switzerland); ETH Zürich, Swiss Federal Institute of Technology, Department of Chemistry and Applied Biosciences, 8093 Zürich (Switzerland); Suter, Marc J.-F. [Eawag, Swiss Federal Institute of Aquatic Science and Technology, 8600 Dübendorf (Switzerland); ETH Zürich, Swiss Federal Institute of Technology, Department of Environmental Systems Science, 8092 Zürich (Switzerland)

    2015-02-15

    Highlights: • We compared reported proteome changes induced by various stressors in zebrafish. • Several proteins groups frequently responding to diverse stressors were identified. • These included energy metabolism enzymes, heat shock and cytoskeletal proteins. • Insufficient proteome coverage impedes identification of more specific responses. • Further research needs for proteomics in ecotoxicology are discussed. - Abstract: Proteomics approaches are being increasingly applied in ecotoxicology on the premise that the identification of specific protein expression changes in response to a particular chemical would allow elucidation of the underlying molecular pathways leading to an adverse effect. This in turn is expected to promote the development of focused testing strategies for specific groups of toxicants. Although both gel-based and gel-free global characterization techniques provide limited proteome coverage, the conclusions regarding the cellular processes affected are still being drawn based on the few changes detected. To investigate how specific the detected responses are, we analyzed a set of studies that characterized proteome alterations induced by various physiological, chemical and biological stressors in zebrafish, a popular model organism. Our analysis highlights several proteins and protein groups, including heat shock and oxidative stress defense proteins, energy metabolism enzymes and cytoskeletal proteins, to be most frequently identified as responding to diverse stressors. In contrast, other potentially more specifically responding protein groups are detected much less frequently. Thus, zebrafish proteome responses to stress reported by different studies appear to depend mostly on the level of stress rather than on the specific stressor itself. This suggests that the most broadly used current proteomics technologies do not provide sufficient proteome coverage to allow in-depth investigation of specific mechanisms of toxicant action

  15. Stressor-induced proteome alterations in zebrafish: A meta-analysis of response patterns

    International Nuclear Information System (INIS)

    Groh, Ksenia J.; Suter, Marc J.-F.

    2015-01-01

    Highlights: • We compared reported proteome changes induced by various stressors in zebrafish. • Several proteins groups frequently responding to diverse stressors were identified. • These included energy metabolism enzymes, heat shock and cytoskeletal proteins. • Insufficient proteome coverage impedes identification of more specific responses. • Further research needs for proteomics in ecotoxicology are discussed. - Abstract: Proteomics approaches are being increasingly applied in ecotoxicology on the premise that the identification of specific protein expression changes in response to a particular chemical would allow elucidation of the underlying molecular pathways leading to an adverse effect. This in turn is expected to promote the development of focused testing strategies for specific groups of toxicants. Although both gel-based and gel-free global characterization techniques provide limited proteome coverage, the conclusions regarding the cellular processes affected are still being drawn based on the few changes detected. To investigate how specific the detected responses are, we analyzed a set of studies that characterized proteome alterations induced by various physiological, chemical and biological stressors in zebrafish, a popular model organism. Our analysis highlights several proteins and protein groups, including heat shock and oxidative stress defense proteins, energy metabolism enzymes and cytoskeletal proteins, to be most frequently identified as responding to diverse stressors. In contrast, other potentially more specifically responding protein groups are detected much less frequently. Thus, zebrafish proteome responses to stress reported by different studies appear to depend mostly on the level of stress rather than on the specific stressor itself. This suggests that the most broadly used current proteomics technologies do not provide sufficient proteome coverage to allow in-depth investigation of specific mechanisms of toxicant action

  16. State-of-the-art nanoplatform-integrated MALDI-MS impacting resolutions in urinary proteomics.

    Science.gov (United States)

    Gopal, Judy; Muthu, Manikandan; Chun, Se-Chul; Wu, Hui-Fen

    2015-06-01

    Urine proteomics has become a subject of interest, since it has led to a number of breakthroughs in disease diagnostics. Urine contains information not only from the kidney and the urinary tract but also from other organs, thus urinary proteome analysis allows for identification of biomarkers for both urogenital and systemic diseases. The following review gives a brief overview of the analytical techniques that have been in practice for urinary proteomics. MALDI-MS technique and its current application status in this area of clinical research have been discussed. The review comments on the challenges facing the conventional MALDI-MS technique and the upgradation of this technique with the introduction of nanotechnology. This review projects nano-based techniques such as nano-MALDI-MS, surface-assisted laser desorption/ionization, and nanostructure-initiator MS as the platforms that have the potential in trafficking MALDI-MS from the lab to the bedside. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  17. Differential proteomic analysis reveals novel links between primary metabolism and antibiotic production in Amycolatopsis balhimycina

    DEFF Research Database (Denmark)

    Gallo, G.; Renzone, G.; Alduina, R.

    2010-01-01

    A differential proteomic analysis, based on 2-DE and MS procedures, was performed on Amycolatopsis balhimycina DSM5908, the actinomycete producing the vancomycin-like antibiotic balhimycin. A comparison of proteomic profiles before and during balhimycin production characterized differentially...... available over the World Wide Web as interactive web pages (http://www.unipa.it/ampuglia/Abal-proteome-maps). Functional clustering analysis revealed that differentially expressed proteins belong to functional groups involved in central carbon metabolism, amino acid metabolism and protein biosynthesis...... intermediates, were upregulated during antibiotic production. qRT-PCR analysis revealed that 8 out of 14 upregulated genes showed a positive correlation between changes at translational and transcriptional expression level. Furthermore, proteomic analysis of two nonproducing mutants, restricted to a sub...

  18. The analysis of proteome changes in sunflower seeds induced by N ...

    Indian Academy of Sciences (India)

    Madhu

    Key Laboratory of Photosynthesis and Environmental Molecular Physiology, Institute of Botany, Chinese ... ion, a proteomic analysis of N+ ion implantation seeds was developed. ... implantation were validated in course of plant development,.

  19. Non-synonymous variations in cancer and their effects on the human proteome: workflow for NGS data biocuration and proteome-wide analysis of TCGA data.

    Science.gov (United States)

    Cole, Charles; Krampis, Konstantinos; Karagiannis, Konstantinos; Almeida, Jonas S; Faison, William J; Motwani, Mona; Wan, Quan; Golikov, Anton; Pan, Yang; Simonyan, Vahan; Mazumder, Raja

    2014-01-27

    Next-generation sequencing (NGS) technologies have resulted in petabytes of scattered data, decentralized in archives, databases and sometimes in isolated hard-disks which are inaccessible for browsing and analysis. It is expected that curated secondary databases will help organize some of this Big Data thereby allowing users better navigate, search and compute on it. To address the above challenge, we have implemented a NGS biocuration workflow and are analyzing short read sequences and associated metadata from cancer patients to better understand the human variome. Curation of variation and other related information from control (normal tissue) and case (tumor) samples will provide comprehensive background information that can be used in genomic medicine research and application studies. Our approach includes a CloudBioLinux Virtual Machine which is used upstream of an integrated High-performance Integrated Virtual Environment (HIVE) that encapsulates Curated Short Read archive (CSR) and a proteome-wide variation effect analysis tool (SNVDis). As a proof-of-concept, we have curated and analyzed control and case breast cancer datasets from the NCI cancer genomics program - The Cancer Genome Atlas (TCGA). Our efforts include reviewing and recording in CSR available clinical information on patients, mapping of the reads to the reference followed by identification of non-synonymous Single Nucleotide Variations (nsSNVs) and integrating the data with tools that allow analysis of effect nsSNVs on the human proteome. Furthermore, we have also developed a novel phylogenetic analysis algorithm that uses SNV positions and can be used to classify the patient population. The workflow described here lays the foundation for analysis of short read sequence data to identify rare and novel SNVs that are not present in dbSNP and therefore provides a more comprehensive understanding of the human variome. Variation results for single genes as well as the entire study are available

  20. Proteome analysis of the almond kernel (Prunus dulcis).

    Science.gov (United States)

    Li, Shugang; Geng, Fang; Wang, Ping; Lu, Jiankang; Ma, Meihu

    2016-08-01

    Almond (Prunus dulcis) is a popular tree nut worldwide and offers many benefits to human health. However, the importance of almond kernel proteins in the nutrition and function in human health requires further evaluation. The present study presents a systematic evaluation of the proteins in the almond kernel using proteomic analysis. The nutrient and amino acid content in almond kernels from Xinjiang is similar to that of American varieties; however, Xinjiang varieties have a higher protein content. Two-dimensional electrophoresis analysis demonstrated a wide distribution of molecular weights and isoelectric points of almond kernel proteins. A total of 434 proteins were identified by LC-MS/MS, and most were proteins that were experimentally confirmed for the first time. Gene ontology (GO) analysis of the 434 proteins indicated that proteins involved in primary biological processes including metabolic processes (67.5%), cellular processes (54.1%), and single-organism processes (43.4%), the main molecular function of almond kernel proteins are in catalytic activity (48.0%), binding (45.4%) and structural molecule activity (11.9%), and proteins are primarily distributed in cell (59.9%), organelle (44.9%), and membrane (22.8%). Almond kernel is a source of a wide variety of proteins. This study provides important information contributing to the screening and identification of almond proteins, the understanding of almond protein function, and the development of almond protein products. © 2015 Society of Chemical Industry. © 2015 Society of Chemical Industry.

  1. Proteomic analysis of three gonad types of swamp eel reveals genes differentially expressed during sex reversal

    OpenAIRE

    Sheng, Yue; Zhao, Wei; Song, Ying; Li, Zhigang; Luo, Majing; Lei, Quan; Cheng, Hanhua; Zhou, Rongjia

    2015-01-01

    A variety of mechanisms are engaged in sex determination in vertebrates. The teleost fish swamp eel undergoes sex reversal naturally and is an ideal model for vertebrate sexual development. However, the importance of proteome-wide scanning for gonad reversal was not previously determined. We report a 2-D electrophoresis analysis of three gonad types of proteomes during sex reversal. MS/MS analysis revealed a group of differentially expressed proteins during ovary to ovotestis to testis transf...

  2. Proteomic analysis of minute amount of colonic biopsies by enteroscopy sampling

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Xing [Department of Analytical Chemistry and CAS Key Laboratory of Receptor Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences (China); Xu, Yanli [Fuyang People’s Hospital (China); Meng, Qian [Department of Analytical Chemistry and CAS Key Laboratory of Receptor Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences (China); Zheng, Qingqing [Digestive Endoscopic Center, Shanghai Jiaotong University Affiliated Sixth People’s Hospital (China); Wu, Jianhong [Department of Analytical Chemistry and CAS Key Laboratory of Receptor Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences (China); Wang, Chen; Jia, Weiping [Shanghai Key Laboratory of Diabetes Mellitus, Department of Endocrinology and Metabolism, Shanghai Diabetes Institute, Shanghai Clinical Center for Diabetes, Shanghai Jiao Tong University Affiliated Sixth People’s Hospital (China); Figeys, Daniel [Department of Biochemistry, Microbiology and Immunology, and Department of Chemistry and Biomolecular Sciences, University of Ottawa (Canada); Chang, Ying, E-mail: emulan@163.com [Digestive Endoscopic Center, Shanghai Jiaotong University Affiliated Sixth People’s Hospital (China); Zhou, Hu, E-mail: zhouhu@simm.ac.cn [Department of Analytical Chemistry and CAS Key Laboratory of Receptor Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences (China)

    2016-08-05

    Colorectal cancer (CRC) is one of the most common types of malignant tumor worldwide. Currently, although many researchers have been devoting themselves in CRC studies, the process of locating biomarkers for CRC early diagnosis and prognostic is still very slow. Using a centrifugal proteomic reactor-based proteomic analysis of minute amount of colonic biopsies by enteroscopy sampling, 2620 protein groups were quantified between cancer mucosa and adjacent normal colorectal mucosa. Of which, 403 protein groups were differentially expressed with statistic significance between cancer and normal tissues, including 195 up-regulated and 208 down-regulated proteins in cancer tissues. Three proteins (SOD3, PRELP and NGAL) were selected for further Western blot validation. And the resulting Western blot experimental results were consistent with the quantitative proteomic data. SOD3 and PRELP are down-regulated in CRC mucosa comparing to adjacent normal tissue, while NGAL is up-regulated in CRC mucosa. In conclusion, the centrifugal proteomic reactor-based label-free quantitative proteomic approach provides a highly sensitive and powerful tool for analyzing minute protein sample from tiny colorectal biopsies, which may facilitate CRC biomarkers discovery for diagnoses and prognoses. -- Highlights: •Minute amount of colonic biopsies by endoscopy is suitable for proteomic analysis. •Centrifugal proteomic reactor can be used for processing tiny clinic biopsy sample. •SOD3 and PRELP are down-regulated in CRC, while NGAL is up-regulated in CRC.

  3. Proteomic analysis of minute amount of colonic biopsies by enteroscopy sampling

    International Nuclear Information System (INIS)

    Liu, Xing; Xu, Yanli; Meng, Qian; Zheng, Qingqing; Wu, Jianhong; Wang, Chen; Jia, Weiping; Figeys, Daniel; Chang, Ying; Zhou, Hu

    2016-01-01

    Colorectal cancer (CRC) is one of the most common types of malignant tumor worldwide. Currently, although many researchers have been devoting themselves in CRC studies, the process of locating biomarkers for CRC early diagnosis and prognostic is still very slow. Using a centrifugal proteomic reactor-based proteomic analysis of minute amount of colonic biopsies by enteroscopy sampling, 2620 protein groups were quantified between cancer mucosa and adjacent normal colorectal mucosa. Of which, 403 protein groups were differentially expressed with statistic significance between cancer and normal tissues, including 195 up-regulated and 208 down-regulated proteins in cancer tissues. Three proteins (SOD3, PRELP and NGAL) were selected for further Western blot validation. And the resulting Western blot experimental results were consistent with the quantitative proteomic data. SOD3 and PRELP are down-regulated in CRC mucosa comparing to adjacent normal tissue, while NGAL is up-regulated in CRC mucosa. In conclusion, the centrifugal proteomic reactor-based label-free quantitative proteomic approach provides a highly sensitive and powerful tool for analyzing minute protein sample from tiny colorectal biopsies, which may facilitate CRC biomarkers discovery for diagnoses and prognoses. -- Highlights: •Minute amount of colonic biopsies by endoscopy is suitable for proteomic analysis. •Centrifugal proteomic reactor can be used for processing tiny clinic biopsy sample. •SOD3 and PRELP are down-regulated in CRC, while NGAL is up-regulated in CRC.

  4. Evaluation of sample extraction methods for proteomics analysis of green algae Chlorella vulgaris.

    Science.gov (United States)

    Gao, Yan; Lim, Teck Kwang; Lin, Qingsong; Li, Sam Fong Yau

    2016-05-01

    Many protein extraction methods have been developed for plant proteome analysis but information is limited on the optimal protein extraction method from algae species. This study evaluated four protein extraction methods, i.e. direct lysis buffer method, TCA-acetone method, phenol method, and phenol/TCA-acetone method, using green algae Chlorella vulgaris for proteome analysis. The data presented showed that phenol/TCA-acetone method was superior to the other three tested methods with regards to shotgun proteomics. Proteins identified using shotgun proteomics were validated using sequential window acquisition of all theoretical fragment-ion spectra (SWATH) technique. Additionally, SWATH provides protein quantitation information from different methods and protein abundance using different protein extraction methods was evaluated. These results highlight the importance of green algae protein extraction method for subsequent MS analysis and identification. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  5. Proteomic biomarkers for ovarian cancer risk in women with polycystic ovary syndrome: a systematic review and biomarker database integration.

    Science.gov (United States)

    Galazis, Nicolas; Olaleye, Olalekan; Haoula, Zeina; Layfield, Robert; Atiomo, William

    2012-12-01

    To review and identify possible biomarkers for ovarian cancer (OC) in women with polycystic ovary syndrome (PCOS). Systematic literature searches of MEDLINE, EMBASE, and Cochrane using the search terms "proteomics," "proteomic," and "ovarian cancer" or "ovarian carcinoma." Proteomic biomarkers for OC were then integrated with an updated previously published database of all proteomic biomarkers identified to date in patients with PCOS. Academic department of obstetrics and gynecology in the United Kingdom. A total of 180 women identified in the six studies. Tissue samples from women with OC vs. tissue samples from women without OC. Proteomic biomarkers, proteomic technique used, and methodologic quality score. A panel of six biomarkers was overexpressed both in women with OC and in women with PCOS. These biomarkers include calreticulin, fibrinogen-γ, superoxide dismutase, vimentin, malate dehydrogenase, and lamin B2. These biomarkers could help improve our understanding of the links between PCOS and OC and could potentially be used to identify subgroups of women with PCOS at increased risk of OC. More studies are required to further evaluate the role these biomarkers play in women with PCOS and OC. Copyright © 2012 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

  6. Quantitative and temporal proteome analysis of butyrate-treated colorectal cancer cells.

    Science.gov (United States)

    Tan, Hwee Tong; Tan, Sandra; Lin, Qingsong; Lim, Teck Kwang; Hew, Choy Leong; Chung, Maxey C M

    2008-06-01

    Colorectal cancer is one of the most common cancers in developed countries, and its incidence is negatively associated with high dietary fiber intake. Butyrate, a short-chain fatty acid fermentation by-product of fiber induces cell maturation with the promotion of growth arrest, differentiation, and/or apoptosis of cancer cells. The stimulation of cell maturation by butyrate in colonic cancer cells follows a temporal progression from the early phase of growth arrest to the activation of apoptotic cascades. Previously we performed two-dimensional DIGE to identify differentially expressed proteins induced by 24-h butyrate treatment of HCT-116 colorectal cancer cells. Herein we used quantitative proteomics approaches using iTRAQ (isobaric tags for relative and absolute quantitation), a stable isotope labeling methodology that enables multiplexing of four samples, for a temporal study of HCT-116 cells treated with butyrate. In addition, cleavable ICAT, which selectively tags cysteine-containing proteins, was also used, and the results complemented those obtained from the iTRAQ strategy. Selected protein targets were validated by real time PCR and Western blotting. A model is proposed to illustrate our findings from this temporal analysis of the butyrate-responsive proteome that uncovered several integrated cellular processes and pathways involved in growth arrest, apoptosis, and metastasis. These signature clusters of butyrate-regulated pathways are potential targets for novel chemopreventive and therapeutic drugs for treatment of colorectal cancer.

  7. A proteomic analysis of the functional effects of fatty acids in NIH 3T3 fibroblasts

    LENUS (Irish Health Repository)

    Magdalon, Juliana

    2011-11-24

    Abstract Previous studies have demonstrated that long chain fatty acids influence fibroblast function at sub-lethal concentrations. This study is the first to assess the effects of oleic, linoleic or palmitic acids on protein expression of fibroblasts, as determined by standard proteomic techniques. The fatty acids were not cytotoxic at the concentration used in this work as assessed by membrane integrity, DNA fragmentation and the MTT assay but significantly increased cell proliferation. Subsequently, a proteomic analysis was performed using two dimensional difference gel electrophoresis (2D-DIGE) and MS based identification. Cells treated with 50 μM oleic, linoleic or palmitic acid for 24 h were associated with 24, 22, 16 spots differentially expressed, respectively. Among the identified proteins, α-enolase and far upstream element binding protein 1 (FBP-1) are of importance due to their function in fibroblast-associated diseases. However, modulation of α-enolase and FBP-1 expression by fatty acids was not validated by the Western blot technique.

  8. Analysis of the biofilm proteome of Xylella fastidiosa

    Directory of Open Access Journals (Sweden)

    Labate Carlos A

    2011-09-01

    Full Text Available Abstract Background Xylella fastidiosa is limited to the xylem of the plant host and the foregut of insect vectors (sharpshooters. The mechanism of pathogenicity of this bacterium differs from other plant pathogens, since it does not present typical genes that confer specific interactions between plant and pathogens (avr and/or hrp. The bacterium is injected directly into the xylem vessels where it adheres and colonizes. The whole process leads to the formation of biofilms, which are considered the main mechanism of pathogenicity. Cells in biofilms are metabolically and phenotypically different from their planktonic condition. The mature biofilm stage (phase of higher cell density presents high virulence and resistance to toxic substances such as antibiotics and detergents. Here we performed proteomic analysis of proteins expressed exclusively in the mature biofilm of X. fastidiosa strain 9a5c, in comparison to planktonic growth condition. Results We found a total of 456 proteins expressed in the biofilm condition, which correspond to approximately 10% of total protein in the genome. The biofilm showed 37% (or 144 proteins different protein than we found in the planktonic growth condition. The large difference in protein pattern in the biofilm condition may be responsible for the physiological changes of the cells in the biofilm of X. fastidiosa. Mass spectrometry was used to identify these proteins, while real-time quantitative polymerase chain reaction monitored expression of genes encoding them. Most of proteins expressed in the mature biofilm growth were associated with metabolism, adhesion, pathogenicity and stress conditions. Even though the biofilm cells in this work were not submitted to any stress condition, some stress related proteins were expressed only in the biofilm condition, suggesting that the biofilm cells would constitutively express proteins in different adverse environments. Conclusions We observed overexpression of proteins

  9. Comparative proteomic analysis of ductal and lobular invasive breast carcinoma.

    Science.gov (United States)

    Oliveira, N C S; Gomig, T H B; Milioli, H H; Cordeiro, F; Costa, G G; Urban, C A; Lima, R S; Cavalli, I J; Ribeiro, E M S F

    2016-04-04

    Breast cancer is the second most common cancer worldwide and the first among women. Invasive ductal carcinoma (IDC) and invasive lobular carcinoma (ILC) are the two major histological subtypes, and the clinical and molecular differences between them justify the search for new markers to distinguish them. As proteomic analysis allows for a powerful and analytical approach to identify potential biomarkers, we performed a comparative analysis of IDC and ILC samples by using two-dimensional electrophoresis and mass spectrometry. Twenty-three spots were identified corresponding to 10 proteins differentially expressed between the two subtypes. ACTB, ACTG, TPM3, TBA1A, TBA1B, VIME, TPIS, PDIA3, PDIA6, and VTDB were upregulated in ductal carcinoma compared to in lobular carcinoma samples. Overall, these 10 proteins have a key role in oncogenesis. Their specific functions and relevance in cancer initiation and progression are further discussed in this study. The identified peptides represent promising biomarkers for the differentiation of ductal and lobular breast cancer subtypes, and for future interventions based on tailored therapy.

  10. Proteomic analysis of honeybee worker (Apis mellifera hypopharyngeal gland development

    Directory of Open Access Journals (Sweden)

    Li Jianke

    2009-12-01

    Full Text Available Abstract Background Hypopharyngeal glands (HG of honeybee workers play an important role in honeybee nutrition and caste differentiation. Previous research mainly focused on age-dependent morphological, physiological, biochemical and genomic characters of the HG. Here proteomics and biochemical network analysis were used to follow protein changes during the HG development. Results A total of 87, 76, 85, 74, 71, and 55 proteins were unambiguously identified on day 1, 3, 6, 12, 15 and 20, respectively. These proteins were major royal jelly proteins (MRJPs, metabolism of carbohydrates, lipids and proteins, cytoskeleton, development regulation, antioxidant, molecule transporter, regulation of transcription/translation, proteins with folding functions. The most interesting is that MRJP's that have been detected in the HG of the newly emerged worker bees. The MRJP's expression is at peak level from 6-12 days, was validated by western blot analysis of MRJP1, 2 and 3. Moreover, 35 key node proteins were found in the biochemical networks of the HG. Conclusions HG secretes RJ at peak level within 6-12 days, but the worker bee can secrete royal jelly (RJ since birth, which is a new finding. Several key node proteins play an important role in the biochemical networks of the developing HG. This provides us some target proteins when genetically manipulating honeybees.

  11. Proteomic analysis of the fish pathogen Flavobacterium columnare

    Directory of Open Access Journals (Sweden)

    Lawrence Mark L

    2010-06-01

    Full Text Available Abstract Background Flavobacterium columnare causes columnaris disease in cultured and wild fish populations worldwide. Columnaris is the second most prevalent bacterial disease of commercial channel catfish industry in the United States. Despite its economic importance, little is known about the expressed proteins and virulence mechanisms of F. columnare. Here, we report the first high throughput proteomic analysis of F. columnare using 2-D LC ESI MS/MS and 2-DE MALDI TOF/TOF MS. Results Proteins identified in this study and predicted from the draft F. columnare genome were clustered into functional groups using clusters of orthologous groups (COGs, and their subcellular locations were predicted. Possible functional relations among the identified proteins were determined using pathway analysis. The total number of unique F. columnare proteins identified using both 2-D LC and 2-DE approaches was 621, of which 10.95% (68 were identified by both methods, while 77.29% (480 and 11.76% (73 were unique in 2-D LC and 2-DE, respectively. COG groupings and subcellular localizations were similar between our data set and proteins predicted from the whole genome. Twenty eight pathways were significantly represented in our dataset (P Conclusion Results from this study provide experimental evidence for many proteins that were predicted from the F. columnare genome annotation, and they should accelerate functional and comparative studies aimed at understanding virulence mechanisms of this important pathogen.

  12. Functional complexity of the axonal growth cone: a proteomic analysis.

    Directory of Open Access Journals (Sweden)

    Adriana Estrada-Bernal

    Full Text Available The growth cone, the tip of the emerging neurite, plays a crucial role in establishing the wiring of the developing nervous system. We performed an extensive proteomic analysis of axonal growth cones isolated from the brains of fetal Sprague-Dawley rats. Approximately 2000 proteins were identified at ≥ 99% confidence level. Using informatics, including functional annotation cluster and KEGG pathway analysis, we found great diversity of proteins involved in axonal pathfinding, cytoskeletal remodeling, vesicular traffic and carbohydrate metabolism, as expected. We also found a large and complex array of proteins involved in translation, protein folding, posttranslational processing, and proteasome/ubiquitination-dependent degradation. Immunofluorescence studies performed on hippocampal neurons in culture confirmed the presence in the axonal growth cone of proteins representative of these processes. These analyses also provide evidence for rough endoplasmic reticulum and reveal a reticular structure equipped with Golgi-like functions in the axonal growth cone. Furthermore, Western blot revealed the growth cone enrichment, relative to fetal brain homogenate, of some of the proteins involved in protein synthesis, folding and catabolism. Our study provides a resource for further research and amplifies the relatively recently developed concept that the axonal growth cone is equipped with proteins capable of performing a highly diverse range of functions.

  13. Quantitative proteomics analysis using 2D-PAGE to investigate the effects of cigarette smoke and aerosol of a prototypic modified risk tobacco product on the lung proteome in C57BL/6 mice.

    Science.gov (United States)

    Elamin, Ashraf; Titz, Bjoern; Dijon, Sophie; Merg, Celine; Geertz, Marcel; Schneider, Thomas; Martin, Florian; Schlage, Walter K; Frentzel, Stefan; Talamo, Fabio; Phillips, Blaine; Veljkovic, Emilija; Ivanov, Nikolai V; Vanscheeuwijck, Patrick; Peitsch, Manuel C; Hoeng, Julia

    2016-08-11

    Smoking is associated with several serious diseases, such as lung cancer and chronic obstructive pulmonary disease (COPD). Within our systems toxicology framework, we are assessing whether potential modified risk tobacco products (MRTP) can reduce smoking-related health risks compared to conventional cigarettes. In this article, we evaluated to what extent 2D-PAGE/MALDI MS/MS (2D-PAGE) can complement the iTRAQ LC-MS/MS results from a previously reported mouse inhalation study, in which we assessed a prototypic MRTP (pMRTP). Selected differentially expressed proteins identified by both LC-MS/MS and 2D-PAGE approaches were further verified using reverse-phase protein microarrays. LC-MS/MS captured the effects of cigarette smoke (CS) on the lung proteome more comprehensively than 2D-PAGE. However, an integrated analysis of both proteomics data sets showed that 2D-PAGE data complement the LC-MS/MS results by supporting the overall trend of lower effects of pMRTP aerosol than CS on the lung proteome. Biological effects of CS exposure supported by both methods included increases in immune-related, surfactant metabolism, proteasome, and actin cytoskeleton protein clusters. Overall, while 2D-PAGE has its value, especially as a complementary method for the analysis of effects on intact proteins, LC-MS/MS approaches will likely be the method of choice for proteome analysis in systems toxicology investigations. Quantitative proteomics is anticipated to play a growing role within systems toxicology assessment frameworks in the future. To further understand how different proteomics technologies can contribute to toxicity assessment, we conducted a quantitative proteomics analysis using 2D-PAGE and isobaric tag-based LC-MS/MS approaches and compared the results produced from the 2 approaches. Using a prototypic modified risk tobacco product (pMRTP) as our test item, we show compared with cigarette smoke, how 2D-PAGE results can complement and support LC-MS/MS data, demonstrating

  14. Proteome reference map of Drosophila melanogaster head.

    Science.gov (United States)

    Lee, Tian-Ren; Huang, Shun-Hong; Lee, Chi-Ching; Lee, Hsiao-Yun; Chan, Hsin-Tzu; Lin, Kuo-Sen; Chan, Hong-Lin; Lyu, Ping-Chiang

    2012-06-01

    Drosophila melanogaster has been used as a genetic model organism to understand the fundamental molecular mechanisms in human biology including memory formation that has been reported involving protein synthesis and/or post-translational modification. In this study, we employed a proteomic platform based on fluorescent 2DE and MALDI-TOF MS to build a standard D. melanogaster head proteome map for proteome-proteome comparison. In order to facilitate the comparison, an interactive database has been constructed for systematically integrating and analyzing the proteomes from different conditions and further implicated to study human diseases related to D. melanogaster model. In summary, the fundamental head proteomic database and bioinformatic analysis will be useful for further elucidating the biological mechanisms such as memory formation and neurodegenerative diseases. © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  15. MASPECTRAS: a platform for management and analysis of proteomics LC-MS/MS data

    Directory of Open Access Journals (Sweden)

    Rader Robert

    2007-06-01

    Full Text Available Abstract Background The advancements of proteomics technologies have led to a rapid increase in the number, size and rate at which datasets are generated. Managing and extracting valuable information from such datasets requires the use of data management platforms and computational approaches. Results We have developed the MAss SPECTRometry Analysis System (MASPECTRAS, a platform for management and analysis of proteomics LC-MS/MS data. MASPECTRAS is based on the Proteome Experimental Data Repository (PEDRo relational database schema and follows the guidelines of the Proteomics Standards Initiative (PSI. Analysis modules include: 1 import and parsing of the results from the search engines SEQUEST, Mascot, Spectrum Mill, X! Tandem, and OMSSA; 2 peptide validation, 3 clustering of proteins based on Markov Clustering and multiple alignments; and 4 quantification using the Automated Statistical Analysis of Protein Abundance Ratios algorithm (ASAPRatio. The system provides customizable data retrieval and visualization tools, as well as export to PRoteomics IDEntifications public repository (PRIDE. MASPECTRAS is freely available at http://genome.tugraz.at/maspectras Conclusion Given the unique features and the flexibility due to the use of standard software technology, our platform represents significant advance and could be of great interest to the proteomics community.

  16. Proteomic analysis of Sydney Rock oysters (Saccostrea glomerata) exposed to metal contamination in the field

    International Nuclear Information System (INIS)

    Thompson, Emma L.; Taylor, Daisy A.; Nair, Sham V.; Birch, Gavin; Hose, Grant C.; Raftos, David A.

    2012-01-01

    This study used proteomics to assess the impacts of metal contamination in the field on Sydney Rock oysters. Oysters were transplanted into Lake Macquarie, NSW, for two weeks in both 2009 and 2010. Two-dimensional electrophoresis identified changes in protein expression profiles of oyster haemolymph between control and metal contaminated sites. There were unique protein expression profiles for each field trial. Principal components analysis attributed these differences in oyster proteomes to the different combinations and concentrations of metals and other environmental variables present during the three field trials. Identification of differentially expressed proteins showed that proteins associated with cytoskeletal activity and stress responses were the most commonly affected biological functions in the Sydney Rock oyster. Overall, the data show that proteomics combined with multivariate analysis has the potential to link the effects of contaminants with biological consequences. - Highlights: ► Sydney Rock oyster haemolymph was analysed by proteomics after metal exposure in 3 field trials. ► 2-DE analysis was used to compare protein profiles between control and contaminated sites. ► Different protein expression profiles were revealed per field trial. ► Principal components analysis attributed profiles to different suites of metals and environmental variables per trial. ► The study highlights the need to do multiple field trials and to combine proteomic and enviro. data. - This study used proteomics to analyse impacts of metal contamination on Sydney Rock oyster (Saccostrea glomerata) haemolymph in multiple field trials.

  17. MUMAL: Multivariate analysis in shotgun proteomics using machine learning techniques

    Directory of Open Access Journals (Sweden)

    Cerqueira Fabio R

    2012-10-01

    Full Text Available Abstract Background The shotgun strategy (liquid chromatography coupled with tandem mass spectrometry is widely applied for identification of proteins in complex mixtures. This method gives rise to thousands of spectra in a single run, which are interpreted by computational tools. Such tools normally use a protein database from which peptide sequences are extracted for matching with experimentally derived mass spectral data. After the database search, the correctness of obtained peptide-spectrum matches (PSMs needs to be evaluated also by algorithms, as a manual curation of these huge datasets would be impractical. The target-decoy database strategy is largely used to perform spectrum evaluation. Nonetheless, this method has been applied without considering sensitivity, i.e., only error estimation is taken into account. A recently proposed method termed MUDE treats the target-decoy analysis as an optimization problem, where sensitivity is maximized. This method demonstrates a significant increase in the retrieved number of PSMs for a fixed error rate. However, the MUDE model is constructed in such a way that linear decision boundaries are established to separate correct from incorrect PSMs. Besides, the described heuristic for solving the optimization problem has to be executed many times to achieve a significant augmentation in sensitivity. Results Here, we propose a new method, termed MUMAL, for PSM assessment that is based on machine learning techniques. Our method can establish nonlinear decision boundaries, leading to a higher chance to retrieve more true positives. Furthermore, we need few iterations to achieve high sensitivities, strikingly shortening the running time of the whole process. Experiments show that our method achieves a considerably higher number of PSMs compared with standard tools such as MUDE, PeptideProphet, and typical target-decoy approaches. Conclusion Our approach not only enhances the computational performance, and

  18. InSilico Proteomics System: Integration and Application of Protein and Protein-Protein Interaction Data using Microsoft .NET

    Directory of Open Access Journals (Sweden)

    Straßer Wolfgang

    2006-12-01

    Full Text Available In the last decades, biological databases became the major knowledge resource for researchers in the field of molecular biology. The distribution of information among these databases is one of the major problems. An overview about the subject area of data access and representation of protein and protein-protein interaction data within public biological databases is described. For a comprehensive and consistent way of searching and analysing integrated protein and protein-protein interaction data, the InSilico Proteomics (ISP project has been initiated. Its three main objectives are (1 to provide an integrated knowledge pool for data investigation and global network analysis functions for a better understanding of a cell’s interactome, (2 employment of public data for plausibility analysis and validation of in-house experimental data and (3 testing the applicability of Microsoft’s .NET architecture for bioinformatics applications. Data integrated into the ISP database can be queried through the Web portal PRIMOS (PRotein Interaction and MOlecule Search which is freely available at http://biomis.fh-hagenberg.at/isp/primos.

  19. Proteomic Analysis of Bovine Pregnancy-specific Serum Proteins by 2D Fluorescence Difference Gel Electrophoresis

    OpenAIRE

    Lee, Jae Eun; Lee, Jae Young; Kim, Hong Rye; Shin, Hyun Young; Lin, Tao; Jin, Dong Il

    2015-01-01

    Two dimensional-fluorescence difference gel electrophoresis (2D DIGE) is an emerging technique for comparative proteomics, which improves the reproducibility and reliability of differential protein expression analysis between samples. The purpose of this study was to investigate bovine pregnancy-specific proteins in the proteome between bovine pregnant and non-pregnant serum using DIGE technique. Serums of 2 pregnant Holstein dairy cattle at day 21 after artificial insemination and those of 2...

  20. Proteomic analysis of the EhV-86 virion

    Directory of Open Access Journals (Sweden)

    Lilley Kathryn S

    2008-03-01

    Full Text Available Abstract Background Emiliania huxleyi virus 86 (EhV-86 is the type species of the genus Coccolithovirus within the family Phycodnaviridae. The fully sequenced 407,339 bp genome is predicted to encode 473 protein coding sequences (CDSs and is the largest Phycodnaviridae sequenced to date. The majority of EhV-86 CDSs exhibit no similarity to proteins in the public databases. Results Proteomic analysis by 1-DE and then LC-MS/MS determined that the virion of EhV-86 is composed of at least 28 proteins, 23 of which are predicted to be membrane proteins. Besides the major capsid protein, putative function can be assigned to 4 other components of the virion: two lectin proteins, a thioredoxin and a serine/threonine protein kinase. Conclusion This study represents the first steps toward the identification of the protein components that make up the EhV-86 virion. Aside from the major capsid protein, whose function in the virion is well known and defined, the nature of the other proteins suggest roles involved with viral budding, caspase activation, signalling, anti-oxidation, virus adsorption and host range determination.

  1. A proteomic analysis of seed development in Brassica campestri L.

    Directory of Open Access Journals (Sweden)

    Wenlan Li

    Full Text Available To gain insights into the protein dynamics during seed development, a proteomic study on the developing Brassica campestri L. seeds with embryos in different embryogenesis stages was carried out. The seed proteins at 10, 16, 20, 25 and 35 DAP (days after pollination, respectively, were separated using two-dimensional gel electrophoresis and identities of 209 spots with altered abundance were determined by matrix-assisted laser desorption ionization time-of-flight/time-of-flight mass spectrometry (MALDI-TOF/TOF MS. These proteins were classified into 16 groups according to their functions. The most abundant proteins were related to primary metabolism, indicating the heavy demand of materials for rapid embryo growth. Besides, the high amount of proteins involved in protein processing and destination indicated importance of protein renewal during seed development. The remaining were those participated in oxidation/detoxification, energy, defense, transcription, protein synthesis, transporter, cell structure, signal transduction, secondary metabolism, transposition, DNA repair, storage and so on. Protein abundance profiles of each functional class were generated and hierarchical cluster analysis established 8 groups of dynamic patterns. Our results revealed novel characters of protein dynamics in seed development in Brassica campestri L. and provided valuable information about the complex process of seed development in plants.

  2. Proteomics Analysis for Finding Serum Markers of Ovarian Cancer

    Directory of Open Access Journals (Sweden)

    Yushan Cheng

    2014-01-01

    Full Text Available A combination of peptide ligand library beads (PLLB and 1D gel liquid chromatography-mass spectrometry/mass spectrometry (1DGel-LC-MS/MS was employed to analyze serum samples from patients with ovarian cancer and from healthy controls. Proteomic analysis identified 1200 serum proteins, among which 57 proteins were upregulated and 10 were downregulated in the sera from cancer patients. Retinol binding protein 4 (RBP4 is highly upregulated in the ovarian cancer serum samples. ELISA was employed to measure plasma concentrations of RBP4 in 80 samples from ovarian cancer patients, healthy individuals, myoma patients, and patients with benign ovarian tumor, respectively. The plasma concentrations of RBP4 ranging from 76.91 to 120.08 ng/mL with the mean value 89.13±1.67 ng/mL in ovarian cancer patients are significantly higher than those in healthy individuals (10.85±2.38 ng/mL. Results were further confirmed with immunohistochemistry, demonstrating that RBP4 expression levels in normal ovarian tissue were lower than those in ovarian cancer tissues. Our results suggested that RBP4 is a potential biomarker for diagnostic of screening ovarian cancer.

  3. Comprehensive proteomic analysis of the wheat pathogenic fungus Zymoseptoria tritici.

    Science.gov (United States)

    Yang, Fen; Yin, Qi

    2016-01-01

    Zymoseptoria tritici causes Septoria tritici blotch disease of wheat. To obtain a comprehensive protein dataset of this fungal pathogen, proteomes of Z. tritici growing in nutrient-limiting and rich media and in vivo at a late stage of wheat infection were fractionated by 1D gel or strong cation exchange (SCX) chromatography and analyzed by LC-MS/MS. A total of 5731, 5376 and 3168 Z. tritici proteins were confidently identified from these conditions, respectively. Of these in vitro and in planta proteins, 9 and 11% were predicted to contain signal peptides, respectively. Functional classification analysis revealed the proteins were involved in the various cellular activities. Comparison of three distinct protein expression profiles demonstrates the elevated carbohydrate, lipid and secondary metabolisms, transport, protein processing and energy production specifically in the host environment, in contrast to the enhancement of signaling, defense, replication, transcription and cell division in vitro. The data provide useful targets towards a better understanding of the molecular basis of Z. tritici growth, development, stress response and pathogenicity. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  4. Global Proteome Analysis Identifies Active Immunoproteasome Subunits in Human Platelets*

    Science.gov (United States)

    Klockenbusch, Cordula; Walsh, Geraldine M.; Brown, Lyda M.; Hoffman, Michael D.; Ignatchenko, Vladimir; Kislinger, Thomas; Kast, Juergen

    2014-01-01

    The discovery of new functions for platelets, particularly in inflammation and immunity, has expanded the role of these anucleate cell fragments beyond their primary hemostatic function. Here, four in-depth human platelet proteomic data sets were generated to explore potential new functions for platelets based on their protein content and this led to the identification of 2559 high confidence proteins. During a more detailed analysis, consistently high expression of the proteasome was discovered, and the composition and function of this complex, whose role in platelets has not been thoroughly investigated, was examined. Data set mining resulted in identification of nearly all members of the 26S proteasome in one or more data sets, except the β5 subunit. However, β5i, a component of the immunoproteasome, was identified. Biochemical analyses confirmed the presence of all catalytically active subunits of the standard 20S proteasome and immunoproteasome in human platelets, including β5, which was predominantly found in its precursor form. It was demonstrated that these components were assembled into the proteasome complex and that standard proteasome as well as immunoproteasome subunits were constitutively active in platelets. These findings suggest potential new roles for platelets in the immune system. For example, the immunoproteasome may be involved in major histocompatibility complex I (MHC I) peptide generation, as the MHC I machinery was also identified in our data sets. PMID:25146974

  5. Global proteome analysis identifies active immunoproteasome subunits in human platelets.

    Science.gov (United States)

    Klockenbusch, Cordula; Walsh, Geraldine M; Brown, Lyda M; Hoffman, Michael D; Ignatchenko, Vladimir; Kislinger, Thomas; Kast, Juergen

    2014-12-01

    The discovery of new functions for platelets, particularly in inflammation and immunity, has expanded the role of these anucleate cell fragments beyond their primary hemostatic function. Here, four in-depth human platelet proteomic data sets were generated to explore potential new functions for platelets based on their protein content and this led to the identification of 2559 high confidence proteins. During a more detailed analysis, consistently high expression of the proteasome was discovered, and the composition and function of this complex, whose role in platelets has not been thoroughly investigated, was examined. Data set mining resulted in identification of nearly all members of the 26S proteasome in one or more data sets, except the β5 subunit. However, β5i, a component of the immunoproteasome, was identified. Biochemical analyses confirmed the presence of all catalytically active subunits of the standard 20S proteasome and immunoproteasome in human platelets, including β5, which was predominantly found in its precursor form. It was demonstrated that these components were assembled into the proteasome complex and that standard proteasome as well as immunoproteasome subunits were constitutively active in platelets. These findings suggest potential new roles for platelets in the immune system. For example, the immunoproteasome may be involved in major histocompatibility complex I (MHC I) peptide generation, as the MHC I machinery was also identified in our data sets. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  6. Proteomics analysis of egg white proteins from different egg varieties.

    Science.gov (United States)

    Wang, Jiapei; Liang, Yue; Omana, Dileep A; Kav, Nat N V; Wu, Jianping

    2012-01-11

    The market of specialty eggs, such as omega-3-enriched eggs, organic eggs, and free-range eggs, is continuously growing. The nutritional composition of egg yolk can be manipulated by feed diet; however, it is not known if there is any difference in the composition of egg white proteins among different egg varieties. The purpose of the study was to compare the egg white proteins among six different egg varieties using proteomics analysis. Egg white proteins were analyzed using two-dimensional gel electrophoresis (2-DE), and 89 protein spots were subjected to LC-MS/MS. A total of 23 proteins, belonging to Gallus gallus , were identified from 72 detected protein spots. A quiescence-specific protein precursor in egg white was identified for the first time in this study. Significant differences in the abundant levels of 19 proteins (from 65 protein spots) were observed among six egg varieties. Four proteins, ovalbumin-related protein Y, cystatin, avidin, and albumin precursor, were not different among these six egg varieties. These findings suggest that the abundance, but not the composition, of egg white proteins varied among the egg varieties.

  7. Mastitomics, the integrated omics of bovine milk in an experimental model of Streptococcus uberis mastitis: 2. Label-free relative quantitative proteomics.

    Science.gov (United States)

    Mudaliar, Manikhandan; Tassi, Riccardo; Thomas, Funmilola C; McNeilly, Tom N; Weidt, Stefan K; McLaughlin, Mark; Wilson, David; Burchmore, Richard; Herzyk, Pawel; Eckersall, P David; Zadoks, Ruth N

    2016-08-16

    Mastitis, inflammation of the mammary gland, is the most common and costly disease of dairy cattle in the western world. It is primarily caused by bacteria, with Streptococcus uberis as one of the most prevalent causative agents. To characterize the proteome during Streptococcus uberis mastitis, an experimentally induced model of intramammary infection was used. Milk whey samples obtained from 6 cows at 6 time points were processed using label-free relative quantitative proteomics. This proteomic analysis complements clinical, bacteriological and immunological studies as well as peptidomic and metabolomic analysis of the same challenge model. A total of 2552 non-redundant bovine peptides were identified, and from these, 570 bovine proteins were quantified. Hierarchical cluster analysis and principal component analysis showed clear clustering of results by stage of infection, with similarities between pre-infection and resolution stages (0 and 312 h post challenge), early infection stages (36 and 42 h post challenge) and late infection stages (57 and 81 h post challenge). Ingenuity pathway analysis identified upregulation of acute phase protein pathways over the course of infection, with dominance of different acute phase proteins at different time points based on differential expression analysis. Antimicrobial peptides, notably cathelicidins and peptidoglycan recognition protein, were upregulated at all time points post challenge and peaked at 57 h, which coincided with 10 000-fold decrease in average bacterial counts. The integration of clinical, bacteriological, immunological and quantitative proteomics and other-omic data provides a more detailed systems level view of the host response to mastitis than has been achieved previously.

  8. Changes in cod muscle proteins during frozen storage revealed by proteome analysis and multivariate data analysis

    DEFF Research Database (Denmark)

    Kjærsgård, Inger Vibeke Holst; Nørrelykke, M.R.; Jessen, Flemming

    2006-01-01

    Multivariate data analysis has been combined with proteomics to enhance the recovery of information from 2-DE of cod muscle proteins during different storage conditions. Proteins were extracted according to 11 different storage conditions and samples were resolved by 2-DE. Data generated by 2-DE...... was subjected to principal component analysis (PCA) and discriminant partial least squares regression (DPLSR). Applying PCA to 2-DE data revealed the samples to form groups according to frozen storage time, whereas differences due to different storage temperatures or chilled storage in modified atmosphere...... light chain 1, 2 and 3, triose-phosphate isomerase, glyceraldehyde-3-phosphate dehydrogenase, aldolase A and two ?-actin fragments, and a nuclease diphosphate kinase B fragment to change in concentration, during frozen storage. Application of proteomics, multivariate data analysis and MS/MS to analyse...

  9. Proteome analysis of ofloxacin and moxifloxacin induced mycobacterium tuberculosis isolates by proteomic approach.

    Science.gov (United States)

    Lata, Manju; Sharma, Divakar; Kumar, Bhavnesh; Deo, Nirmala; Tiwari, Pramod Kumar; Bisht, Deepa; Venkatesan, Krishnamurthy

    2015-01-01

    Ofloxacin (OFX) and moxifloxacin (MOX) are the most promising second line drugs for tuberculosis treatment. Although the primary mechanism of action of OFX and MOX is gyrase inhibition, other possible mechanisms cannot be ruled out. Being the functional moiety of cell, the proteins act as primary targets for developing drugs, diagnostics and therapeutics. In this study we have investigated the proteomic changes of Mycobacterium tuberculosis isolates induced by OFX and MOX by applying comparative proteomic approaches based on two-dinensional gel electrophoresis (2DE) along with matrix assisted laser desorption ionisation time of flight mass spectrometry (MALDI TOF/TOF-MS) and bioinformatic tools. The findings are likely to provide new understanding of OFX and MOX mechanisms that might be helpful in exploring new diagnostics and drug targets. Our study explored eleven proteins (Rv2889c, Rv2623, Rv0952, Rv1827, Rv1932, Rv0054, Rv1080c, Rv3418c, Rv3914, Rv1636 and Rv0009) that were overexpressed in the presence of drugs. Among them, Rv2623, Rv1827 and Rv1636 were identified as proteins with unknown function. InterProScan and molecular docking revealed that the conserved domain of hypothetical proteins interact with OFX and MOX which indicate a probable inhibition/modulation of the functioning of these proteins by both drugs, which might be overexpressed to overcome this effect.

  10. Top-down proteomics for the analysis of proteolytic events - Methods, applications and perspectives.

    Science.gov (United States)

    Tholey, Andreas; Becker, Alexander

    2017-11-01

    Mass spectrometry based proteomics is an indispensable tool for almost all research areas relevant for the understanding of proteolytic processing, ranging from the identification of substrates, products and cleavage sites up to the analysis of structural features influencing protease activity. The majority of methods for these studies are based on bottom-up proteomics performing analysis at peptide level. As this approach is characterized by a number of pitfalls, e.g. loss of molecular information, there is an ongoing effort to establish top-down proteomics, performing separation and MS analysis both at intact protein level. We briefly introduce major approaches of bottom-up proteomics used in the field of protease research and highlight the shortcomings of these methods. We then discuss the present state-of-the-art of top-down proteomics. Together with the discussion of known challenges we show the potential of this approach and present a number of successful applications of top-down proteomics in protease research. This article is part of a Special Issue entitled: Proteolysis as a Regulatory Event in Pathophysiology edited by Stefan Rose-John. Copyright © 2017 Elsevier B.V. All rights reserved.

  11. Integration of deep transcriptome and proteome analyses reveals the components of alkaloid metabolism in opium poppy cell cultures

    Directory of Open Access Journals (Sweden)

    Schriemer David C

    2010-11-01

    Full Text Available Abstract Background Papaver somniferum (opium poppy is the source for several pharmaceutical benzylisoquinoline alkaloids including morphine, the codeine and sanguinarine. In response to treatment with a fungal elicitor, the biosynthesis and accumulation of sanguinarine is induced along with other plant defense responses in opium poppy cell cultures. The transcriptional induction of alkaloid metabolism in cultured cells provides an opportunity to identify components of this process via the integration of deep transcriptome and proteome databases generated using next-generation technologies. Results A cDNA library was prepared for opium poppy cell cultures treated with a fungal elicitor for 10 h. Using 454 GS-FLX Titanium pyrosequencing, 427,369 expressed sequence tags (ESTs with an average length of 462 bp were generated. Assembly of these sequences yielded 93,723 unigenes, of which 23,753 were assigned Gene Ontology annotations. Transcripts encoding all known sanguinarine biosynthetic enzymes were identified in the EST database, 5 of which were represented among the 50 most abundant transcripts. Liquid chromatography-tandem mass spectrometry (LC-MS/MS of total protein extracts from cell cultures treated with a fungal elicitor for 50 h facilitated the identification of 1,004 proteins. Proteins were fractionated by one-dimensional SDS-PAGE and digested with trypsin prior to LC-MS/MS analysis. Query of an opium poppy-specific EST database substantially enhanced peptide identification. Eight out of 10 known sanguinarine biosynthetic enzymes and many relevant primary metabolic enzymes were represented in the peptide database. Conclusions The integration of deep transcriptome and proteome analyses provides an effective platform to catalogue the components of secondary metabolism, and to identify genes encoding uncharacterized enzymes. The establishment of corresponding transcript and protein databases generated by next-generation technologies in a

  12. Integration of deep transcriptome and proteome analyses reveals the components of alkaloid metabolism in opium poppy cell cultures.

    Science.gov (United States)

    Desgagné-Penix, Isabel; Khan, Morgan F; Schriemer, David C; Cram, Dustin; Nowak, Jacek; Facchini, Peter J

    2010-11-18

    Papaver somniferum (opium poppy) is the source for several pharmaceutical benzylisoquinoline alkaloids including morphine, the codeine and sanguinarine. In response to treatment with a fungal elicitor, the biosynthesis and accumulation of sanguinarine is induced along with other plant defense responses in opium poppy cell cultures. The transcriptional induction of alkaloid metabolism in cultured cells provides an opportunity to identify components of this process via the integration of deep transcriptome and proteome databases generated using next-generation technologies. A cDNA library was prepared for opium poppy cell cultures treated with a fungal elicitor for 10 h. Using 454 GS-FLX Titanium pyrosequencing, 427,369 expressed sequence tags (ESTs) with an average length of 462 bp were generated. Assembly of these sequences yielded 93,723 unigenes, of which 23,753 were assigned Gene Ontology annotations. Transcripts encoding all known sanguinarine biosynthetic enzymes were identified in the EST database, 5 of which were represented among the 50 most abundant transcripts. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) of total protein extracts from cell cultures treated with a fungal elicitor for 50 h facilitated the identification of 1,004 proteins. Proteins were fractionated by one-dimensional SDS-PAGE and digested with trypsin prior to LC-MS/MS analysis. Query of an opium poppy-specific EST database substantially enhanced peptide identification. Eight out of 10 known sanguinarine biosynthetic enzymes and many relevant primary metabolic enzymes were represented in the peptide database. The integration of deep transcriptome and proteome analyses provides an effective platform to catalogue the components of secondary metabolism, and to identify genes encoding uncharacterized enzymes. The establishment of corresponding transcript and protein databases generated by next-generation technologies in a system with a well-defined metabolite profile facilitates

  13. Hepatic Proteomic Analysis Revealed Altered Metabolic Pathways in Insulin Resistant Akt1+/-/Akt2-/-Mice

    Science.gov (United States)

    Pedersen, Brian A; Wang, Weiwen; Taylor, Jared F; Khattab, Omar S; Chen, Yu-Han; Edwards, Robert A; Yazdi, Puya G; Wang, Ping H

    2015-01-01

    Objective The aim of this study was to identify liver proteome changes in a mouse model of severe insulin resistance and markedly decreased leptin levels. Methods Two-dimensional differential gel electrophoresis was utilized to identify liver proteome changes in AKT1+/-/AKT2-/- mice. Proteins with altered levels were identified with tandem mass spectrometry. Ingenuity Pathway analysis was performed for the interpretation of the biological significance of the observed proteomic changes. Results 11 proteins were identified from 2 biological replicates to be differentially expressed by a ratio of at least 1.3 between age-matched insulin resistant (Akt1+/-/Akt2-/-) and wild type mice. Albumin and mitochondrial ornithine aminotransferase were detected from multiple spots, which suggest post-translational modifications. Enzymes of the urea cycle were common members of top regulated pathways. Conclusion Our results help to unveil the regulation of the liver proteome underlying altered metabolism in an animal model of severe insulin resistance. PMID:26455965

  14. Integrated proteomic and N-glycoproteomic analyses of doxorubicin sensitive and resistant ovarian cancer cells reveal glycoprotein alteration in protein abundance and glycosylation

    Science.gov (United States)

    Hou, Junjie; Zhang, Chengqian; Xue, Peng; Wang, Jifeng; Chen, Xiulan; Guo, Xiaojing; Yang, Fuquan

    2017-01-01

    Ovarian cancer is one of the most common cancer among women in the world, and chemotherapy remains the principal treatment for patients. However, drug resistance is a major obstacle to the effective treatment of ovarian cancers and the underlying mechanism is not clear. An increased understanding of the mechanisms that underline the pathogenesis of drug resistance is therefore needed to develop novel therapeutics and diagnostic. Herein, we report the comparative analysis of the doxorubicin sensitive OVCAR8 cells and its doxorubicin-resistant variant NCI/ADR-RES cells using integrated global proteomics and N-glycoproteomics. A total of 1525 unique N-glycosite-containing peptides from 740 N-glycoproteins were identified and quantified, of which 253 N-glycosite-containing peptides showed significant change in the NCI/ADR-RES cells. Meanwhile, stable isotope labeling by amino acids in cell culture (SILAC) based comparative proteomic analysis of the two ovarian cancer cells led to the quantification of 5509 proteins. As about 50% of the identified N-glycoproteins are low-abundance membrane proteins, only 44% of quantified unique N-glycosite-containing peptides had corresponding protein expression ratios. The comparison and calibration of the N-glycoproteome versus the proteome classified 14 change patterns of N-glycosite-containing peptides, including 8 up-regulated N-glycosite-containing peptides with the increased glycosylation sites occupancy, 35 up-regulated N-glycosite-containing peptides with the unchanged glycosylation sites occupancy, 2 down-regulated N-glycosite-containing peptides with the decreased glycosylation sites occupancy, 46 down-regulated N-glycosite-containing peptides with the unchanged glycosylation sites occupancy. Integrated proteomic and N-glycoproteomic analyses provide new insights, which can help to unravel the relationship of N-glycosylation and multidrug resistance (MDR), understand the mechanism of MDR, and discover the new diagnostic and

  15. Integrated proteomic and transcriptomic investigation of the acetaminophen toxicity in liver microfluidic biochip.

    Directory of Open Access Journals (Sweden)

    Jean Matthieu Prot

    Full Text Available Microfluidic bioartificial organs allow the reproduction of in vivo-like properties such as cell culture in a 3D dynamical micro environment. In this work, we established a method and a protocol for performing a toxicogenomic analysis of HepG2/C3A cultivated in a microfluidic biochip. Transcriptomic and proteomic analyses have shown the induction of the NRF2 pathway and the related drug metabolism pathways when the HepG2/C3A cells were cultivated in the biochip. The induction of those pathways in the biochip enhanced the metabolism of the N-acetyl-p-aminophenol drug (acetaminophen-APAP when compared to Petri cultures. Thus, we observed 50% growth inhibition of cell proliferation at 1 mM in the biochip, which appeared similar to human plasmatic toxic concentrations reported at 2 mM. The metabolic signature of APAP toxicity in the biochip showed similar biomarkers as those reported in vivo, such as the calcium homeostasis, lipid metabolism and reorganization of the cytoskeleton, at the transcriptome and proteome levels (which was not the case in Petri dishes. These results demonstrate a specific molecular signature for acetaminophen at transcriptomic and proteomic levels closed to situations found in vivo. Interestingly, a common component of the signature of the APAP molecule was identified in Petri and biochip cultures via the perturbations of the DNA replication and cell cycle. These findings provide an important insight into the use of microfluidic biochips as new tools in biomarker research in pharmaceutical drug studies and predictive toxicity investigations.

  16. Redox proteomic analysis of the gastrocnemius muscle from adult and old mice

    Directory of Open Access Journals (Sweden)

    Brian McDonagh

    2015-09-01

    Full Text Available The data provides information in support of the research article, “Differential Cysteine Labeling and Global Label-Free Proteomics Reveals an Altered Metabolic State in Skeletal Muscle Aging”, Journal of Proteome Research, 2014, 13 (11, 2008–21 [1]. Raw data is available from ProteomeXchange [2] with identifier PDX001054. The proteome of gastrocnemius muscle from adult and old mice was analyzed by global label-free proteomics and the relative quantification of specific reduced and reversibly oxidized Cysteine (Cys residues was performed using Skyline [3]. Briefly, reduced Cysteine (Cys containing peptides was alkylated using N-ethylmalemide (d0-NEM. Samples were desalted and reversibly oxidized Cys residues were reduced using tris(2-carboxyethylphosphine (TCEP and the newly formed reduced Cys residues were labeled with heavy NEM( d5-NEM. Label-free analysis of the global proteome of adult (n=5 and old (n=4 gastrocnemius muscles was performed using Peaks7™ mass spectrometry data analysis software [4]. Relative quantification of Cys containing peptides that were identified as reduced (d(0 NEM labeled and reversibly oxidized d(5–NEM labeled was performed using the intensity of their precursor ions in Skyline. Results indicate that muscles from old mice show reduced redox flexibility particularly in proteins involved in the generation of precursor metabolites and energy metabolism, indicating a loss in the flexibility of the redox energy response.

  17. Sample preparation and fractionation for proteome analysis and cancer biomarker discovery by mass spectrometry.

    Science.gov (United States)

    Ahmed, Farid E

    2009-03-01

    Sample preparation and fractionation technologies are one of the most crucial processes in proteomic analysis and biomarker discovery in solubilized samples. Chromatographic or electrophoretic proteomic technologies are also available for separation of cellular protein components. There are, however, considerable limitations in currently available proteomic technologies as none of them allows for the analysis of the entire proteome in a simple step because of the large number of peptides, and because of the wide concentration dynamic range of the proteome in clinical blood samples. The results of any undertaken experiment depend on the condition of the starting material. Therefore, proper experimental design and pertinent sample preparation is essential to obtain meaningful results, particularly in comparative clinical proteomics in which one is looking for minor differences between experimental (diseased) and control (nondiseased) samples. This review discusses problems associated with general and specialized strategies of sample preparation and fractionation, dealing with samples that are solution or suspension, in a frozen tissue state, or formalin-preserved tissue archival samples, and illustrates how sample processing might influence detection with mass spectrometric techniques. Strategies that dramatically improve the potential for cancer biomarker discovery in minimally invasive, blood-collected human samples are also presented.

  18. Proteomics analysis of alfalfa response to heat stress.

    Directory of Open Access Journals (Sweden)

    Weimin Li

    Full Text Available The proteome responses to heat stress have not been well understood. In this study, alfalfa (Medicago sativa L. cv. Huaiyin seedlings were exposed to 25 °C (control and 40 °C (heat stress in growth chambers, and leaves were collected at 24, 48 and 72 h after treatment, respectively. The morphological, physiological and proteomic processes were negatively affected under heat stress. Proteins were extracted and separated by two-dimensional polyacrylamide gel electrophoresis (2-DE, and differentially expressed protein spots were identified by mass spectrometry (MS. Totally, 81 differentially expressed proteins were identified successfully by MALDI-TOF/TOF. These proteins were categorized into nine classes: including metabolism, energy, protein synthesis, protein destination/storage, transporters, intracellular traffic, cell structure, signal transduction and disease/defence. Five proteins were further analyzed for mRNA levels. The results of the proteomics analyses provide a better understanding of the molecular basis of heat-stress responses in alfalfa.

  19. Integration of metabolomics and proteomics in molecular plant physiology--coping with the complexity by data-dimensionality reduction.

    Science.gov (United States)

    Weckwerth, Wolfram

    2008-02-01

    In recent years, genomics has been extended to functional genomics. Toward the characterization of organisms or species on the genome level, changes on the metabolite and protein level have been shown to be essential to assign functions to genes and to describe the dynamic molecular phenotype. Gas chromatography (GC) and liquid chromatography coupled to mass spectrometry (GC- and LC-MS) are well suited for the fast and comprehensive analysis of ultracomplex metabolite samples. For the integration of metabolite profiles with quantitative protein profiles, a high throughput (HTP) shotgun proteomics approach using LC-MS and label-free quantification of unique proteins in a complex protein digest is described. Multivariate statistics are applied to examine sample pattern recognition based on data-dimensionality reduction and biomarker identification in plant systems biology. The integration of the data reveal multiple correlative biomarkers providing evidence for an increase of information in such holistic approaches. With computational simulation of metabolic networks and experimental measurements, it can be shown that biochemical regulation is reflected by metabolite network dynamics measured in a metabolomics approach. Examples in molecular plant physiology are presented to substantiate the integrative approach.

  20. Proteomic analysis of africanized bee venom: a comparison of protein extraction methods

    Directory of Open Access Journals (Sweden)

    Yessica Pineda Guerra

    2016-09-01

    Full Text Available The Africanised bee is the most common type of bee in Colombia, and therapeutic properties for different diseases have been attributed to its venom, without much scientific support. A literature search of reports on the proteomic analysis of honeybee venom yielded four different methods for extracting proteins from bee venom. The first method consists in resuspending the venom in 7 M Urea, followed by precipitation with acetone and finally resuspending the pellet in 7 M Urea and 4 % CHAPS. For the second method, the venom is resuspended in lysis buffer, precipitated with trichloroacetic acid, and then resuspended in 7 M Urea and 4 % CHAPS. The third method is similar to the previous one, except that the precipitation step is performed with acetone instead of trichloroacetic acid. Finally, the fourth method is to resuspend the venom in distilled water, precipitate with acetone and resuspend in 7 M Urea and 4 % CHAPS. This work focused on comparing the performance of these four extraction methods, in order to determine the method with the best results in terms of concentration and integrity of the proteins obtained. Of the four methods evaluated, the best results in terms of protein concentration and yield were obtained by resuspending the bee venom in lysis buffer followed by precipitation with acetone (method 3, and by resuspending in distilled water followed by precipitation with acetone (method 4. Of these, the method that maintained protein integrity and yielded the best proteomic profile was that in which the bee venom was resuspended in lysis buffer followed by precipitation with acetone (method 3.

  1. Understanding pea resistance mechanisms in response to Fusarium oxysporum through proteomic analysis.

    Science.gov (United States)

    Castillejo, María Ángeles; Bani, Moustafa; Rubiales, Diego

    2015-07-01

    Fusarium oxysporum f. sp. pisi (Fop) is an important and destructive pathogen affecting pea crop (Pisum sativum) throughout the world. Control of this disease is achieved mainly by integration of different disease management procedures. However, the constant evolution of the pathogen drives the necessity to broaden the molecular basis of resistance to Fop. Our proteomic study was performed on pea with the aim of identifying proteins involved in different resistance mechanisms operating during F. oxysporum infection. For such purpose, we used a two-dimensional electrophoresis (2-DE) coupled to mass spectrometry (MALDI-TOF/TOF) analysis to study the root proteome of three pea genotypes showing different resistance response to Fop race 2. Multivariate statistical analysis identified 132 differential protein spots under the experimental conditions (genotypes/treatments). All of these protein spots were subjected to mass spectrometry analysis to deduce their possible functions. A total of 53 proteins were identified using a combination of peptide mass fingerprinting (PMF) and MSMS fragmentation. The following main functional categories were assigned to the identified proteins: carbohydrate and energy metabolism, nucleotides and aminoacid metabolism, signal transduction and cellular process, folding and degradation, redox and homeostasis, defense, biosynthetic process and transcription/translation. Results obtained in this work suggest that the most susceptible genotypes have increased levels of enzymes involved in the production of reducing power which could then be used as cofactor for enzymes of the redox reactions. This is in concordance with the fact that a ROS burst occurred in the same genotypes, as well as an increase of PR proteins. Conversely, in the resistant genotype proteins responsible to induce changes in the membrane and cell wall composition related to reinforcement were identified. Results are discussed in terms of the differential response to Fop

  2. Predicting co-complexed protein pairs using genomic and proteomic data integration

    Directory of Open Access Journals (Sweden)

    King Oliver D

    2004-04-01

    Full Text Available Abstract Background Identifying all protein-protein interactions in an organism is a major objective of proteomics. A related goal is to know which protein pairs are present in the same protein complex. High-throughput methods such as yeast two-hybrid (Y2H and affinity purification coupled with mass spectrometry (APMS have been used to detect interacting proteins on a genomic scale. However, both Y2H and APMS methods have substantial false-positive rates. Aside from high-throughput interaction screens, other gene- or protein-pair characteristics may also be informative of physical interaction. Therefore it is desirable to integrate multiple datasets and utilize their different predictive value for more accurate prediction of co-complexed relationship. Results Using a supervised machine learning approach – probabilistic decision tree, we integrated high-throughput protein interaction datasets and other gene- and protein-pair characteristics to predict co-complexed pairs (CCP of proteins. Our predictions proved more sensitive and specific than predictions based on Y2H or APMS methods alone or in combination. Among the top predictions not annotated as CCPs in our reference set (obtained from the MIPS complex catalogue, a significant fraction was found to physically interact according to a separate database (YPD, Yeast Proteome Database, and the remaining predictions may potentially represent unknown CCPs. Conclusions We demonstrated that the probabilistic decision tree approach can be successfully used to predict co-complexed protein (CCP pairs from other characteristics. Our top-scoring CCP predictions provide testable hypotheses for experimental validation.

  3. Transcriptome and quantitative proteome analysis reveals molecular processes associated with larval metamorphosis in the polychaete pseudopolydora vexillosa

    KAUST Repository

    Chandramouli, Kondethimmanahalli; Sun, Jin; Mok, FloraSy; Liu, Lingli; Qiu, Jianwen; Ravasi, Timothy; Qian, Peiyuan

    2013-01-01

    Larval growth of the polychaete worm Pseudopolydora vexillosa involves the formation of segment-specific structures. When larvae attain competency to settle, they discard swimming chaetae and secrete mucus. The larvae build tubes around themselves and metamorphose into benthic juveniles. Understanding the molecular processes, which regulate this complex and unique transition, remains a major challenge because of the limited molecular information available. To improve this situation, we conducted high-throughput RNA sequencing and quantitative proteome analysis of the larval stages of P. vexillosa. Based on gene ontology (GO) analysis, transcripts related to cellular and metabolic processes, binding, and catalytic activities were highly represented during larval-adult transition. Mitogen-activated protein kinase (MAPK), calcium-signaling, Wnt/β-catenin, and notch signaling metabolic pathways were enriched in transcriptome data. Quantitative proteomics identified 107 differentially expressed proteins in three distinct larval stages. Fourteen and 53 proteins exhibited specific differential expression during competency and metamorphosis, respectively. Dramatic up-regulation of proteins involved in signaling, metabolism, and cytoskeleton functions were found during the larval-juvenile transition. Several proteins involved in cell signaling, cytoskeleton and metabolism were up-regulated, whereas proteins related to transcription and oxidative phosphorylation were down-regulated during competency. The integration of high-throughput RNA sequencing and quantitative proteomics allowed a global scale analysis of larval transcripts/proteins associated molecular processes in the metamorphosis of polychaete worms. Further, transcriptomic and proteomic insights provide a new direction to understand the fundamental mechanisms that regulate larval metamorphosis in polychaetes. © 2013 American Chemical Society.

  4. Transcriptome and quantitative proteome analysis reveals molecular processes associated with larval metamorphosis in the polychaete pseudopolydora vexillosa

    KAUST Repository

    Chandramouli, Kondethimmanahalli

    2013-03-01

    Larval growth of the polychaete worm Pseudopolydora vexillosa involves the formation of segment-specific structures. When larvae attain competency to settle, they discard swimming chaetae and secrete mucus. The larvae build tubes around themselves and metamorphose into benthic juveniles. Understanding the molecular processes, which regulate this complex and unique transition, remains a major challenge because of the limited molecular information available. To improve this situation, we conducted high-throughput RNA sequencing and quantitative proteome analysis of the larval stages of P. vexillosa. Based on gene ontology (GO) analysis, transcripts related to cellular and metabolic processes, binding, and catalytic activities were highly represented during larval-adult transition. Mitogen-activated protein kinase (MAPK), calcium-signaling, Wnt/β-catenin, and notch signaling metabolic pathways were enriched in transcriptome data. Quantitative proteomics identified 107 differentially expressed proteins in three distinct larval stages. Fourteen and 53 proteins exhibited specific differential expression during competency and metamorphosis, respectively. Dramatic up-regulation of proteins involved in signaling, metabolism, and cytoskeleton functions were found during the larval-juvenile transition. Several proteins involved in cell signaling, cytoskeleton and metabolism were up-regulated, whereas proteins related to transcription and oxidative phosphorylation were down-regulated during competency. The integration of high-throughput RNA sequencing and quantitative proteomics allowed a global scale analysis of larval transcripts/proteins associated molecular processes in the metamorphosis of polychaete worms. Further, transcriptomic and proteomic insights provide a new direction to understand the fundamental mechanisms that regulate larval metamorphosis in polychaetes. © 2013 American Chemical Society.

  5. Comparative proteomic analysis of brains of naturally aging mice.

    Science.gov (United States)

    Yang, S; Liu, T; Li, S; Zhang, X; Ding, Q; Que, H; Yan, X; Wei, K; Liu, S

    2008-06-26

    We used comparative proteomic techniques to identify aging-related brain proteins in normal mice from neonate to old age. By 2-dimensional electrophoresis (2-DE), matrix assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry (MS) and peptide mass fingerprint (PMF) analysis, 39 proteins were identified, among which 6 stayed unchanged since 3 months, 6 increased and 27 decreased in various manners during aging. They are mainly involved in processes usually with destructive changes during aging, such as metabolism, transport, signaling, stress response and apoptosis. The 27 proteins' decrease may be responsible for brain aging. In particular, decrease of proteasome alpha subunits 3/6, ubiquitin carboxyl-terminal esterase L3, valosin-containing protein and calreticulin may be responsible for the declination of protein quality control; glutamate dehydrogenase 1, isocitrate dehydrogenase 1 and ubiquinol cytochrome c reductase core protein 2 for the shortage of energy and reducing agent; ubiquitin-conjugating enzyme E2N and heterogeneous nuclear ribonucleoprotein A2/B1 for the increase of DNA damage and transcription detuning; calbindin 1 and amphiphysin for the disturbance of synaptic transport and ion signals. The six proteins' increase may be involved in anti-aging processes. In particular, transketolase, mitochondrial creatine kinase 1 and ribosomal protein L37 may help to enhance energy metabolism; triosephosphate isomerase 1 may help to resist oxidative stress. Moreover, most of these proteins were found for the first time to be involved in the natural senescence of brain, which would provide new clues about the mechanism of brain aging.

  6. Proteomic analysis of zebrafish embryos exposed to simulated-microgravity

    Science.gov (United States)

    Hang, Xiaoming; Ma, Wenwen; Wang, Wei; Liu, Cong; Sun, Yeqing

    Microgravity can induce a serial of physiological and pathological changes in human body, such as cardiovascular functional disorder, bone loss, muscular atrophy and impaired immune system function, etc. In this research, we focus on the influence of microgravity to vertebrate embryo development. As a powerful model for studying vertebrate development, zebrafish embryos at 8 hpf (hour past fertilization) and 24 hpf were placed into a NASA developed bioreac-tor (RCCS) to simulate microgravity for 64 and 48 hours, respectively. The same number of control embryos from the same parents were placed in a tissue culture dish at the same temper-ature of 28° C. Each experiment was repeated 3 times and analyzed by two-dimensional (2-D) gel electrophoresis. Image analysis of silver stained 2-D gels revealed that 64 from total 292 protein spots showed quantitative and qualitative variations that were significantly (P<0.05) and reproducibly different between simulate-microgravity treatment and the stationary control samples. 4 protein spots with significant expression alteration (P<0.01) were excised from 2-D gels and analyzed by MALDI-TOF/TOF mass spectra primarily. Of these proteins, 3 down-regulated proteins were identified as bectin 2, centrosomal protein of 135kDa and tropomyosin 4, while the up-regulated protein was identified as creatine kinase muscle B. Other protein spots showed significant expression alteration will be identified successively and the corresponding genes expression will also be measured by Q-PCR method at different development stages. The data presented in this study illustrate that zebrafish embryo can be significantly induced by microgravity on the expression of proteins involved in bone and muscle formation. Key Words: Danio rerio; Simulated-microgravity; Proteomics

  7. GeLC-MS: A Sample Preparation Method for Proteomics Analysis of Minimal Amount of Tissue.

    Science.gov (United States)

    Makridakis, Manousos; Vlahou, Antonia

    2017-10-10

    Application of various proteomics methodologies have been implemented for the global and targeted proteome analysis of many different types of biological samples such as tissue, urine, plasma, serum, blood, and cell lines. Among the aforementioned biological samples, tissue has an exceptional role into clinical research and practice. Disease initiation and progression is usually located at the tissue level of different organs, making the analysis of this material very important for the understanding of the disease pathophysiology. Despite the significant advances in the mass spectrometry instrumentation, tissue proteomics still faces several challenges mainly due to increased sample complexity and heterogeneity. However, the most prominent challenge is attributed to the invasive procedure of tissue sampling which restricts the availability of fresh frozen tissue to minimal amounts and limited number of samples. Application of GeLC-MS sample preparation protocol for tissue proteomics analysis can greatly facilitate making up for these difficulties. In this chapter, a step by step guide for the proteomics analysis of minute amounts of tissue samples using the GeLC-MS sample preparation protocol, as applied by our group in the analysis of multiple different types of tissues (vessels, kidney, bladder, prostate, heart) is provided.

  8. Proteomic Analysis of Male-Fertility Restoration in CMS Onion

    Science.gov (United States)

    The production of hybrid-onion seed is dependent on cytoplasmic-genic male sterility (CMS) systems. For the most commonly used CMS, male-sterile (S) cytoplasm interacts with a dominant allele at one nuclear male-fertility restoration locus (Ms) to condition male fertility. We are using proteomics ...

  9. Comprehensive proteomic analysis of the wheat pathogenic fungus Zymoseptoria tritici

    DEFF Research Database (Denmark)

    Yang, Fen; Yin, Qi

    2016-01-01

    Zymoseptoria tritici causes Septoria tritici blotch disease of wheat. To obtain a comprehensive protein dataset of this fungal pathogen, proteomes of Z. tritici growing in nutrient-limiting and rich media and in vivo at a late stage of wheat infection were fractionated by 1D gel or strong cation...

  10. Proteome analysis of Bordetella pertussis isolated from human macrophages

    Czech Academy of Sciences Publication Activity Database

    Lamberti, Y.; Cafiero, J.H.; Surmann, K.; Valdez, H.; Holubová, Jana; Večerek, Branislav; Šebo, Peter; Schmidt, F.; Völker, U.; Rodriguez, M.E.

    2016-01-01

    Roč. 136, MAY16 (2016), s. 55-67 ISSN 1874-3919 R&D Projects: GA MŠk(CZ) 7AMB14AR028 Institutional support: RVO:61388971 Keywords : Bordetella pertussis * Intracellular survival * Proteomics Subject RIV: EE - Microbiology, Virology Impact factor: 3.914, year: 2016

  11. Integrated multi-level quality control for proteomic profiling studies using mass spectrometry

    Directory of Open Access Journals (Sweden)

    Barrett Jennifer H

    2008-12-01

    Full Text Available Abstract Background Proteomic profiling using mass spectrometry (MS is one of the most promising methods for the analysis of complex biological samples such as urine, serum and tissue for biomarker discovery. Such experiments are often conducted using MALDI-TOF (matrix-assisted laser desorption/ionisation time-of-flight and SELDI-TOF (surface-enhanced laser desorption/ionisation time-of-flight MS. Using such profiling methods it is possible to identify changes in protein expression that differentiate disease states and individual proteins or patterns that may be useful as potential biomarkers. However, the incorporation of quality control (QC processes that allow the identification of low quality spectra reliably and hence allow the removal of such data before further analysis is often overlooked. In this paper we describe rigorous methods for the assessment of quality of spectral data. These procedures are presented in a user-friendly, web-based program. The data obtained post-QC is then examined using variance components analysis to quantify the amount of variance due to some of the factors in the experimental design. Results Using data from a SELDI profiling study of serum from patients with different levels of renal function, we show how the algorithms described in this paper may be used to detect systematic variability within and between sample replicates, pooled samples and SELDI chips and spots. Manual inspection of those spectral data that were identified as being of poor quality confirmed the efficacy of the algorithms. Variance components analysis demonstrated the relatively small amount of technical variance attributable to day of profile generation and experimental array. Conclusion Using the techniques described in this paper it is possible to reliably detect poor quality data within proteomic profiling experiments undertaken by MS. The removal of these spectra at the initial stages of the analysis substantially improves the

  12. Proteomic analysis of pig (Sus scrofa olfactory soluble proteome reveals O-GlcNAcylation of secreted odorant-binding proteins

    Directory of Open Access Journals (Sweden)

    Patricia eNAGNAN-LE MEILLOUR

    2014-12-01

    Full Text Available The diversity of olfactory binding proteins (OBPs is a key point to understand their role in molecular olfaction. Since only few different sequences were characterized in each mammalian species, they have been considered as passive carriers of odors and pheromones. We have explored the soluble proteome of pig nasal mucus, taking benefit of the powerful tools of proteomics. Combining two-dimensional electrophoresis, mass spectrometry and western-blot with specific antibodies, our analyses revealed for the first time that the pig nasal mucus is mainly composed of secreted OBP isoforms, some of them being potentially modified by O-GlcNAcylation. An ortholog gene of the glycosyltransferase responsible of the O-GlcNAc linking on extracellular proteins in Drosophila and Mouse (EOGT was amplified from tissues of pigs of different ages and sex. The sequence was used in a phylogenetic analysis, which evidenced conservation of EOGT in insect and mammalian models studied in molecular olfaction. Extracellular O-GlcNAcylation of secreted OBPs could finely modulate their binding specificities to odors and pheromones. This constitutes a new mechanism for extracellular signaling by OBPs, suggesting that they act as the first step of odor discrimination.

  13. Streamlined Membrane Proteome Preparation for Shotgun Proteomics Analysis with Triton X-100 Cloud Point Extraction and Nanodiamond Solid Phase Extraction

    Directory of Open Access Journals (Sweden)

    Minh D. Pham

    2016-05-01

    Full Text Available While mass spectrometry (MS plays a key role in proteomics research, characterization of membrane proteins (MP by MS has been a challenging task because of the presence of a host of interfering chemicals in the hydrophobic protein extraction process, and the low protease digestion efficiency. We report a sample preparation protocol, two-phase separation with Triton X-100, induced by NaCl, with coomassie blue added for visualizing the detergent-rich phase, which streamlines MP preparation for SDS-PAGE analysis of intact MP and shot-gun proteomic analyses. MP solubilized in the detergent-rich milieu were then sequentially extracted and fractionated by surface-oxidized nanodiamond (ND at three pHs. The high MP affinity of ND enabled extensive washes for removal of salts, detergents, lipids, and other impurities to ensure uncompromised ensuing purposes, notably enhanced proteolytic digestion and down-stream mass spectrometric (MS analyses. Starting with a typical membranous cellular lysate fraction harvested with centrifugation/ultracentrifugation, MP purities of 70%, based on number (not weight of proteins identified by MS, was achieved; the weight-based purity can be expected to be much higher.

  14. 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...

  15. Elevated host lipid metabolism revealed by iTRAQ-based quantitative proteomic analysis of cerebrospinal fluid of tuberculous meningitis patients

    Energy Technology Data Exchange (ETDEWEB)

    Mu, Jun [Department of Neurology, The First Affiliated Hospital of Chongqing Medical University, Chongqing (China); Institute of Neuroscience and the Collaborative Innovation Center for Brain Science, Chongqing Medical University, Chongqing (China); Chongqing Key Laboratory of Neurobiology, Chongqing (China); Yang, Yongtao [Institute of Neuroscience and the Collaborative Innovation Center for Brain Science, Chongqing Medical University, Chongqing (China); Chongqing Key Laboratory of Neurobiology, Chongqing (China); Department of Neurology, Yongchuan Hospital of Chongqing Medical University, Chongqing (China); Chen, Jin; Cheng, Ke; Li, Qi; Wei, Yongdong; Zhu, Dan; Shao, Weihua; Zheng, Peng [Department of Neurology, The First Affiliated Hospital of Chongqing Medical University, Chongqing (China); Institute of Neuroscience and the Collaborative Innovation Center for Brain Science, Chongqing Medical University, Chongqing (China); Chongqing Key Laboratory of Neurobiology, Chongqing (China); Xie, Peng, E-mail: xiepeng@cqmu.edu.cn [Department of Neurology, The First Affiliated Hospital of Chongqing Medical University, Chongqing (China); Institute of Neuroscience and the Collaborative Innovation Center for Brain Science, Chongqing Medical University, Chongqing (China); Chongqing Key Laboratory of Neurobiology, Chongqing (China); Department of Neurology, Yongchuan Hospital of Chongqing Medical University, Chongqing (China)

    2015-10-30

    Purpose: Tuberculous meningitis (TBM) remains to be one of the most deadly infectious diseases. The pathogen interacts with the host immune system, the process of which is largely unknown. Various cellular processes of Mycobacterium tuberculosis (MTB) centers around lipid metabolism. To determine the lipid metabolism related proteins, a quantitative proteomic study was performed here to identify differential proteins in the cerebrospinal fluid (CSF) obtained from TBM patients (n = 12) and healthy controls (n = 12). Methods: CSF samples were desalted, concentrated, labelled with isobaric tags for relative and absolute quantitation (iTRAQ™), and analyzed by multi-dimensional liquid chromatography-tandem mass spectrometry (LC-MS/MS). Gene ontology and proteomic phenotyping analysis of the differential proteins were conducted using Database for Annotation, Visualization, and Integrated Discovery (DAVID) Bioinformatics Resources. ApoE and ApoB were selected for validation by ELISA. Results: Proteomic phenotyping of the 4 differential proteins was invloved in the lipid metabolism. ELISA showed significantly increased ApoB levels in TBM subjects compared to healthy controls. Area under the receiver operating characteristic curve analysis demonstrated ApoB levels could distinguish TBM subjects from healthy controls and viral meningitis subjects with 89.3% sensitivity and 92% specificity. Conclusions: CSF lipid metabolism disregulation, especially elevated expression of ApoB, gives insights into the pathogenesis of TBM. Further evaluation of these findings in larger studies including anti-tuberculosis medicated and unmedicated patient cohorts with other center nervous system infectious diseases is required for successful clinical translation. - Highlights: • The first proteomic study on the cerebrospinal fluid of tuberculous meningitis patients using iTRAQ. • Identify 4 differential proteins invloved in the lipid metabolism. • Elevated expression of ApoB gives

  16. Elevated host lipid metabolism revealed by iTRAQ-based quantitative proteomic analysis of cerebrospinal fluid of tuberculous meningitis patients

    International Nuclear Information System (INIS)

    Mu, Jun; Yang, Yongtao; Chen, Jin; Cheng, Ke; Li, Qi; Wei, Yongdong; Zhu, Dan; Shao, Weihua; Zheng, Peng; Xie, Peng

    2015-01-01

    Purpose: Tuberculous meningitis (TBM) remains to be one of the most deadly infectious diseases. The pathogen interacts with the host immune system, the process of which is largely unknown. Various cellular processes of Mycobacterium tuberculosis (MTB) centers around lipid metabolism. To determine the lipid metabolism related proteins, a quantitative proteomic study was performed here to identify differential proteins in the cerebrospinal fluid (CSF) obtained from TBM patients (n = 12) and healthy controls (n = 12). Methods: CSF samples were desalted, concentrated, labelled with isobaric tags for relative and absolute quantitation (iTRAQ™), and analyzed by multi-dimensional liquid chromatography-tandem mass spectrometry (LC-MS/MS). Gene ontology and proteomic phenotyping analysis of the differential proteins were conducted using Database for Annotation, Visualization, and Integrated Discovery (DAVID) Bioinformatics Resources. ApoE and ApoB were selected for validation by ELISA. Results: Proteomic phenotyping of the 4 differential proteins was invloved in the lipid metabolism. ELISA showed significantly increased ApoB levels in TBM subjects compared to healthy controls. Area under the receiver operating characteristic curve analysis demonstrated ApoB levels could distinguish TBM subjects from healthy controls and viral meningitis subjects with 89.3% sensitivity and 92% specificity. Conclusions: CSF lipid metabolism disregulation, especially elevated expression of ApoB, gives insights into the pathogenesis of TBM. Further evaluation of these findings in larger studies including anti-tuberculosis medicated and unmedicated patient cohorts with other center nervous system infectious diseases is required for successful clinical translation. - Highlights: • The first proteomic study on the cerebrospinal fluid of tuberculous meningitis patients using iTRAQ. • Identify 4 differential proteins invloved in the lipid metabolism. • Elevated expression of ApoB gives

  17. Integrated genetic analysis microsystems

    International Nuclear Information System (INIS)

    Lagally, Eric T; Mathies, Richard A

    2004-01-01

    With the completion of the Human Genome Project and the ongoing DNA sequencing of the genomes of other animals, bacteria, plants and others, a wealth of new information about the genetic composition of organisms has become available. However, as the demand for sequence information grows, so does the workload required both to generate this sequence and to use it for targeted genetic analysis. Microfabricated genetic analysis systems are well poised to assist in the collection and use of these data through increased analysis speed, lower analysis cost and higher parallelism leading to increased assay throughput. In addition, such integrated microsystems may point the way to targeted genetic experiments on single cells and in other areas that are otherwise very difficult. Concomitant with these advantages, such systems, when fully integrated, should be capable of forming portable systems for high-speed in situ analyses, enabling a new standard in disciplines such as clinical chemistry, forensics, biowarfare detection and epidemiology. This review will discuss the various technologies available for genetic analysis on the microscale, and efforts to integrate them to form fully functional robust analysis devices. (topical review)

  18. Multidimensional proteomics analysis of amniotic fluid to provide insight into the mechanisms of idiopathic preterm birth.

    Directory of Open Access Journals (Sweden)

    Irina A Buhimschi

    2008-04-01

    Full Text Available Though recent advancement in proteomics has provided a novel perspective on several distinct pathogenetic mechanisms leading to preterm birth (inflammation, bleeding, the etiology of most preterm births still remains elusive. We conducted a multidimensional proteomic analysis of the amniotic fluid to identify pathways related to preterm birth in the absence of inflammation or bleeding.A proteomic fingerprint was generated from fresh amniotic fluid using surface-enhanced laser desorbtion ionization time of flight (SELDI-TOF mass spectrometry in a total of 286 consecutive samples retrieved from women who presented with signs or symptoms of preterm labor or preterm premature rupture of the membranes. Inflammation and/or bleeding proteomic patterns were detected in 32% (92/286 of the SELDI tracings. In the remaining tracings, a hierarchical algorithm was applied based on descriptors quantifying similarity/dissimilarity among proteomic fingerprints. This allowed identification of a novel profile (Q-profile based on the presence of 5 SELDI peaks in the 10-12.5 kDa mass area. Women displaying the Q-profile (mean+/-SD, gestational age: 25+/-4 weeks, n = 40 were more likely to deliver preterm despite expectant management in the context of intact membranes and normal amniotic fluid clinical results. Utilizing identification-centered proteomics techniques (fluorescence two-dimensional differential gel electrophoresis, robotic tryptic digestion and mass spectrometry coupled with Protein ANalysis THrough Evolutionary Relationships (PANTHER ontological classifications, we determined that in amniotic fluids with Q-profile the differentially expressed proteins are primarily involved in non-inflammatory biological processes such as protein metabolism, signal transduction and transport.Proteomic profiling of amniotic fluid coupled with non-hierarchical bioinformatics algorithms identified a subgroup of patients at risk for preterm birth in the absence of intra

  19. Integrated Proteomic Pipeline Using Multiple Search Engines for a Proteogenomic Study with a Controlled Protein False Discovery Rate.

    Science.gov (United States)

    Park, Gun Wook; Hwang, Heeyoun; Kim, Kwang Hoe; Lee, Ju Yeon; Lee, Hyun Kyoung; Park, Ji Yeong; Ji, Eun Sun; Park, Sung-Kyu Robin; Yates, John R; Kwon, Kyung-Hoon; Park, Young Mok; Lee, Hyoung-Joo; Paik, Young-Ki; Kim, Jin Young; Yoo, Jong Shin

    2016-11-04

    In the Chromosome-Centric Human Proteome Project (C-HPP), false-positive identification by peptide spectrum matches (PSMs) after database searches is a major issue for proteogenomic studies using liquid-chromatography and mass-spectrometry-based large proteomic profiling. Here we developed a simple strategy for protein identification, with a controlled false discovery rate (FDR) at the protein level, using an integrated proteomic pipeline (IPP) that consists of four engrailed steps as follows. First, using three different search engines, SEQUEST, MASCOT, and MS-GF+, individual proteomic searches were performed against the neXtProt database. Second, the search results from the PSMs were combined using statistical evaluation tools including DTASelect and Percolator. Third, the peptide search scores were converted into E-scores normalized using an in-house program. Last, ProteinInferencer was used to filter the proteins containing two or more peptides with a controlled FDR of 1.0% at the protein level. Finally, we compared the performance of the IPP to a conventional proteomic pipeline (CPP) for protein identification using a controlled FDR of <1% at the protein level. Using the IPP, a total of 5756 proteins (vs 4453 using the CPP) including 477 alternative splicing variants (vs 182 using the CPP) were identified from human hippocampal tissue. In addition, a total of 10 missing proteins (vs 7 using the CPP) were identified with two or more unique peptides, and their tryptic peptides were validated using MS/MS spectral pattern from a repository database or their corresponding synthetic peptides. This study shows that the IPP effectively improved the identification of proteins, including alternative splicing variants and missing proteins, in human hippocampal tissues for the C-HPP. All RAW files used in this study were deposited in ProteomeXchange (PXD000395).

  20. 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

  1. Comparative proteomic and metabolomic analysis reveal the antiosteoporotic molecular mechanism of icariin from Epimedium brevicornu maxim.

    Science.gov (United States)

    Xue, Liming; Jiang, Yiping; Han, Ting; Zhang, Naidan; Qin, Luping; Xin, Hailiang; Zhang, Qiaoyan

    2016-11-04

    Icariin, a principal flavonoid glycoside of Epimedium brevicornu Maxim, has been widely proved to possess antiosteoporotic activity with promoting bone formation and decreasing bone resorption. However, the involving mechanisms remain unclear. To clear a global insight of signal pathways involved in anti-osteoporotic mechanism of icariin at proteins and metabolites level by integrating the proteomics and NMR metabonomics, in a systems biology approach. Mice were divided into sham, OVX model and icariin-treated OVX group, after 90 days treatment, difference gel electrophoresis combined with MALDI-TOF/TOF proteomics analysis on bone femur and serum metabolomics were carried out for monitor intracellular processes and elucidate anti-osteoporotic mechanism of icariin. Osteoblast and osteoclast were applied to evaluate the potential signal pathways. Twenty three proteins in bone femur, and 8 metabolites in serum, were significantly altered and identified, involving in bone remodeling, energy metabolism, cytoskeleton, lipid metabolism, MAPK signaling, Ca 2+ signaling et, al. Furthermore, animal experiment show icariin could enhance the BMD and BMC, decrease CTX-I level in ovariectomized mice. The mitochondrial membrane potential and the intracellular ATP levels were increased significantly, and the cytoskeleton were improved in icariin-treatment osteoblast and osteoclast. Icariin also increased mRNA expression of Runx2 and osterix of OB, decreased CTR and CAII mRNA expression and protein expression of P38 and JNK. However, icariin did not reveal any inhibition of the collagenolytic activity of cathepsin K, mRNA expression of MMP-9 and protein expression of ERK in osteoclast. we consider icariin as multi-targeting compounds for treating with osteoporosis, involve initiating osteoblastogenesis, inhibiting adipogenesis, and preventing osteoclast differentiation. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  2. Centrosome isolation and analysis by mass spectrometry-based proteomics

    DEFF Research Database (Denmark)

    Jakobsen, Lis; Schrøder, Jacob Morville; Larsen, Katja M

    2013-01-01

    Centrioles are microtubule-based scaffolds that are essential for the formation of centrosomes, cilia, and flagella with important functions throughout the cell cycle, in physiology and during development. The ability to purify centriole-containing organelles on a large scale, combined with advan...... to isolate centrosomes from human cells and strategies to selectively identify and study the properties of the associated proteins using quantitative mass spectrometry-based proteomics.......Centrioles are microtubule-based scaffolds that are essential for the formation of centrosomes, cilia, and flagella with important functions throughout the cell cycle, in physiology and during development. The ability to purify centriole-containing organelles on a large scale, combined...... with advances in protein identification using mass spectrometry-based proteomics, have revealed multiple centriole-associated proteins that are conserved during evolution in eukaryotes. Despite these advances, the molecular basis for the plethora of processes coordinated by cilia and centrosomes is not fully...

  3. [Proteome analysis on interaction between Anoectochilus roxburghii and Mycorrhizal fungus].

    Science.gov (United States)

    Gao, Chuan; Guo, Shun-Xing; Zhang, Jing; Chen, Juan; Zhang, Li-Chun

    2012-12-01

    To study the mechanism of plant growing promoted by Mycorrhizal fungus through the difference of proteomes. The differential proteomes between uninoculated and inoculated endophytic fungi, Epulorhiza sp. on Anoectochilus roxburghii were analyzed by two-dimensional gel electrophoresis and MALDI-TOF/TOF mass spectrum. Twenty-seven protein spots were analyzed by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS). Twenty-two candidate proteins were identified by database comparisons. The function of these proteins mostly involved in signal transduction, metabolic regulation, as well as photosynthesis and substance metabolism. The results indicate that the regulator control system of plant is influenced by fungi action, and the positive regulation improves substance metabolism and photosynthesis, which results in strong plant and higher resistance. It is also deduced that silent genes may exist in endosymbiosis plants.

  4. Current perspectives in proteomic analysis of abiotic stress in Grapevines

    Directory of Open Access Journals (Sweden)

    Iniga Seraphina George

    2014-12-01

    Full Text Available Grapes are an important crop plant which forms the basis of a globally important industry. Grape and wine production is particularly vulnerable to environmental and climatic fluctuations, which makes it essential for us to develop a greater understanding of the molecular level responses of grape plants to various abiotic stresses. The completion of the initial grape genome sequence in 2007 has led to a significant increase in research on grapes using proteomics approaches. In this article, we discuss some of the current research on abiotic stress in grapevines, in the context of abiotic stress research in other plant species. We also highlight some of the current limitations in grapevine proteomics and identify areas with promising scope for potential future research.

  5. Analysis of the plasmodium falciparum proteome by high-accuracy mass spectrometry

    DEFF Research Database (Denmark)

    Lasonder, Edwin; Ishihama, Yasushi; Andersen, Jens S

    2002-01-01

    -accuracy (average deviation less than 0.02 Da at 1,000 Da) mass spectrometric proteome analysis of selected stages of the human malaria parasite Plasmodium falciparum. The analysis revealed 1,289 proteins of which 714 proteins were identified in asexual blood stages, 931 in gametocytes and 645 in gametes. The last...

  6. Analysis of proteomic changes of the serum of irradiated mice

    International Nuclear Information System (INIS)

    Wang Zhidong; Chen Xiaohua; Dong Bo; Zhang Junquan; Rao Yalan; Gao Ronglian; Hou Lili; Mao Bingzhi

    2005-01-01

    To explore the early diagnostic factors, new therapeutic targets and mechanisms of acute radiation disease. Proteomic changes of the serum of irradiated mice were studied using 2-DE and Q-TOF-MS approaches. One higher level expressed protein after the irradiation was found, and it was identified as α chain of haptoglobin by Q-TOF-MS. The authors confirmed the result by Western blotting with anti-haptoglobin antibody. Haptoglobin may involve in the process of acute radiation injury. (authors)

  7. Proteome Analysis of Borrelia burgdorferi Response to Environmental Change

    Energy Technology Data Exchange (ETDEWEB)

    Angel, Thomas E.; Luft, Benjamin J.; Yang, Xiaohua; Nicora, Carrie D.; Camp, David G.; Jacobs, Jon M.; Smith, Richard D.

    2010-11-02

    We examined global changes in protein expression in the B31 strain of Borrelia burgdorferi, in response to two environmental cues (pH and temperature) chosen for their reported similarity to those encountered at different stages of the organism’s life cycle. Multidimensional nano-liquid chromatographic separations coupled with tandem mass spectrometry were used to examine the array of proteins (i.e., the proteome) of B. burgdorferi for different pH and temperature culture conditions. Changes in pH and temperature elicited in vitro adaptations of this spirochete known to cause Lyme disease and led to alterations in protein expression that are associated with increased microbial pathogenesis. We identified 1031 proteins that represent 59% of the annotated genome of B. burgdorferi and elucidated a core proteome of 414 proteins that were present in all environmental conditions investigated. Observed changes in protein abundances indicated varied replicon usage, as well as proteome functional distributions between the in vitro cell culture conditions. Surprisingly, the pH and temperature conditions that mimicked B. burgdorferi residing in the gut of a fed tick showed a marked reduction in protein diversity. Additionally, the results provide us with leading candidates for exploring how B. burgdorferi adapts to and is able to survive in a wide variety of environmental conditions and lay a foundation for planned in situ studies of B. burgdorferi isolated from the tick midgut and infected animals.

  8. Proteomic analysis uncovers a metabolic phenotype in C. elegans after nhr-40 reduction of function

    International Nuclear Information System (INIS)

    Pohludka, Michal; Simeckova, Katerina; Vohanka, Jaroslav; Yilma, Petr; Novak, Petr; Krause, Michael W.; Kostrouchova, Marta; Kostrouch, Zdenek

    2008-01-01

    Caenorhabditis elegans has an unexpectedly large number (284) of genes encoding nuclear hormone receptors, most of which are nematode-specific and are of unknown function. We have exploited comparative two-dimensional chromatography of synchronized cultures of wild type C. elegans larvae and a mutant in nhr-40 to determine if proteomic approaches will provide additional insight into gene function. Chromatofocusing, followed by reversed-phase chromatography and mass spectrometry, identified altered chromatographic patterns for a set of proteins, many of which function in muscle and metabolism. Prompted by the proteomic analysis, we find that the penetrance of the developmental phenotypes in the mutant is enhanced at low temperatures and by food restriction. The combination of our phenotypic and proteomic analysis strongly suggests that NHR-40 provides a link between metabolism and muscle development. Our results highlight the utility of comparative two-dimensional chromatography to provide a relatively rapid method to gain insight into gene function

  9. RAPID PROCESSING OF ARCHIVAL TISSUE SAMPLES FOR PROTEOMIC ANALYSIS USING PRESSURE-CYCLING TECHNOLOGY

    Directory of Open Access Journals (Sweden)

    Vinuth N. Puttamallesh1,2

    2017-06-01

    Full Text Available Advent of mass spectrometry based proteomics has revolutionized our ability to study proteins from biological specimen in a high-throughput manner. Unlike cell line based studies, biomedical research involving tissue specimen is often challenging due to limited sample availability. In addition, investigation of clinically relevant research questions often requires enormous amount of time for sample collection prospectively. Formalin fixed paraffin embedded (FFPE archived tissue samples are a rich source of tissue specimen for biomedical research. However, there are several challenges associated with analysing FFPE samples. Protein cross-linking and degradation of proteins particularly affects proteomic analysis. We demonstrate that barocycler that uses pressure-cycling technology enables efficient protein extraction and processing of small amounts of FFPE tissue samples for proteomic analysis. We identified 3,525 proteins from six 10µm esophageal squamous cell carcinoma (ESCC tissue sections. Barocycler allows efficient protein extraction and proteolytic digestion of proteins from FFPE tissue sections at par with conventional methods.

  10. Selective proteomic analysis of antibiotic-tolerant cellular subpopulations in pseudomonas aeruginosa biofilms

    DEFF Research Database (Denmark)

    Babin, Brett M.; Atangcho, Lydia; van Eldijk, Mark B.

    2017-01-01

    involved in central carbon metabolism. We differentiated the immediate proteomic response, characterized by an increase in flagellar motility, from the long-term adaptive strategy, which included the upregulation of purine synthesis. This targeted, selective analysis of a bacterial subpopulation...... amino acid tagging (BONCAT) method to enable selective proteomic analysis of a Pseudomonas aeruginosa biofilm subpopulation. Through controlled expression of a mutant methionyl-tRNA synthetase, we targeted BONCAT labeling to cells in the regions of biofilm microcolonies that showed increased tolerance...... demonstrates how the study of proteome dynamics can enhance our understanding of biofilm heterogeneity and antibiotic tolerance. IMPORTANCE Bacterial growth is frequently characterized by behavioral heterogeneity at the single-cell level. Heterogeneity is especially evident in the physiology of biofilms...

  11. Proteomic analysis of three gonad types of swamp eel reveals genes differentially expressed during sex reversal.

    Science.gov (United States)

    Sheng, Yue; Zhao, Wei; Song, Ying; Li, Zhigang; Luo, Majing; Lei, Quan; Cheng, Hanhua; Zhou, Rongjia

    2015-05-18

    A variety of mechanisms are engaged in sex determination in vertebrates. The teleost fish swamp eel undergoes sex reversal naturally and is an ideal model for vertebrate sexual development. However, the importance of proteome-wide scanning for gonad reversal was not previously determined. We report a 2-D electrophoresis analysis of three gonad types of proteomes during sex reversal. MS/MS analysis revealed a group of differentially expressed proteins during ovary to ovotestis to testis transformation. Cbx3 is up-regulated during gonad reversal and is likely to have a role in spermatogenesis. Rab37 is down-regulated during the reversal and is mainly associated with oogenesis. Both Cbx3 and Rab37 are linked up in a protein network. These datasets in gonadal proteomes provide a new resource for further studies in gonadal development.

  12. Integrated Network Analysis and Effective Tools in Plant Systems Biology

    Directory of Open Access Journals (Sweden)

    Atsushi eFukushima

    2014-11-01

    Full Text Available One of the ultimate goals in plant systems biology is to elucidate the genotype-phenotype relationship in plant cellular systems. Integrated network analysis that combines omics data with mathematical models has received particular attention. Here we focus on the latest cutting-edge computational advances that facilitate their combination. We highlight (1 network visualization tools, (2 pathway analyses, (3 genome-scale metabolic reconstruction, and (4 the integration of high-throughput experimental data and mathematical models. Multi-omics data that contain the genome, transcriptome, proteome, and metabolome and mathematical models are expected to integrate and expand our knowledge of complex plant metabolisms.

  13. Network analysis of quantitative proteomics on asthmatic bronchi: effects of inhaled glucocorticoid treatment

    Directory of Open Access Journals (Sweden)

    Sihlbom Carina

    2011-09-01

    Full Text Available Abstract Background Proteomic studies of respiratory disorders have the potential to identify protein biomarkers for diagnosis and disease monitoring. Utilisation of sensitive quantitative proteomic methods creates opportunities to determine individual patient proteomes. The aim of the current study was to determine if quantitative proteomics of bronchial biopsies from asthmatics can distinguish relevant biological functions and whether inhaled glucocorticoid treatment affects these functions. Methods Endobronchial biopsies were taken from untreated asthmatic patients (n = 12 and healthy controls (n = 3. Asthmatic patients were randomised to double blind treatment with either placebo or budesonide (800 μg daily for 3 months and new biopsies were obtained. Proteins extracted from the biopsies were digested and analysed using isobaric tags for relative and absolute quantitation combined with a nanoLC-LTQ Orbitrap mass spectrometer. Spectra obtained were used to identify and quantify proteins. Pathways analysis was performed using Ingenuity Pathway Analysis to identify significant biological pathways in asthma and determine how the expression of these pathways was changed by treatment. Results More than 1800 proteins were identified and quantified in the bronchial biopsies of subjects. The pathway analysis revealed acute phase response signalling, cell-to-cell signalling and tissue development associations with proteins expressed in asthmatics compared to controls. The functions and pathways associated with placebo and budesonide treatment showed distinct differences, including the decreased association with acute phase proteins as a result of budesonide treatment compared to placebo. Conclusions Proteomic analysis of bronchial biopsy material can be used to identify and quantify proteins using highly sensitive technologies, without the need for pooling of samples from several patients. Distinct pathophysiological features of asthma can be

  14. Mass spectrometry-based serum proteome pattern analysis in molecular diagnostics of early stage breast cancer

    Directory of Open Access Journals (Sweden)

    Stobiecki Maciej

    2009-07-01

    Full Text Available Abstract Background Mass spectrometric analysis of the blood proteome is an emerging method of clinical proteomics. The approach exploiting multi-protein/peptide sets (fingerprints detected by mass spectrometry that reflect overall features of a specimen's proteome, termed proteome pattern analysis, have been already shown in several studies to have applicability in cancer diagnostics. We aimed to identify serum proteome patterns specific for early stage breast cancer patients using MALDI-ToF mass spectrometry. Methods Blood samples were collected before the start of therapy in a group of 92 patients diagnosed at stages I and II of the disease, and in a group of age-matched healthy controls (104 women. Serum specimens were purified and the low-molecular-weight proteome fraction was examined using MALDI-ToF mass spectrometry after removal of albumin and other high-molecular-weight serum proteins. Protein ions registered in a mass range between 2,000 and 10,000 Da were analyzed using a new bioinformatic tool created in our group, which included modeling spectra as a sum of Gaussian bell-shaped curves. Results We have identified features of serum proteome patterns that were significantly different between blood samples of healthy individuals and early stage breast cancer patients. The classifier built of three spectral components that differentiated controls and cancer patients had 83% sensitivity and 85% specificity. Spectral components (i.e., protein ions that were the most frequent in such classifiers had approximate m/z values of 2303, 2866 and 3579 Da (a biomarker built from these three components showed 88% sensitivity and 78% specificity. Of note, we did not find a significant correlation between features of serum proteome patterns and established prognostic or predictive factors like tumor size, nodal involvement, histopathological grade, estrogen and progesterone receptor expression. In addition, we observed a significantly (p = 0

  15. GProX, a User-Friendly Platform for Bioinformatics Analysis and Visualization of Quantitative Proteomics Data

    DEFF Research Database (Denmark)

    Rigbolt, Kristoffer T G; Vanselow, Jens T; Blagoev, Blagoy

    2011-01-01

    -friendly platform for comprehensive analysis, inspection and visualization of quantitative proteomics data we developed the Graphical Proteomics Data Explorer (GProX)(1). The program requires no special bioinformatics training, as all functions of GProX are accessible within its graphical user-friendly interface...... such as database querying, clustering based on abundance ratios, feature enrichment tests for e.g. GO terms and pathway analysis tools. A number of plotting options for visualization of quantitative proteomics data is available and most analysis functions in GProX create customizable high quality graphical...... displays in both vector and bitmap formats. The generic import requirements allow data originating from essentially all mass spectrometry platforms, quantitation strategies and software to be analyzed in the program. GProX represents a powerful approach to proteomics data analysis providing proteomics...

  16. Proteomic analysis during larval development and metamorphosis of the spionid polychaete Pseudopolydora vexillosa

    KAUST Repository

    Mok, Flora SY; Thiyagarajan, Vengatesen; Qian, Pei-Yuan

    2009-01-01

    Background: While the larval-juvenile transition (metamorphosis) in the spionid polychaete Pseudopolydora vexillosa involves gradual morphological changes and does not require substantial development of juvenile organs, the opposite occurs in the barnacle Balanus amphitrite. We hypothesized that the proteome changes during metamorphosis in the spionids are less drastic than that in the barnacles. To test this, proteomes of pre-competent larvae, competent larvae (ready to metamorphose), and juveniles of P. vexillosa were compared using 2-dimensional gel electrophoresis (2-DE), and they were then compared to those of the barnacle.Results: Unlike the significant changes found during barnacle metamorphosis, proteomes of competent P. vexillosa larvae were more similar to those of their juveniles. Pre-competent larvae had significantly fewer protein spots (384 spots), while both competent larvae and juveniles expressed about 660 protein spots each. Proteins up-regulated during competence identified by MALDI-TOF/TOF analysis included a molecular chaperon (calreticulin), a signal transduction regulator (tyrosin activation protein), and a tissue-remodeling enzyme (metallopeptidase).Conclusions: This was the first time to study the protein expression patterns during the metamorphosis of a marine polychaete and to compare the proteomes of marine invertebrates that have different levels of morphological changes during metamorphosis. The findings provide promising initial steps towards the development of a proteome database for marine invertebrate metamorphosis, thus deciphering the possible mechanisms underlying larval metamorphosis in non-model marine organisms. © 2009 Mok et al; licensee BioMed Central Ltd.

  17. Proteomic analysis during larval development and metamorphosis of the spionid polychaete Pseudopolydora vexillosa

    KAUST Repository

    Mok, Flora SY

    2009-12-14

    Background: While the larval-juvenile transition (metamorphosis) in the spionid polychaete Pseudopolydora vexillosa involves gradual morphological changes and does not require substantial development of juvenile organs, the opposite occurs in the barnacle Balanus amphitrite. We hypothesized that the proteome changes during metamorphosis in the spionids are less drastic than that in the barnacles. To test this, proteomes of pre-competent larvae, competent larvae (ready to metamorphose), and juveniles of P. vexillosa were compared using 2-dimensional gel electrophoresis (2-DE), and they were then compared to those of the barnacle.Results: Unlike the significant changes found during barnacle metamorphosis, proteomes of competent P. vexillosa larvae were more similar to those of their juveniles. Pre-competent larvae had significantly fewer protein spots (384 spots), while both competent larvae and juveniles expressed about 660 protein spots each. Proteins up-regulated during competence identified by MALDI-TOF/TOF analysis included a molecular chaperon (calreticulin), a signal transduction regulator (tyrosin activation protein), and a tissue-remodeling enzyme (metallopeptidase).Conclusions: This was the first time to study the protein expression patterns during the metamorphosis of a marine polychaete and to compare the proteomes of marine invertebrates that have different levels of morphological changes during metamorphosis. The findings provide promising initial steps towards the development of a proteome database for marine invertebrate metamorphosis, thus deciphering the possible mechanisms underlying larval metamorphosis in non-model marine organisms. © 2009 Mok et al; licensee BioMed Central Ltd.

  18. An Automated High Throughput Proteolysis and Desalting Platform for Quantitative Proteomic Analysis

    Directory of Open Access Journals (Sweden)

    Albert-Baskar Arul

    2013-06-01

    Full Text Available Proteomics for biomarker validation needs high throughput instrumentation to analyze huge set of clinical samples for quantitative and reproducible analysis at a minimum time without manual experimental errors. Sample preparation, a vital step in proteomics plays a major role in identification and quantification of proteins from biological samples. Tryptic digestion a major check point in sample preparation for mass spectrometry based proteomics needs to be more accurate with rapid processing time. The present study focuses on establishing a high throughput automated online system for proteolytic digestion and desalting of proteins from biological samples quantitatively and qualitatively in a reproducible manner. The present study compares online protein digestion and desalting of BSA with conventional off-line (in-solution method and validated for real time sample for reproducibility. Proteins were identified using SEQUEST data base search engine and the data were quantified using IDEALQ software. The present study shows that the online system capable of handling high throughput samples in 96 well formats carries out protein digestion and peptide desalting efficiently in a reproducible and quantitative manner. Label free quantification showed clear increase of peptide quantities with increase in concentration with much linearity compared to off line method. Hence we would like to suggest that inclusion of this online system in proteomic pipeline will be effective in quantification of proteins in comparative proteomics were the quantification is really very crucial.

  19. A Pilot Proteomic Analysis of Salivary Biomarkers in Autism Spectrum Disorder.

    Science.gov (United States)

    Ngounou Wetie, Armand G; Wormwood, Kelly L; Russell, Stefanie; Ryan, Jeanne P; Darie, Costel C; Woods, Alisa G

    2015-06-01

    Autism spectrum disorder (ASD) prevalence is increasing, with current estimates at 1/68-1/50 individuals diagnosed with an ASD. Diagnosis is based on behavioral assessments. Early diagnosis and intervention is known to greatly improve functional outcomes in people with ASD. Diagnosis, treatment monitoring and prognosis of ASD symptoms could be facilitated with biomarkers to complement behavioral assessments. Mass spectrometry (MS) based proteomics may help reveal biomarkers for ASD. In this pilot study, we have analyzed the salivary proteome in individuals with ASD compared to neurotypical control subjects, using MS-based proteomics. Our goal is to optimize methods for salivary proteomic biomarker discovery and to identify initial putative biomarkers in people with ASDs. The salivary proteome is virtually unstudied in ASD, and saliva could provide an easily accessible biomaterial for analysis. Using nano liquid chromatography-tandem mass spectrometry, we found statistically significant differences in several salivary proteins, including elevated prolactin-inducible protein, lactotransferrin, Ig kappa chain C region, Ig gamma-1 chain C region, Ig lambda-2 chain C regions, neutrophil elastase, polymeric immunoglobulin receptor and deleted in malignant brain tumors 1. Our results indicate that this is an effective method for identification of salivary protein biomarkers, support the concept that immune system and gastrointestinal disturbances may be present in individuals with ASDs and point toward the need for larger studies in behaviorally-characterized individuals. © 2015 International Society for Autism Research, Wiley Periodicals, Inc.

  20. Data from proteomic analysis of the skin of Chinese giant salamander (Andrias davidianus

    Directory of Open Access Journals (Sweden)

    Xiaofang Geng

    2015-06-01

    Full Text Available The Chinese giant salamander (Andrias davidianus, renowned as a living fossil, is the largest and longest-lived amphibian species in the world. Its skin is rich in collagens, and has developed mucous gland which could secrete a large amount of mucus under the scraping and electric stimulation. The molting is the degraded skin stratum corneum. To establish the functional skin proteome of Chinese giant salamander, two-dimensional gel electrophoresis (2DE and mass spectrometry (MS were applied to detect the composition and relative abundance of the proteins in the skin, mucus and molting. The determination of the general proteome in the skin can potentially serve as a foundation for future studies characterizing the skin proteomes from diseased salamander to provide molecular and mechanistic insights into various disease states and potential therapeutic interventions. Data presented here are also related to the research article “Proteomic analysis of the skin of Chinese giant salamander (Andrias davidianus” in the Journal of Proteomics [1].

  1. Proteomic analysis during larval development and metamorphosis of the spionid polychaete Pseudopolydora vexillosa

    Directory of Open Access Journals (Sweden)

    Qian Pei-Yuan

    2009-12-01

    Full Text Available Abstract Background While the larval-juvenile transition (metamorphosis in the spionid polychaete Pseudopolydora vexillosa involves gradual morphological changes and does not require substantial development of juvenile organs, the opposite occurs in the barnacle Balanus amphitrite. We hypothesized that the proteome changes during metamorphosis in the spionids are less drastic than that in the barnacles. To test this, proteomes of pre-competent larvae, competent larvae (ready to metamorphose, and juveniles of P. vexillosa were compared using 2-dimensional gel electrophoresis (2-DE, and they were then compared to those of the barnacle. Results Unlike the significant changes found during barnacle metamorphosis, proteomes of competent P. vexillosa larvae were more similar to those of their juveniles. Pre-competent larvae had significantly fewer protein spots (384 spots, while both competent larvae and juveniles expressed about 660 protein spots each. Proteins up-regulated during competence identified by MALDI-TOF/TOF analysis included a molecular chaperon (calreticulin, a signal transduction regulator (tyrosin activation protein, and a tissue-remodeling enzyme (metallopeptidase. Conclusions This was the first time to study the protein expression patterns during the metamorphosis of a marine polychaete and to compare the proteomes of marine invertebrates that have different levels of morphological changes during metamorphosis. The findings provide promising initial steps towards the development of a proteome database for marine invertebrate metamorphosis, thus deciphering the possible mechanisms underlying larval metamorphosis in non-model marine organisms.

  2. Bringing metabolic networks to life: integration of kinetic, metabolic, and proteomic data

    Directory of Open Access Journals (Sweden)

    Klipp Edda

    2006-12-01

    Full Text Available Abstract Background Translating a known metabolic network into a dynamic model requires reasonable guesses of all enzyme parameters. In Bayesian parameter estimation, model parameters are described by a posterior probability distribution, which scores the potential parameter sets, showing how well each of them agrees with the data and with the prior assumptions made. Results We compute posterior distributions of kinetic parameters within a Bayesian framework, based on integration of kinetic, thermodynamic, metabolic, and proteomic data. The structure of the metabolic system (i.e., stoichiometries and enzyme regulation needs to be known, and the reactions are modelled by convenience kinetics with thermodynamically independent parameters. The parameter posterior is computed in two separate steps: a first posterior summarises the available data on enzyme kinetic parameters; an improved second posterior is obtained by integrating metabolic fluxes, concentrations, and enzyme concentrations for one or more steady states. The data can be heterogenous, incomplete, and uncertain, and the posterior is approximated by a multivariate log-normal distribution. We apply the method to a model of the threonine synthesis pathway: the integration of metabolic data has little effect on the marginal posterior distributions of individual model parameters. Nevertheless, it leads to strong correlations between the parameters in the joint posterior distribution, which greatly improve the model predictions by the following Monte-Carlo simulations. Conclusion We present a standardised method to translate metabolic networks into dynamic models. To determine the model parameters, evidence from various experimental data is combined and weighted using Bayesian parameter estimation. The resulting posterior parameter distribution describes a statistical ensemble of parameter sets; the parameter variances and correlations can account for missing knowledge, measurement

  3. A single lysis solution for the analysis of tissue samples by different proteomic technologies

    DEFF Research Database (Denmark)

    Gromov, P.; Celis, J.E.; Gromova, I.

    2008-01-01

    -based proteomics (reverse-phase lysate arrays or direct antibody arrays), allowing the direct comparison of qualitative and quantitative data yielded by these technologies when applied to the same samples. The usefulness of the CLB1 solution for gel-based proteomics was further established by 2D PAGE analysis...... dissease, is driving scientists to increasingly use clinically relevant samples for biomarker and target discovery. Tissues are heterogeneous and as a result optimization of sample preparation is critical for generating accurate, representative, and highly reproducible quantitative data. Although a large...... number of protocols for preparation of tissue lysates has been published, so far no single recipe is able to provide a "one-size fits all" solubilization procedure that can be used to analyse the same lysate using different proteomics technologies. Here we present evidence showing that cell lysis buffer...

  4. P-MartCancer–Interactive Online Software to Enable Analysis of Shotgun Cancer Proteomic Datasets

    Energy Technology Data Exchange (ETDEWEB)

    Webb-Robertson, Bobbie-Jo M.; Bramer, Lisa M.; Jensen, Jeffrey L.; Kobold, Markus A.; Stratton, Kelly G.; White, Amanda M.; Rodland, Karin D.

    2017-10-31

    P-MartCancer is a new interactive web-based software environment that enables biomedical and biological scientists to perform in-depth analyses of global proteomics data without requiring direct interaction with the data or with statistical software. P-MartCancer offers a series of statistical modules associated with quality assessment, peptide and protein statistics, protein quantification and exploratory data analyses driven by the user via customized workflows and interactive visualization. Currently, P-MartCancer offers access to multiple cancer proteomic datasets generated through the Clinical Proteomics Tumor Analysis Consortium (CPTAC) at the peptide, gene and protein levels. P-MartCancer is deployed using Azure technologies (http://pmart.labworks.org/cptac.html), the web-service is alternatively available via Docker Hub (https://hub.docker.com/r/pnnl/pmart-web/) and many statistical functions can be utilized directly from an R package available on GitHub (https://github.com/pmartR).

  5. P-MartCancer-Interactive Online Software to Enable Analysis of Shotgun Cancer Proteomic Datasets.

    Science.gov (United States)

    Webb-Robertson, Bobbie-Jo M; Bramer, Lisa M; Jensen, Jeffrey L; Kobold, Markus A; Stratton, Kelly G; White, Amanda M; Rodland, Karin D

    2017-11-01

    P-MartCancer is an interactive web-based software environment that enables statistical analyses of peptide or protein data, quantitated from mass spectrometry-based global proteomics experiments, without requiring in-depth knowledge of statistical programming. P-MartCancer offers a series of statistical modules associated with quality assessment, peptide and protein statistics, protein quantification, and exploratory data analyses driven by the user via customized workflows and interactive visualization. Currently, P-MartCancer offers access and the capability to analyze multiple cancer proteomic datasets generated through the Clinical Proteomics Tumor Analysis Consortium at the peptide, gene, and protein levels. P-MartCancer is deployed as a web service (https://pmart.labworks.org/cptac.html), alternatively available via Docker Hub (https://hub.docker.com/r/pnnl/pmart-web/). Cancer Res; 77(21); e47-50. ©2017 AACR . ©2017 American Association for Cancer Research.

  6. Global analysis of the yeast osmotic stress response by quantitative proteomics

    DEFF Research Database (Denmark)

    Soufi, Boumediene; Kelstrup, C.D.; Stoehr, G.

    2009-01-01

    a comprehensive, quantitative, and time-resolved analysis using high-resolution mass spectrometry of phospho-proteome and proteome changes in response to osmotic stress in yeast. We identified 5534 unique phosphopeptide variants and 3383 yeast proteins. More than 15% of the detected phosphorylation site status...... changed more than two-fold within 5 minutes of treatment. Many of the corresponding phosphoproteins are involved in the early response to environmental stress. Surprisingly, we find that 158 regulated phosphorylation sites are potential substrates of basophilic kinases as opposed to the classical proline......-directed MAP kinase network implicated in stress response mechanisms such as p38 and HOG pathways. Proteome changes reveal an increase in abundance of more than one hundred proteins after 20 min of salt stress. Many of these are involved in the cellular response to increased osmolarity, which include proteins...

  7. Data set for the proteomic inventory and quantitative analysis of chicken uterine fluid during eggshell biomineralization

    Directory of Open Access Journals (Sweden)

    Pauline Marie

    2014-12-01

    Full Text Available Chicken eggshell is the protective barrier of the egg. It is a biomineral composed of 95% calcium carbonate on calcitic form and 3.5% organic matrix proteins. Mineralization process occurs in uterus into the uterine fluid. This acellular fluid contains ions and organic matrix proteins precursors which are interacting with the mineral phase and control crystal growth, eggshell structure and mechanical properties. We performed a proteomic approach and identified 308 uterine fluid proteins. Gene Ontology terms enrichments were determined to investigate their potential functions. Mass spectrometry analyses were also combined to label free quantitative analysis to determine the relative abundance of 96 proteins at initiation, rapid growth phase and termination of shell calcification. Sixty four showed differential abundance according to the mineralization stage. Their potential functions have been annotated. The complete proteomic, bioinformatic and functional analyses are reported in Marie et al., J. Proteomics (2015 [1].

  8. 2 D gel based analysis of biological variability of the human plasma proteome

    DEFF Research Database (Denmark)

    Rentsch, Maria Louise; Jessen, Flemming

    individuals and within an individual changes will also happen over time (e.g. after meal intake). Thus, the aim of the present study was to examine the inter-individual variability of plasma protein levels in humans after meal intake. Five subjects consumed three single meals in a randomised order separated...... by one-week interval. Blood samples were drawn before the meal intake and five times during 24 hours for proteome analysis. Plasma was fractionated by use of IgY-12 spin column depleting the 12 highly abundant proteins and further processed for two-dimensional gel electrophoresis. The plasma proteome...

  9. Proteome analysis of the purine stimulon from Lactococcus lactis

    DEFF Research Database (Denmark)

    Beyer, N.H.; Roepstorff, P.; Hammer, Karin

    2003-01-01

    kinase) and translation elongation factors (GTPases: EF-TU, EF-G). Two Dcu proteins could not be identified. Out of 28 proteins subjected to mass spectrometry, 19 could be readily identified despite the fact that only the genome sequence of a strain of L. lactis subsp. lactis was available. The two...... subspecies share about 85% sequence identity, comparable to the genetic distance between Escherichia coli and Salmonella typhimurium. A success rate of 68% indicates that it may be feasible to perform proteomics based upon genomic sequences of relatives outside the genus....

  10. Comparative proteome analysis of Brettanomyces bruxellensis under hydroxycinnamic acid growth

    Directory of Open Access Journals (Sweden)

    Lourdes Carmona

    2016-09-01

    Conclusions: The proteomic profile of B. bruxellensis cultivated in the presence of p-coumaric acid in synthetic wine, agrees with the hypothesis of metabolic flux regulation, allowing a better conditioning to an adverse environment. This study involved the translational level of B. bruxellensis in the production of ethylphenols and corroborated that this yeast presented an advantage in these stress conditions. Thus, this work will allow an understanding of the regulation and processes involved in the production of ethyl-derivate compounds by B. bruxellensis. Furthermore, it allows the development of newer and better techniques for spoilage yeast control.

  11. Proteomic analysis of protein phosphatase Z1 from Candida albicans.

    Directory of Open Access Journals (Sweden)

    Bernadett Márkus

    Full Text Available Protein phosphatase Z is a "novel type" fungus specific serine/threonine protein phosphatase. Previously our research group identified the CaPPZ1 gene in the opportunistic pathogen Candida albicans and reported that the gene deletion had several important physiological consequences. In order to reveal the protein targets and the associated mechanisms behind the functions of the phosphatase a proteomic method was adopted for the comparison of the cappz1 deletion mutant and the genetically matching QMY23 control strain. Proteins extracted from the control and deletion mutant strains were separated by two-dimensional gel electrophoresis and the protein spots were stained with RuBPS and Pro-Q Diamond in order to visualize the total proteome and the phosphoproteome, respectively. The alterations in spot intensities were determined by densitometry and were analysed with the Delta2D (Decodon software. Spots showing significantly different intensities between the mutant and control strains were excised from the gels and were digested with trypsin. The resulting peptides were identified by LC-MS/MS mass spectrometry. As many as 15 protein spots were found that exhibited significant changes in their intensity upon the deletion of the phosphatase and 20 phosphoproteins were identified in which the level of phosphorylation was modified significantly in the mutant. In agreement with previous findings we found that the affected proteins function in protein synthesis, oxidative stress response, regulation of morphology and metabolism. Among these proteins we identified two potential CaPpz1 substrates (Eft2 and Rpp0 that may regulate the elongation step of translation. RT-qPCR experiments revealed that the expression of the genes coding for the affected proteins was not altered significantly. Thus, the absence of CaPpz1 exerted its effects via protein synthesis/degradation and phosphorylation/dephosphorylation. In addition, our proteomics data strongly

  12. Proteomic analysis of protein phosphatase Z1 from Candida albicans

    Science.gov (United States)

    Pfliegler, Walter P.; Petrényi, Katalin; Boros, Enikő; Pócsi, István; Tőzsér, József; Dombrádi, Viktor

    2017-01-01

    Protein phosphatase Z is a “novel type” fungus specific serine/threonine protein phosphatase. Previously our research group identified the CaPPZ1 gene in the opportunistic pathogen Candida albicans and reported that the gene deletion had several important physiological consequences. In order to reveal the protein targets and the associated mechanisms behind the functions of the phosphatase a proteomic method was adopted for the comparison of the cappz1 deletion mutant and the genetically matching QMY23 control strain. Proteins extracted from the control and deletion mutant strains were separated by two-dimensional gel electrophoresis and the protein spots were stained with RuBPS and Pro-Q Diamond in order to visualize the total proteome and the phosphoproteome, respectively. The alterations in spot intensities were determined by densitometry and were analysed with the Delta2D (Decodon) software. Spots showing significantly different intensities between the mutant and control strains were excised from the gels and were digested with trypsin. The resulting peptides were identified by LC-MS/MS mass spectrometry. As many as 15 protein spots were found that exhibited significant changes in their intensity upon the deletion of the phosphatase and 20 phosphoproteins were identified in which the level of phosphorylation was modified significantly in the mutant. In agreement with previous findings we found that the affected proteins function in protein synthesis, oxidative stress response, regulation of morphology and metabolism. Among these proteins we identified two potential CaPpz1 substrates (Eft2 and Rpp0) that may regulate the elongation step of translation. RT-qPCR experiments revealed that the expression of the genes coding for the affected proteins was not altered significantly. Thus, the absence of CaPpz1 exerted its effects via protein synthesis/degradation and phosphorylation/dephosphorylation. In addition, our proteomics data strongly suggested a role for

  13. Integrative proteomics, genomics, and translational immunology approaches reveal mutated forms of Proteolipid Protein 1 (PLP1) and mutant-specific immune response in multiple sclerosis.

    Science.gov (United States)

    Qendro, Veneta; Bugos, Grace A; Lundgren, Debbie H; Glynn, John; Han, May H; Han, David K

    2017-03-01

    In order to gain mechanistic insights into multiple sclerosis (MS) pathogenesis, we utilized a multi-dimensional approach to test the hypothesis that mutations in myelin proteins lead to immune activation and central nervous system autoimmunity in MS. Mass spectrometry-based proteomic analysis of human MS brain lesions revealed seven unique mutations of PLP1; a key myelin protein that is known to be destroyed in MS. Surprisingly, in-depth genomic analysis of two MS patients at the genomic DNA and mRNA confirmed mutated PLP1 in RNA, but not in the genomic DNA. Quantification of wild type and mutant PLP RNA levels by qPCR further validated the presence of mutant PLP RNA in the MS patients. To seek evidence linking mutations in abundant myelin proteins and immune-mediated destruction of myelin, specific immune response against mutant PLP1 in MS patients was examined. Thus, we have designed paired, wild type and mutant peptide microarrays, and examined antibody response to multiple mutated PLP1 in sera from MS patients. Consistent with the idea of different patients exhibiting unique mutation profiles, we found that 13 out of 20 MS patients showed antibody responses against specific but not against all the mutant-PLP1 peptides. Interestingly, we found mutant PLP-directed antibody response against specific mutant peptides in the sera of pre-MS controls. The results from integrative proteomic, genomic, and immune analyses reveal a possible mechanism of mutation-driven pathogenesis in human MS. The study also highlights the need for integrative genomic and proteomic analyses for uncovering pathogenic mechanisms of human diseases. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  14. Proteomics Core

    Data.gov (United States)

    Federal Laboratory Consortium — Proteomics Core is the central resource for mass spectrometry based proteomics within the NHLBI. The Core staff help collaborators design proteomics experiments in a...

  15. Integrative proteomics and biochemical analyses define Ptc6p as the Saccharomyces cerevisiae pyruvate dehydrogenase phosphatase.

    Science.gov (United States)

    Guo, Xiao; Niemi, Natalie M; Coon, Joshua J; Pagliarini, David J

    2017-07-14

    The pyruvate dehydrogenase complex (PDC) is the primary metabolic checkpoint connecting glycolysis and mitochondrial oxidative phosphorylation and is important for maintaining cellular and organismal glucose homeostasis. Phosphorylation of the PDC E1 subunit was identified as a key inhibitory modification in bovine tissue ∼50 years ago, and this regulatory process is now known to be conserved throughout evolution. Although Saccharomyces cerevisiae is a pervasive model organism for investigating cellular metabolism and its regulation by signaling processes, the phosphatase(s) responsible for activating the PDC in S. cerevisiae has not been conclusively defined. Here, using comparative mitochondrial phosphoproteomics, analyses of protein-protein interactions by affinity enrichment-mass spectrometry, and in vitro biochemistry, we define Ptc6p as the primary PDC phosphatase in S. cerevisiae Our analyses further suggest additional substrates for related S. cerevisiae phosphatases and describe the overall phosphoproteomic changes that accompany mitochondrial respiratory dysfunction. In summary, our quantitative proteomics and biochemical analyses have identified Ptc6p as the primary-and likely sole- S. cerevisiae PDC phosphatase, closing a key knowledge gap about the regulation of yeast mitochondrial metabolism. Our findings highlight the power of integrative omics and biochemical analyses for annotating the functions of poorly characterized signaling proteins. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  16. Integrated proteomics and metabolomics suggests symbiotic metabolism and multimodal regulation in a fungal-endobacterial system.

    Science.gov (United States)

    Li, Zhou; Yao, Qiuming; Dearth, Stephen P; Entler, Matthew R; Castro Gonzalez, Hector F; Uehling, Jessie K; Vilgalys, Rytas J; Hurst, Gregory B; Campagna, Shawn R; Labbé, Jessy L; Pan, Chongle

    2017-03-01

    Many plant-associated fungi host endosymbiotic endobacteria with reduced genomes. While endobacteria play important roles in these tri-partite plant-fungal-endobacterial systems, the active physiology of fungal endobacteria has not been characterized extensively by systems biology approaches. Here, we use integrated proteomics and metabolomics to characterize the relationship between the endobacterium Mycoavidus sp. and the root-associated fungus Mortierella elongata. In nitrogen-poor media, M. elongata had decreased growth but hosted a large and growing endobacterial population. The active endobacterium likely extracted malate from the fungal host as the primary carbon substrate for energy production and biosynthesis of phospho-sugars, nucleobases, peptidoglycan and some amino acids. The endobacterium obtained nitrogen by importing a variety of nitrogen-containing compounds. Further, nitrogen limitation significantly perturbed the carbon and nitrogen flows in the fungal metabolic network. M. elongata regulated many pathways by concordant changes on enzyme abundances, post-translational modifications, reactant concentrations and allosteric effectors. Such multimodal regulations may be a general mechanism for metabolic modulation. © 2016 Society for Applied Microbiology and John Wiley & Sons Ltd.

  17. Integrative Identification of Arabidopsis Mitochondrial Proteome and Its Function Exploitation through Protein Interaction Network

    Science.gov (United States)

    Cui, Jian; Liu, Jinghua; Li, Yuhua; Shi, Tieliu

    2011-01-01

    Mitochondria are major players on the production of energy, and host several key reactions involved in basic metabolism and biosynthesis of essential molecules. Currently, the majority of nucleus-encoded mitochondrial proteins are unknown even for model plant Arabidopsis. We reported a computational framework for predicting Arabidopsis mitochondrial proteins based on a probabilistic model, called Naive Bayesian Network, which integrates disparate genomic data generated from eight bioinformatics tools, multiple orthologous mappings, protein domain properties and co-expression patterns using 1,027 microarray profiles. Through this approach, we predicted 2,311 candidate mitochondrial proteins with 84.67% accuracy and 2.53% FPR performances. Together with those experimental confirmed proteins, 2,585 mitochondria proteins (named CoreMitoP) were identified, we explored those proteins with unknown functions based on protein-protein interaction network (PIN) and annotated novel functions for 26.65% CoreMitoP proteins. Moreover, we found newly predicted mitochondrial proteins embedded in particular subnetworks of the PIN, mainly functioning in response to diverse environmental stresses, like salt, draught, cold, and wound etc. Candidate mitochondrial proteins involved in those physiological acitivites provide useful targets for further investigation. Assigned functions also provide comprehensive information for Arabidopsis mitochondrial proteome. PMID:21297957

  18. Functional proteomic analysis of corticosteroid pharmacodynamics in rat liver: Relationship to hepatic stress, signaling, energy regulation, and drug metabolism.

    Science.gov (United States)

    Ayyar, Vivaswath S; Almon, Richard R; DuBois, Debra C; Sukumaran, Siddharth; Qu, Jun; Jusko, William J

    2017-05-08

    Corticosteroids (CS) are anti-inflammatory agents that cause extensive pharmacogenomic and proteomic changes in multiple tissues. An understanding of the proteome-wide effects of CS in liver and its relationships to altered hepatic and systemic physiology remains incomplete. Here, we report the application of a functional pharmacoproteomic approach to gain integrated insight into the complex nature of CS responses in liver in vivo. An in-depth functional analysis was performed using rich pharmacodynamic (temporal-based) proteomic data measured over 66h in rat liver following a single dose of methylprednisolone (MPL). Data mining identified 451 differentially regulated proteins. These proteins were analyzed on the basis of temporal regulation, cellular localization, and literature-mined functional information. Of the 451 proteins, 378 were clustered into six functional groups based on major clinically-relevant effects of CS in liver. MPL-responsive proteins were highly localized in the mitochondria (20%) and cytosol (24%). Interestingly, several proteins were related to hepatic stress and signaling processes, which appear to be involved in secondary signaling cascades and in protecting the liver from CS-induced oxidative damage. Consistent with known adverse metabolic effects of CS, several rate-controlling enzymes involved in amino acid metabolism, gluconeogenesis, and fatty-acid metabolism were altered by MPL. In addition, proteins involved in the metabolism of endogenous compounds, xenobiotics, and therapeutic drugs including cytochrome P450 and Phase-II enzymes were differentially regulated. Proteins related to the inflammatory acute-phase response were up-regulated in response to MPL. Functionally-similar proteins showed large diversity in their temporal profiles, indicating complex mechanisms of regulation by CS. Clinical use of corticosteroid (CS) therapy is frequent and chronic. However, current knowledge on the proteome-level effects of CS in liver and

  19. Genomic, proteomic and biochemical analysis of the organohalide respiratory pathway in Desulfitobacterium dehalogenans

    NARCIS (Netherlands)

    Kruse, T.; Pas, van de B.A.; Atteia, A.; Krab, K.; Hagen, W.R.; Goodwin, L.; Chain, P.; Boeren, S.; Maphosa, F.; Schraa, G.; Vos, de W.M.; Oost, van der J.; Smidt, H.; Stams, A.J.M.

    2015-01-01

    Desulfitobacterium dehalogenans is able to grow by organohalide respiration using 3-chloro-4-hydroxyphenyl acetate (Cl-OHPA) as an electron acceptor. We used a combination of genome sequencing, biochemical analysis of redox active components and shotgun proteomics to study elements of the

  20. Integration of genomic, transcriptomic and proteomic data identifies two biologically distinct subtypes of invasive lobular breast cancer.

    Science.gov (United States)

    Michaut, Magali; Chin, Suet-Feung; Majewski, Ian; Severson, Tesa M; Bismeijer, Tycho; de Koning, Leanne; Peeters, Justine K; Schouten, Philip C; Rueda, Oscar M; Bosma, Astrid J; Tarrant, Finbarr; Fan, Yue; He, Beilei; Xue, Zheng; Mittempergher, Lorenza; Kluin, Roelof J C; Heijmans, Jeroen; Snel, Mireille; Pereira, Bernard; Schlicker, Andreas; Provenzano, Elena; Ali, Hamid Raza; Gaber, Alexander; O'Hurley, Gillian; Lehn, Sophie; Muris, Jettie J F; Wesseling, Jelle; Kay, Elaine; Sammut, Stephen John; Bardwell, Helen A; Barbet, Aurélie S; Bard, Floriane; Lecerf, Caroline; O'Connor, Darran P; Vis, Daniël J; Benes, Cyril H; McDermott, Ultan; Garnett, Mathew J; Simon, Iris M; Jirström, Karin; Dubois, Thierry; Linn, Sabine C; Gallagher, William M; Wessels, Lodewyk F A; Caldas, Carlos; Bernards, Rene

    2016-01-05

    Invasive lobular carcinoma (ILC) is the second most frequently occurring histological breast cancer subtype after invasive ductal carcinoma (IDC), accounting for around 10% of all breast cancers. The molecular processes that drive the development of ILC are still largely unknown. We have performed a comprehensive genomic, transcriptomic and proteomic analysis of a large ILC patient cohort and present here an integrated molecular portrait of ILC. Mutations in CDH1 and in the PI3K pathway are the most frequent molecular alterations in ILC. We identified two main subtypes of ILCs: (i) an immune related subtype with mRNA up-regulation of PD-L1, PD-1 and CTLA-4 and greater sensitivity to DNA-damaging agents in representative cell line models; (ii) a hormone related subtype, associated with Epithelial to Mesenchymal Transition (EMT), and gain of chromosomes 1q and 8q and loss of chromosome 11q. Using the somatic mutation rate and eIF4B protein level, we identified three groups with different clinical outcomes, including a group with extremely good prognosis. We provide a comprehensive overview of the molecular alterations driving ILC and have explored links with therapy response. This molecular characterization may help to tailor treatment of ILC through the application of specific targeted, chemo- and/or immune-therapies.

  1. Proteome analysis of Acetobacter pasteurianus during acetic acid fermentation.

    Science.gov (United States)

    Andrés-Barrao, Cristina; Saad, Maged M; Chappuis, Marie-Louise; Boffa, Mauro; Perret, Xavier; Ortega Pérez, Ruben; Barja, François

    2012-03-16

    Acetic acid bacteria (AAB) are Gram-negative, strictly aerobic microorganisms that show a unique resistance to ethanol (EtOH) and acetic acid (AcH). Members of the Acetobacter and Gluconacetobacter genera are capable of transforming EtOH into AcH via the alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH) enzymes and are used for the industrial production of vinegar. Several mechanisms have been proposed to explain how AAB resist high concentrations of AcH, such as the assimilation of acetate through the tricarboxylic acid (TCA) cycle, the export of acetate by various transporters and modifications of the outer membrane. However, except for a few acetate-specific proteins, little is known about the global proteome responses to AcH. In this study, we used 2D-DIGE to compare the proteome of Acetobacter pasteurianus LMG 1262(T) when growing in glucose or ethanol and in the presence of acetic acid. Interesting protein spots were selected using the ANOVA p-value of 0.05 as threshold and 1.5-fold as the minimal level of differential expression, and a total of 53 proteins were successfully identified. Additionally, the size of AAB was reduced by approximately 30% in length as a consequence of the acidity. A modification in the membrane polysaccharides was also revealed by PATAg specific staining. Copyright © 2011 Elsevier B.V. All rights reserved.

  2. Proteomics analysis of the endogenous, constitutive, leaf SUMOylome.

    Science.gov (United States)

    Colignon, Bertrand; Delaive, Edouard; Dieu, Marc; Demazy, Catherine; Muhovski, Yordan; Wallon, Cindy; Raes, Martine; Mauro, Sergio

    2017-01-06

    SUMOylation is a post-translational modification which regulates a number of critical biological processes in, for example mammals, yeast and plants. In order to fully understand the functional effects of SUMOylation an essential first step is the identification of endogenous targets for SUMOylation. Here we report the results of using a recently developed proteomic approach based on the use of 3D gels to identify the endogenous SUMO targets in leaves of Solanum tuberosum. By using 3D gels we avoid the problem of co-migration of proteins, which is a major limitation of 2D gels, and we enable the use of the highly sensitive CyDye DIGE fluor saturation dyes. Using this new method we have identified 39 individual proteins as probable SUMO targets in leaves of Solanum tuberosum. The advantages of this method compared with other approaches are discussed, and possible future developments are outlined. The authors have no conflicts of interest to declare. All authors have approved the manuscript and agree with submission to Journal of Proteomics. Copyright © 2016. Published by Elsevier B.V.

  3. Proteomic analysis of the flooding tolerance mechanism in mutant soybean.

    Science.gov (United States)

    Komatsu, Setsuko; Nanjo, Yohei; Nishimura, Minoru

    2013-02-21

    Flooding stress of soybean is a serious problem because it reduces growth; however, flooding-tolerant cultivars have not been identified. To analyze the flooding tolerance mechanism of soybean, the flooding-tolerant mutant was isolated and analyzed using a proteomic technique. Flooding-tolerance tests were repeated five times using gamma-ray irradiated soybeans, whose root growth (M6 stage) was not suppressed even under flooding stress. Two-day-old wild-type and mutant plants were subjected to flooding stress for 2days, and proteins were identified using a gel-based proteomic technique. In wild-type under flooding stress, levels of proteins related to development, protein synthesis/degradation, secondary metabolism, and the cell wall changed; however, these proteins did not markedly differ in the mutant. In contrast, an increased number of fermentation-related proteins were identified in the mutant under flooding stress. The root tips of mutant plants were not affected by flooding stress, even though the wild-type plants had damaged root. Alcohol dehydrogenase activity in the mutant increased at an early stage of flooding stress compared with that of the wild-type. Taken together, these results suggest that activation of the fermentation system in the early stages of flooding may be an important factor for the acquisition of flooding tolerance in soybean. Copyright © 2012 Elsevier B.V. All rights reserved.

  4. Comparative proteomics analysis of oral cancer cell lines: identification of cancer associated proteins

    Science.gov (United States)

    2014-01-01

    Background A limiting factor in performing proteomics analysis on cancerous cells is the difficulty in obtaining sufficient amounts of starting material. Cell lines can be used as a simplified model system for studying changes that accompany tumorigenesis. This study used two-dimensional gel electrophoresis (2DE) to compare the whole cell proteome of oral cancer cell lines vs normal cells in an attempt to identify cancer associated proteins. Results Three primary cell cultures of normal cells with a limited lifespan without hTERT immortalization have been successfully established. 2DE was used to compare the whole cell proteome of these cells with that of three oral cancer cell lines. Twenty four protein spots were found to have changed in abundance. MALDI TOF/TOF was then used to determine the identity of these proteins. Identified proteins were classified into seven functional categories – structural proteins, enzymes, regulatory proteins, chaperones and others. IPA core analysis predicted that 18 proteins were related to cancer with involvements in hyperplasia, metastasis, invasion, growth and tumorigenesis. The mRNA expressions of two proteins – 14-3-3 protein sigma and Stress-induced-phosphoprotein 1 – were found to correlate with the corresponding proteins’ abundance. Conclusions The outcome of this analysis demonstrated that a comparative study of whole cell proteome of cancer versus normal cell lines can be used to identify cancer associated proteins. PMID:24422745

  5. Whole-Proteome Analysis of Twelve Species of Alphaproteobacteria Links Four Pathogens

    Directory of Open Access Journals (Sweden)

    Yunyun Zhou

    2013-11-01

    Full Text Available Thousands of whole-genome and whole-proteome sequences have been made available through advances in sequencing technology, and sequences of millions more organisms will become available in the coming years. This wealth of genetic information will provide numerous opportunities to enhance our understanding of these organisms including a greater understanding of relationships among species. Researchers have used 16S rRNA and other gene sequences to study the evolutionary origins of bacteria, but these strategies do not provide insight into the sharing of genes among bacteria via horizontal transfer. In this work we use an open source software program called pClust to cluster proteins from the complete proteomes of twelve species of Alphaproteobacteria and generate a dendrogram from the resulting orthologous protein clusters. We compare the results with dendrograms constructed using the 16S rRNA gene and multiple sequence alignment of seven housekeeping genes. Analysis of the whole proteomes of these pathogens grouped Rickettsia typhi with three other animal pathogens whereas conventional sequence analysis failed to group these pathogens together. We conclude that whole-proteome analysis can give insight into relationships among species beyond their phylogeny, perhaps reflecting the effects of horizontal gene transfer and potentially providing insight into the functions of shared genes by means of shared phenotypes.

  6. Dynamic Proteomic Characteristics and Network Integration Revealing Key Proteins for Two Kernel Tissue Developments in Popcorn.

    Directory of Open Access Journals (Sweden)

    Yongbin Dong

    Full Text Available The formation and development of maize kernel is a complex dynamic physiological and biochemical process that involves the temporal and spatial expression of many proteins and the regulation of metabolic pathways. In this study, the protein profiles of the endosperm and pericarp at three important developmental stages were analyzed by isobaric tags for relative and absolute quantification (iTRAQ labeling coupled with LC-MS/MS in popcorn inbred N04. Comparative quantitative proteomic analyses among developmental stages and between tissues were performed, and the protein networks were integrated. A total of 6,876 proteins were identified, of which 1,396 were nonredundant. Specific proteins and different expression patterns were observed across developmental stages and tissues. The functional annotation of the identified proteins revealed the importance of metabolic and cellular processes, and binding and catalytic activities for the development of the tissues. The whole, endosperm-specific and pericarp-specific protein networks integrated 125, 9 and 77 proteins, respectively, which were involved in 54 KEGG pathways and reflected their complex metabolic interactions. Confirmation for the iTRAQ endosperm proteins by two-dimensional gel electrophoresis showed that 44.44% proteins were commonly found. However, the concordance between mRNA level and the protein abundance varied across different proteins, stages, tissues and inbred lines, according to the gene cloning and expression analyses of four relevant proteins with important functions and different expression levels. But the result by western blot showed their same expression tendency for the four proteins as by iTRAQ. These results could provide new insights into the developmental mechanisms of endosperm and pericarp, and grain formation in maize.

  7. Automated selected reaction monitoring data analysis workflow for large-scale targeted proteomic studies.

    Science.gov (United States)

    Surinova, Silvia; Hüttenhain, Ruth; Chang, Ching-Yun; Espona, Lucia; Vitek, Olga; Aebersold, Ruedi

    2013-08-01

    Targeted proteomics based on selected reaction monitoring (SRM) mass spectrometry is commonly used for accurate and reproducible quantification of protein analytes in complex biological mixtures. Strictly hypothesis-driven, SRM assays quantify each targeted protein by collecting measurements on its peptide fragment ions, called transitions. To achieve sensitive and accurate quantitative results, experimental design and data analysis must consistently account for the variability of the quantified transitions. This consistency is especially important in large experiments, which increasingly require profiling up to hundreds of proteins over hundreds of samples. Here we describe a robust and automated workflow for the analysis of large quantitative SRM data sets that integrates data processing, statistical protein identification and quantification, and dissemination of the results. The integrated workflow combines three software tools: mProphet for peptide identification via probabilistic scoring; SRMstats for protein significance analysis with linear mixed-effect models; and PASSEL, a public repository for storage, retrieval and query of SRM data. The input requirements for the protocol are files with SRM traces in mzXML format, and a file with a list of transitions in a text tab-separated format. The protocol is especially suited for data with heavy isotope-labeled peptide internal standards. We demonstrate the protocol on a clinical data set in which the abundances of 35 biomarker candidates were profiled in 83 blood plasma samples of subjects with ovarian cancer or benign ovarian tumors. The time frame to realize the protocol is 1-2 weeks, depending on the number of replicates used in the experiment.

  8. HUPO BPP pilot study: a proteomics analysis of the mouse brain of different developmental stages.

    Science.gov (United States)

    Wang, Jing; Gu, Yong; Wang, Lihong; Hang, Xingyi; Gao, Yan; Wang, Hangyan; Zhang, Chenggang

    2007-11-01

    This study is a part of the HUPO Brain Proteome Project (BPP) pilot study, which aims at obtaining a reliable database of mouse brain proteome, at the comparison of techniques, laboratories, and approaches as well as at preparing subsequent proteome studies of neurologic diseases. The C57/Bl6 mouse brains of three developmental stages at embryonic day 16 (E16), postnatal day 7 (P7), and 8 wk (P56) (n = 5 in each group) were provided by the HUPO BPP executive committee. The whole brain proteins of each animal were individually prepared using 2-DE coupled with PDQuest software analysis. The protein spots representing developmentally related or stably expressed proteins were then prepared with in-gel digestion followed with MALDI-TOF/TOF MS/MS and analyzed using the MASCOT search engines to search the Swiss-Prot or NCBInr database. The 2-DE gel maps of the mouse brains of all of the developmental stages were obtained and submitted to the Data Collection Centre (DCC). The proteins alpha-enolase, stathmin, actin, C14orf166 homolog, 28,000 kDa heat- and acid-stable phosphoprotein, 3-mercaptopyruvate sulfurtransferase and 40 S ribosomal protein S3a were successfully identified. A further Western blotting analysis demonstrated that enolase is a protein up-regulated in the mouse brain from embryonic stage to adult stage. These data are helpful for understanding the proteome changes in the development of the mouse brain.

  9. Comparative proteomic analysis reveals heart toxicity induced by chronic arsenic exposure in rats

    International Nuclear Information System (INIS)

    Huang, Qingyu; Xi, Guochen; Alamdar, Ambreen; Zhang, Jie; Shen, Heqing

    2017-01-01

    Arsenic is a widespread metalloid in the environment, which poses a broad spectrum of adverse effects on human health. However, a global view of arsenic-induced heart toxicity is still lacking, and the underlying molecular mechanisms remain unclear. By performing a comparative quantitative proteomic analysis, the present study aims to investigate the alterations of proteome profile in rat heart after long-term exposure to arsenic. As a result, we found that the abundance of 81 proteins were significantly altered by arsenic treatment (35 up-regulated and 46 down-regulated). Among these, 33 proteins were specifically associated with cardiovascular system development and function, including heart development, heart morphology, cardiac contraction and dilation, and other cardiovascular functions. It is further proposed that the aberrant regulation of 14 proteins induced by arsenic would disturb cardiac contraction and relaxation, impair heart morphogenesis and development, and induce thrombosis in rats, which is mediated by the Akt/p38 MAPK signaling pathway. Overall, these findings will augment our knowledge of the involved mechanisms and develop useful biomarkers for cardiotoxicity induced by environmental arsenic exposure. - Highlights: • Arsenic exposure has been associated with a number of adverse health effects. • The molecular mechanisms involved in arsenic-induced cardiotoxicity remain unclear. • Differential proteins were identified in arsenic-exposed rat heart by proteomics. • Arsenic induces heart toxicity through the Akt/p38 MAPK signaling pathway. - Label-free quantitative proteomic analysis of rat heart reveals putative mechanisms and biomarkers for arsenic-induced cardiotoxicity.

  10. Proteomic Analysis of the Cell Cycle of Procylic Form Trypanosoma brucei.

    Science.gov (United States)

    Crozier, Thomas W M; Tinti, Michele; Wheeler, Richard J; Ly, Tony; Ferguson, Michael A J; Lamond, Angus I

    2018-06-01

    We describe a single-step centrifugal elutriation method to produce synchronous Gap1 (G1)-phase procyclic trypanosomes at a scale amenable for proteomic analysis of the cell cycle. Using ten-plex tandem mass tag (TMT) labeling and mass spectrometry (MS)-based proteomics technology, the expression levels of 5325 proteins were quantified across the cell cycle in this parasite. Of these, 384 proteins were classified as cell-cycle regulated and subdivided into nine clusters with distinct temporal regulation. These groups included many known cell cycle regulators in trypanosomes, which validates the approach. In addition, we identify 40 novel cell cycle regulated proteins that are essential for trypanosome survival and thus represent potential future drug targets for the prevention of trypanosomiasis. Through cross-comparison to the TrypTag endogenous tagging microscopy database, we were able to validate the cell-cycle regulated patterns of expression for many of the proteins of unknown function detected in our proteomic analysis. A convenient interface to access and interrogate these data is also presented, providing a useful resource for the scientific community. Data are available via ProteomeXchange with identifier PXD008741 (https://www.ebi.ac.uk/pride/archive/). © 2018 by The American Society for Biochemistry and Molecular Biology, Inc.

  11. Review of software tools for design and analysis of large scale MRM proteomic datasets.

    Science.gov (United States)

    Colangelo, Christopher M; Chung, Lisa; Bruce, Can; Cheung, Kei-Hoi

    2013-06-15

    Selective or Multiple Reaction monitoring (SRM/MRM) is a liquid-chromatography (LC)/tandem-mass spectrometry (MS/MS) method that enables the quantitation of specific proteins in a sample by analyzing precursor ions and the fragment ions of their selected tryptic peptides. Instrumentation software has advanced to the point that thousands of transitions (pairs of primary and secondary m/z values) can be measured in a triple quadrupole instrument coupled to an LC, by a well-designed scheduling and selection of m/z windows. The design of a good MRM assay relies on the availability of peptide spectra from previous discovery-phase LC-MS/MS studies. The tedious aspect of manually developing and processing MRM assays involving thousands of transitions has spurred to development of software tools to automate this process. Software packages have been developed for project management, assay development, assay validation, data export, peak integration, quality assessment, and biostatistical analysis. No single tool provides a complete end-to-end solution, thus this article reviews the current state and discusses future directions of these software tools in order to enable researchers to combine these tools for a comprehensive targeted proteomics workflow. Copyright © 2013 The Authors. Published by Elsevier Inc. All rights reserved.

  12. Proteome Analysis of Disease Resistance against Ralstonia solanacearum in Potato Cultivar CT206-10

    Directory of Open Access Journals (Sweden)

    Sangryeol Park

    2016-02-01

    Full Text Available Potato is one of the most important crops worldwide. Its commercial cultivars are highly susceptible to many fungal and bacterial diseases. Among these, bacterial wilt caused by Ralstonia solanacearum causes significant yield loss. In the present study, integrated proteomics and genomics approaches were used in order to identify bacterial wilt resistant genes from Rs resistance potato cultivar CT-206-10. 2-DE and MALDI-TOF/TOF-MS analysis identified eight differentially abundant proteins including glycine-rich RNA binding protein (GRP, tomato stress induced-1 (TSI-1 protein, pathogenesis-related (STH-2 protein and pentatricopeptide repeat containing (PPR protein in response to Rs infection. Further, semi-quantitative RT-PCR identified up-regulation in transcript levels of all these genes upon Rs infection. Taken together, our results showed the involvement of the identified proteins in the Rs stress tolerance in potato. In the future, it would be interesting to raise the transgenic plants to further validate their involvement in resistance against Rs in potato.

  13. Extraction of intracellular protein from Glaciozyma antarctica for proteomics analysis

    Science.gov (United States)

    Faizura, S. Nor; Farahayu, K.; Faizal, A. B. Mohd; Asmahani, A. A. S.; Amir, R.; Nazalan, N.; Diba, A. B. Farah; Muhammad, M. Nor; Munir, A. M. Abdul

    2013-11-01

    Two preparation methods of crude extracts of psychrophilic yeast Glaciozyma antarctica were compared in order to obtain a good recovery of intracellular proteins. Extraction with mechanical procedures using sonication was found to be more effective for obtaining good yield compare to alkaline treatment method. The procedure is simple, rapid, and produce better yield. A total of 52 proteins were identified by combining both extraction methods. Most of the proteins identified in this study involves in the metabolic process including glycolysis pathway, pentose phosphate pathway, pyruyate decarboxylation and also urea cyle. Several chaperons were identified including probable cpr1-cyclophilin (peptidylprolyl isomerase), macrolide-binding protein fkbp12 and heat shock proteins which were postulate to accelerate proper protein folding. Characteristic of the fundamental cellular processes inferred from the expressed-proteome highlight the evolutionary and functional complexity existing in this domain of life.

  14. Biomechanical and proteomic analysis of INF- {beta}-treated astrocytes

    Energy Technology Data Exchange (ETDEWEB)

    Vergara, Daniele; Leporatti, Stefano; Maruccio, Giuseppe; Cingolani, Roberto; Rinaldi, Ross [National Nanotechnology Laboratory of CNR-INFM, ISUFI, University of Lecce, Italian Institute of Technology (IIT) Research Unit, via Arnesano, I-73100 Lecce (Italy); Martignago, Roberta; Nuccio, Franco De; Nicolardi, Giuseppe; Maffia, Michele [Department of Biological and Environmental Sciences and Technologies, University of Salento, via Monteroni, I-73100 Lecce (Italy); Bonsegna, Stefania; Santino, Angelo, E-mail: michele.maffia@unile.i, E-mail: ross.rinaldi@unile.i [Institute of Sciences of Food Production CNR, Unit of Lecce I-73100 (Italy)

    2009-11-11

    Astrocytes have a key role in the pathogenesis of several diseases including multiple sclerosis and were proposed as the designed target for immunotherapy. In this study we used atomic force microscopy (AFM) and proteomics methods to analyse and correlate the modifications induced in the viscoleastic properties of astrocytes to the changes induced in protein expression after interferon- {beta} (IFN-{beta}) treatment. Our results indicated that IFN-{beta} treatment resulted in a significant decrease in the Young's modulus, a measure of cell elasticity, in comparison with control cells. The molecular mechanisms that trigger these changes were investigated by 2DE (two-dimensional electrophoresis) and confocal analyses and confirmed by western blotting. Altered proteins were found to be involved in cytoskeleton organization and other important physiological processes.

  15. Biomechanical and proteomic analysis of INF- β-treated astrocytes

    International Nuclear Information System (INIS)

    Vergara, Daniele; Leporatti, Stefano; Maruccio, Giuseppe; Cingolani, Roberto; Rinaldi, Ross; Martignago, Roberta; Nuccio, Franco De; Nicolardi, Giuseppe; Maffia, Michele; Bonsegna, Stefania; Santino, Angelo

    2009-01-01

    Astrocytes have a key role in the pathogenesis of several diseases including multiple sclerosis and were proposed as the designed target for immunotherapy. In this study we used atomic force microscopy (AFM) and proteomics methods to analyse and correlate the modifications induced in the viscoleastic properties of astrocytes to the changes induced in protein expression after interferon- β (IFN-β) treatment. Our results indicated that IFN-β treatment resulted in a significant decrease in the Young's modulus, a measure of cell elasticity, in comparison with control cells. The molecular mechanisms that trigger these changes were investigated by 2DE (two-dimensional electrophoresis) and confocal analyses and confirmed by western blotting. Altered proteins were found to be involved in cytoskeleton organization and other important physiological processes.

  16. Proteomic analysis of the cyst stage of Entamoeba histolytica.

    Directory of Open Access Journals (Sweden)

    Ibne Karim M Ali

    Full Text Available The category B agent of bioterrorism, Entamoeba histolytica has a two-stage life cycle: an infective cyst stage, and an invasive trophozoite stage. Due to our inability to effectively induce encystation in vitro, our knowledge about the cyst form remains limited. This also hampers our ability to develop cyst-specific diagnostic tools.Three main aims were (i to identify E. histolytica proteins in cyst samples, (ii to enrich our knowledge about the cyst stage, and (iii to identify candidate proteins to develop cyst-specific diagnostic tools.Cysts were purified from the stool of infected individuals using Percoll (gradient purification. A highly sensitive LC-MS/MS mass spectrometer (Orbitrap was used to identify cyst proteins.A total of 417 non-redundant E. histolytica proteins were identified including 195 proteins that were never detected in trophozoite-derived proteomes or expressed sequence tag (EST datasets, consistent with cyst specificity. Cyst-wall specific glycoproteins Jacob, Jessie and chitinase were positively identified. Antibodies produced against Jacob identified cysts in fecal specimens and have potential utility as a diagnostic reagent. Several protein kinases, small GTPase signaling molecules, DNA repair proteins, epigenetic regulators, and surface associated proteins were also identified. Proteins we identified are likely to be among the most abundant in excreted cysts, and therefore show promise as diagnostic targets.The proteome data generated here are a first for naturally-occurring E. histolytica cysts, and they provide important insights into the infectious cyst form. Additionally, numerous unique candidate proteins were identified which will aid the development of new diagnostic tools for identification of E. histolytica cysts.

  17. Integrative analyses of miRNA and proteomics identify potential biological pathways associated with onset of pulmonary fibrosis in the bleomycin rat model

    International Nuclear Information System (INIS)

    Fukunaga, Satoki; Kakehashi, Anna; Sumida, Kayo; Kushida, Masahiko; Asano, Hiroyuki; Gi, Min; Wanibuchi, Hideki

    2015-01-01

    To determine miRNAs and their predicted target proteins regulatory networks which are potentially involved in onset of pulmonary fibrosis in the bleomycin rat model, we conducted integrative miRNA microarray and iTRAQ-coupled LC-MS/MS proteomic analyses, and evaluated the significance of altered biological functions and pathways. We observed that alterations of miRNAs and proteins are associated with the early phase of bleomycin-induced pulmonary fibrosis, and identified potential target pairs by using ingenuity pathway analysis. Using the data set of these alterations, it was demonstrated that those miRNAs, in association with their predicted target proteins, are potentially involved in canonical pathways reflective of initial epithelial injury and fibrogenic processes, and biofunctions related to induction of cellular development, movement, growth, and proliferation. Prediction of activated functions suggested that lung cells acquire proliferative, migratory, and invasive capabilities, and resistance to cell death especially in the very early phase of bleomycin-induced pulmonary fibrosis. The present study will provide new insights for understanding the molecular pathogenesis of idiopathic pulmonary fibrosis. - Highlights: • We analyzed bleomycin-induced pulmonary fibrosis in the rat. • Integrative analyses of miRNA microarray and proteomics were conducted. • We determined the alterations of miRNAs and their potential target proteins. • The alterations may control biological functions and pathways in pulmonary fibrosis. • Our result may provide new insights of pulmonary fibrosis

  18. Integrative analyses of miRNA and proteomics identify potential biological pathways associated with onset of pulmonary fibrosis in the bleomycin rat model

    Energy Technology Data Exchange (ETDEWEB)

    Fukunaga, Satoki [Department of Molecular Pathology, Osaka City University Graduate School of Medicine, 1-4-3 Asahi-machi, Abeno-ku, Osaka 545-8585 (Japan); Environmental Health Science Laboratory, Sumitomo Chemical Co., Ltd., 3-1-98 Kasugade-Naka, Konohana-ku, Osaka 554-8558 (Japan); Kakehashi, Anna [Department of Molecular Pathology, Osaka City University Graduate School of Medicine, 1-4-3 Asahi-machi, Abeno-ku, Osaka 545-8585 (Japan); Sumida, Kayo; Kushida, Masahiko; Asano, Hiroyuki [Environmental Health Science Laboratory, Sumitomo Chemical Co., Ltd., 3-1-98 Kasugade-Naka, Konohana-ku, Osaka 554-8558 (Japan); Gi, Min [Department of Molecular Pathology, Osaka City University Graduate School of Medicine, 1-4-3 Asahi-machi, Abeno-ku, Osaka 545-8585 (Japan); Wanibuchi, Hideki, E-mail: wani@med.osaka-cu.ac.jp [Department of Molecular Pathology, Osaka City University Graduate School of Medicine, 1-4-3 Asahi-machi, Abeno-ku, Osaka 545-8585 (Japan)

    2015-08-01

    To determine miRNAs and their predicted target proteins regulatory networks which are potentially involved in onset of pulmonary fibrosis in the bleomycin rat model, we conducted integrative miRNA microarray and iTRAQ-coupled LC-MS/MS proteomic analyses, and evaluated the significance of altered biological functions and pathways. We observed that alterations of miRNAs and proteins are associated with the early phase of bleomycin-induced pulmonary fibrosis, and identified potential target pairs by using ingenuity pathway analysis. Using the data set of these alterations, it was demonstrated that those miRNAs, in association with their predicted target proteins, are potentially involved in canonical pathways reflective of initial epithelial injury and fibrogenic processes, and biofunctions related to induction of cellular development, movement, growth, and proliferation. Prediction of activated functions suggested that lung cells acquire proliferative, migratory, and invasive capabilities, and resistance to cell death especially in the very early phase of bleomycin-induced pulmonary fibrosis. The present study will provide new insights for understanding the molecular pathogenesis of idiopathic pulmonary fibrosis. - Highlights: • We analyzed bleomycin-induced pulmonary fibrosis in the rat. • Integrative analyses of miRNA microarray and proteomics were conducted. • We determined the alterations of miRNAs and their potential target proteins. • The alterations may control biological functions and pathways in pulmonary fibrosis. • Our result may provide new insights of pulmonary fibrosis.

  19. GProX, a user-friendly platform for bioinformatics analysis and visualization of quantitative proteomics data.

    Science.gov (United States)

    Rigbolt, Kristoffer T G; Vanselow, Jens T; Blagoev, Blagoy

    2011-08-01

    Recent technological advances have made it possible to identify and quantify thousands of proteins in a single proteomics experiment. As a result of these developments, the analysis of data has become the bottleneck of proteomics experiment. To provide the proteomics community with a user-friendly platform for comprehensive analysis, inspection and visualization of quantitative proteomics data we developed the Graphical Proteomics Data Explorer (GProX)(1). The program requires no special bioinformatics training, as all functions of GProX are accessible within its graphical user-friendly interface which will be intuitive to most users. Basic features facilitate the uncomplicated management and organization of large data sets and complex experimental setups as well as the inspection and graphical plotting of quantitative data. These are complemented by readily available high-level analysis options such as database querying, clustering based on abundance ratios, feature enrichment tests for e.g. GO terms and pathway analysis tools. A number of plotting options for visualization of quantitative proteomics data is available and most analysis functions in GProX create customizable high quality graphical displays in both vector and bitmap formats. The generic import requirements allow data originating from essentially all mass spectrometry platforms, quantitation strategies and software to be analyzed in the program. GProX represents a powerful approach to proteomics data analysis providing proteomics experimenters with a toolbox for bioinformatics analysis of quantitative proteomics data. The program is released as open-source and can be freely downloaded from the project webpage at http://gprox.sourceforge.net.

  20. Integrative analysis to select cancer candidate biomarkers to targeted validation

    Science.gov (United States)

    Heberle, Henry; Domingues, Romênia R.; Granato, Daniela C.; Yokoo, Sami; Canevarolo, Rafael R.; Winck, Flavia V.; Ribeiro, Ana Carolina P.; Brandão, Thaís Bianca; Filgueiras, Paulo R.; Cruz, Karen S. P.; Barbuto, José Alexandre; Poppi, Ronei J.; Minghim, Rosane; Telles, Guilherme P.; Fonseca, Felipe Paiva; Fox, Jay W.; Santos-Silva, Alan R.; Coletta, Ricardo D.; Sherman, Nicholas E.; Paes Leme, Adriana F.

    2015-01-01

    Targeted proteomics has flourished as the method of choice for prospecting for and validating potential candidate biomarkers in many diseases. However, challenges still remain due to the lack of standardized routines that can prioritize a limited number of proteins to be further validated in human samples. To help researchers identify candidate biomarkers that best characterize their samples under study, a well-designed integrative analysis pipeline, comprising MS-based discovery, feature selection methods, clustering techniques, bioinformatic analyses and targeted approaches was performed using discovery-based proteomic data from the secretomes of three classes of human cell lines (carcinoma, melanoma and non-cancerous). Three feature selection algorithms, namely, Beta-binomial, Nearest Shrunken Centroids (NSC), and Support Vector Machine-Recursive Features Elimination (SVM-RFE), indicated a panel of 137 candidate biomarkers for carcinoma and 271 for melanoma, which were differentially abundant between the tumor classes. We further tested the strength of the pipeline in selecting candidate biomarkers by immunoblotting, human tissue microarrays, label-free targeted MS and functional experiments. In conclusion, the proposed integrative analysis was able to pre-qualify and prioritize candidate biomarkers from discovery-based proteomics to targeted MS. PMID:26540631

  1. Comparative proteomics analysis of placenta from pregnant women with intrahepatic cholestasis of pregnancy.

    Science.gov (United States)

    Zhang, Ting; Guo, Yueshuai; Guo, Xuejiang; Zhou, Tao; Chen, Daozhen; Xiang, Jingying; Zhou, Zuomin

    2013-01-01

    Intrahepatic cholestasis of pregnancy (ICP) usually occurs in the third trimester and associated with increased risks in fetal complications. Currently, the exact cause of this disease is unknown. In this study we aim to investigate the potential proteins in placenta, which may participate in the molecular mechanisms of ICP-related fetal complications using iTRAQ-based proteomics approach. The iTRAQ analysis combined with liquid chromatography-tandem mass spectrometry (LC-MS/MS) was performed to separate differentially expressed placental proteins from 4 pregnant women with ICP and 4 healthy pregnant women. Bioinformatics analysis was used to find the relative processes that these differentially expressed proteins were involved in. Three apoptosis related proteins ERp29, PRDX6 and MPO that resulted from iTRAQ-based proteomics were further verified in placenta by Western blotting and immunohistochemistry. Placental apoptosis was also detected by TUNEL assay. Proteomics results showed there were 38 differentially expressed proteins from pregnant women with ICP and healthy pregnant women, 29 were upregulated and 9 were downregulated in placenta from pregnant women with ICP. Bioinformatics analysis showed most of the identified proteins was functionally related to specific cell processes, including apoptosis, oxidative stress, lipid metabolism. The expression levels of ERp29, PRDX6 and MPO were consistent with the proteomics data. The apoptosis index in placenta from ICP patients was significantly increased. This preliminary work provides a better understanding of the proteomic alterations of placenta from pregnant women with ICP and may provide us some new insights into the pathophysiology and potential novel treatment targets for ICP.

  2. Metabolic effects of the iodothyronine functional analogue TRC150094 on the liver and skeletal muscle of high-fat diet fed overweight rats: an integrated proteomic study.

    Science.gov (United States)

    Silvestri, Elena; Glinni, Daniela; Cioffi, Federica; Moreno, Maria; Lombardi, Assunta; de Lange, Pieter; Senese, Rosalba; Ceccarelli, Michele; Salzano, Anna Maria; Scaloni, Andrea; Lanni, Antonia; Goglia, Fernando

    2012-07-06

    A novel functional iodothyronine analogue, TRC150094, which has a much lower potency toward thyroid hormone receptor (α1/β1) activation than triiodothyronine, has been shown to be effective at reducing adiposity in rats simultaneously receiving a high-fat diet (HFD). Here, by combining metabolic, functional and proteomic analysis, we studied how the hepatic and skeletal muscle phenotypes might respond to TRC150094 treatment in HFD-fed overweight rats. Drug treatment increased both the liver and skeletal muscle mitochondrial oxidative capacities without altering mitochondrial efficiency. Coherently, in terms of individual respiratory in-gel activity, blue-native analysis revealed an increased activity of complex V in the liver and of complexes II and V in tibialis muscle in TCR150094-treated animals. Subsequently, the identification of differentially expressed proteins and the analysis of their interrelations gave an integrated view of the phenotypic/metabolic adaptations occurring in the liver and muscle proteomes during drug treatment. TRC150094 significantly altered the expression of several proteins involved in key liver metabolic pathways, including amino acid and nitrogen metabolism, and fructose and mannose metabolism. The canonical pathways most strongly influenced by TRC150094 in tibialis muscle included glycolysis and gluconeogenesis, amino acid, fructose and mannose metabolism, and cell signaling. The phenotypic/metabolic influence of TRC150094 on the liver and skeletal muscle of HFD-fed overweight rats suggests the potential clinical application of this iodothyronine analogue in ameliorating metabolic risk parameters altered by diet regimens.

  3. MitProNet: A knowledgebase and analysis platform of proteome, interactome and diseases for mammalian mitochondria.

    Directory of Open Access Journals (Sweden)

    Jiabin Wang

    Full Text Available Mitochondrion plays a central role in diverse biological processes in most eukaryotes, and its dysfunctions are critically involved in a large number of diseases and the aging process. A systematic identification of mitochondrial proteomes and characterization of functional linkages among mitochondrial proteins are fundamental in understanding the mechanisms underlying biological functions and human diseases associated with mitochondria. Here we present a database MitProNet which provides a comprehensive knowledgebase for mitochondrial proteome, interactome and human diseases. First an inventory of mammalian mitochondrial proteins was compiled by widely collecting proteomic datasets, and the proteins were classified by machine learning to achieve a high-confidence list of mitochondrial proteins. The current version of MitProNet covers 1124 high-confidence proteins, and the remainders were further classified as middle- or low-confidence. An organelle-specific network of functional linkages among mitochondrial proteins was then generated by integrating genomic features encoded by a wide range of datasets including genomic context, gene expression profiles, protein-protein interactions, functional similarity and metabolic pathways. The functional-linkage network should be a valuable resource for the study of biological functions of mitochondrial proteins and human mitochondrial diseases. Furthermore, we utilized the network to predict candidate genes for mitochondrial diseases using prioritization algorithms. All proteins, functional linkages and disease candidate genes in MitProNet were annotated according to the information collected from their original sources including GO, GEO, OMIM, KEGG, MIPS, HPRD and so on. MitProNet features a user-friendly graphic visualization interface to present functional analysis of linkage networks. As an up-to-date database and analysis platform, MitProNet should be particularly helpful in comprehensive studies of

  4. Elevated host lipid metabolism revealed by iTRAQ-based quantitative proteomic analysis of cerebrospinal fluid of tuberculous meningitis patients.

    Science.gov (United States)

    Mu, Jun; Yang, Yongtao; Chen, Jin; Cheng, Ke; Li, Qi; Wei, Yongdong; Zhu, Dan; Shao, Weihua; Zheng, Peng; Xie, Peng

    2015-10-30

    Tuberculous meningitis (TBM) remains to be one of the most deadly infectious diseases. The pathogen interacts with the host immune system, the process of which is largely unknown. Various cellular processes of Mycobacterium tuberculosis (MTB) centers around lipid metabolism. To determine the lipid metabolism related proteins, a quantitative proteomic study was performed here to identify differential proteins in the cerebrospinal fluid (CSF) obtained from TBM patients (n = 12) and healthy controls (n = 12). CSF samples were desalted, concentrated, labelled with isobaric tags for relative and absolute quantitation (iTRAQ™), and analyzed by multi-dimensional liquid chromatography-tandem mass spectrometry (LC-MS/MS). Gene ontology and proteomic phenotyping analysis of the differential proteins were conducted using Database for Annotation, Visualization, and Integrated Discovery (DAVID) Bioinformatics Resources. ApoE and ApoB were selected for validation by ELISA. Proteomic phenotyping of the 4 differential proteins was invloved in the lipid metabolism. ELISA showed significantly increased ApoB levels in TBM subjects compared to healthy controls. Area under the receiver operating characteristic curve analysis demonstrated ApoB levels could distinguish TBM subjects from healthy controls and viral meningitis subjects with 89.3% sensitivity and 92% specificity. CSF lipid metabolism disregulation, especially elevated expression of ApoB, gives insights into the pathogenesis of TBM. Further evaluation of these findings in larger studies including anti-tuberculosis medicated and unmedicated patient cohorts with other center nervous system infectious diseases is required for successful clinical translation. Copyright © 2015 Elsevier Inc. All rights reserved.

  5. Application of survival analysis methodology to the quantitative analysis of LC-MS proteomics data.

    Science.gov (United States)

    Tekwe, Carmen D; Carroll, Raymond J; Dabney, Alan R

    2012-08-01

    Protein abundance in quantitative proteomics is often based on observed spectral features derived from liquid chromatography mass spectrometry (LC-MS) or LC-MS/MS experiments. Peak intensities are largely non-normal in distribution. Furthermore, LC-MS-based proteomics data frequently have large proportions of missing peak intensities due to censoring mechanisms on low-abundance spectral features. Recognizing that the observed peak intensities detected with the LC-MS method are all positive, skewed and often left-censored, we propose using survival methodology to carry out differential expression analysis of proteins. Various standard statistical techniques including non-parametric tests such as the Kolmogorov-Smirnov and Wilcoxon-Mann-Whitney rank sum tests, and the parametric survival model and accelerated failure time-model with log-normal, log-logistic and Weibull distributions were used to detect any differentially expressed proteins. The statistical operating characteristics of each method are explored using both real and simulated datasets. Survival methods generally have greater statistical power than standard differential expression methods when the proportion of missing protein level data is 5% or more. In particular, the AFT models we consider consistently achieve greater statistical power than standard testing procedures, with the discrepancy widening with increasing missingness in the proportions. The testing procedures discussed in this article can all be performed using readily available software such as R. The R codes are provided as supplemental materials. ctekwe@stat.tamu.edu.

  6. Application of survival analysis methodology to the quantitative analysis of LC-MS proteomics data

    KAUST Repository

    Tekwe, C. D.

    2012-05-24

    MOTIVATION: Protein abundance in quantitative proteomics is often based on observed spectral features derived from liquid chromatography mass spectrometry (LC-MS) or LC-MS/MS experiments. Peak intensities are largely non-normal in distribution. Furthermore, LC-MS-based proteomics data frequently have large proportions of missing peak intensities due to censoring mechanisms on low-abundance spectral features. Recognizing that the observed peak intensities detected with the LC-MS method are all positive, skewed and often left-censored, we propose using survival methodology to carry out differential expression analysis of proteins. Various standard statistical techniques including non-parametric tests such as the Kolmogorov-Smirnov and Wilcoxon-Mann-Whitney rank sum tests, and the parametric survival model and accelerated failure time-model with log-normal, log-logistic and Weibull distributions were used to detect any differentially expressed proteins. The statistical operating characteristics of each method are explored using both real and simulated datasets. RESULTS: Survival methods generally have greater statistical power than standard differential expression methods when the proportion of missing protein level data is 5% or more. In particular, the AFT models we consider consistently achieve greater statistical power than standard testing procedures, with the discrepancy widening with increasing missingness in the proportions. AVAILABILITY: The testing procedures discussed in this article can all be performed using readily available software such as R. The R codes are provided as supplemental materials. CONTACT: ctekwe@stat.tamu.edu.

  7. Proteomic-Biostatistic Integrated Approach for Finding the Underlying Molecular Determinants of Hypertension in Human Plasma.

    Science.gov (United States)

    Gajjala, Prathibha R; Jankowski, Vera; Heinze, Georg; Bilo, Grzegorz; Zanchetti, Alberto; Noels, Heidi; Liehn, Elisa; Perco, Paul; Schulz, Anna; Delles, Christian; Kork, Felix; Biessen, Erik; Narkiewicz, Krzysztof; Kawecka-Jaszcz, Kalina; Floege, Juergen; Soranna, Davide; Zidek, Walter; Jankowski, Joachim

    2017-08-01

    Despite advancements in lowering blood pressure, the best approach to lower it remains controversial because of the lack of information on the molecular basis of hypertension. We, therefore, performed plasma proteomics of plasma from patients with hypertension to identify molecular determinants detectable in these subjects but not in controls and vice versa. Plasma samples from hypertensive subjects (cases; n=118) and controls (n=85) from the InGenious HyperCare cohort were used for this study and performed mass spectrometric analysis. Using biostatistical methods, plasma peptides specific for hypertension were identified, and a model was developed using least absolute shrinkage and selection operator logistic regression. The underlying peptides were identified and sequenced off-line using matrix-assisted laser desorption ionization orbitrap mass spectrometry. By comparison of the molecular composition of the plasma samples, 27 molecular determinants were identified differently expressed in cases from controls. Seventy percent of the molecular determinants selected were found to occur less likely in hypertensive patients. In cross-validation, the overall R 2 was 0.434, and the area under the curve was 0.891 with 95% confidence interval 0.8482 to 0.9349, P hypertensive patients were found to be -2.007±0.3568 and 3.383±0.2643, respectively, P hypertensives and normotensives. The identified molecular determinants may be the starting point for further studies to clarify the molecular causes of hypertension. © 2017 American Heart Association, Inc.

  8. Integrated and comparative proteomics of high-oil and high-protein soybean seeds.

    Science.gov (United States)

    Xu, Xiu Ping; Liu, Hui; Tian, Lihong; Dong, Xiang Bai; Shen, Shi Hua; Qu, Le Qing

    2015-04-01

    We analysed the global protein expression in seeds of a high-oil soybean cultivar (Jiyu 73, JY73) by proteomics. More than 700 protein spots were detected and 363 protein spots were successfully identified. Comparison of the protein profile of JY73 with that of a high-protein cultivar (Zhonghuang 13, ZH13) revealed 40 differentially expressed proteins, including oil synthesis, redox/stress, hydrolysis and storage-related proteins. All redox/stress proteins were less or not expressed in JY73, whereas the expression of the major storage proteins, nitrogen and carbon metabolism-related proteins was higher in ZH13. Biochemical analysis of JY73 revealed that it was in a low oxidation state, with a high content of polyunsaturated fatty acids and vitamin E. Vitamin E was more active than antioxidant enzymes and protected the soybean seed in a lower oxidation state. The characteristics of high oil and high protein in soybean, we revealed, might provide a reference for soybean nutrition and soybean breeding. Copyright © 2014 Elsevier Ltd. All rights reserved.

  9. The effect of using an inappropriate protein database for proteomic data analysis.

    Directory of Open Access Journals (Sweden)

    Giselle M Knudsen

    Full Text Available A recent study by Bromenshenk et al., published in PLoS One (2010, used proteomic analysis to identify peptides purportedly of Iridovirus and Nosema origin; however the validity of this finding is controversial. We show here through re-analysis of a subset of this data that many of the spectra identified by Bromenshenk et al. as deriving from Iridovirus and Nosema proteins are actually products from Apis mellifera honey bee proteins. We find no reliable evidence that proteins from Iridovirus and Nosema are present in the samples that were re-analyzed. This article is also intended as a learning exercise for illustrating some of the potential pitfalls of analysis of mass spectrometry proteomic data and to encourage authors to observe MS/MS data reporting guidelines that would facilitate recognition of analysis problems during the review process.

  10. Scientific Workflow Management in Proteomics

    Science.gov (United States)

    de Bruin, Jeroen S.; Deelder, André M.; Palmblad, Magnus

    2012-01-01

    Data processing in proteomics can be a challenging endeavor, requiring extensive knowledge of many different software packages, all with different algorithms, data format requirements, and user interfaces. In this article we describe the integration of a number of existing programs and tools in Taverna Workbench, a scientific workflow manager currently being developed in the bioinformatics community. We demonstrate how a workflow manager provides a single, visually clear and intuitive interface to complex data analysis tasks in proteomics, from raw mass spectrometry data to protein identifications and beyond. PMID:22411703

  11. Integrated Translatome and Proteome: Approach for Accurate Portraying of Widespread Multifunctional Aspects of Trichoderma

    Science.gov (United States)

    Sharma, Vivek; Salwan, Richa; Sharma, P. N.; Gulati, Arvind

    2017-01-01

    Genome-wide studies of transcripts expression help in systematic monitoring of genes and allow targeting of candidate genes for future research. In contrast to relatively stable genomic data, the expression of genes is dynamic and regulated both at time and space level at different level in. The variation in the rate of translation is specific for each protein. Both the inherent nature of an mRNA molecule to be translated and the external environmental stimuli can affect the efficiency of the translation process. In biocontrol agents (BCAs), the molecular response at translational level may represents noise-like response of absolute transcript level and an adaptive response to physiological and pathological situations representing subset of mRNAs population actively translated in a cell. The molecular responses of biocontrol are complex and involve multistage regulation of number of genes. The use of high-throughput techniques has led to rapid increase in volume of transcriptomics data of Trichoderma. In general, almost half of the variations of transcriptome and protein level are due to translational control. Thus, studies are required to integrate raw information from different “omics” approaches for accurate depiction of translational response of BCAs in interaction with plants and plant pathogens. The studies on translational status of only active mRNAs bridging with proteome data will help in accurate characterization of only a subset of mRNAs actively engaged in translation. This review highlights the associated bottlenecks and use of state-of-the-art procedures in addressing the gap to accelerate future accomplishment of biocontrol mechanisms. PMID:28900417

  12. Proteomic analysis of early phase of conidia germination in Aspergillus nidulans.

    Science.gov (United States)

    Oh, Young Taek; Ahn, Chun-Seob; Kim, Jeong Geun; Ro, Hyeon-Su; Lee, Chang-Won; Kim, Jae Won

    2010-03-01

    In order to investigate proteins involved in early phase of conidia germination, proteomic analysis was performed using two-dimensional gel electrophoresis (2D-GE) in conjunction with MALDI-TOF mass spectrometry (MS). The expression levels of 241 proteins varied quantitatively with statistical significance (Pproteomic analysis of early phase of conidia germination and will contribute to a better understanding of the molecular events involved in conidia germination process. Copyright (c) 2009 Elsevier Inc. All rights reserved.

  13. 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

  14. Data in support of proteomic analysis of pneumococcal pediatric clinical isolates to construct a protein array

    Directory of Open Access Journals (Sweden)

    Alfonso Olaya-Abril

    2016-03-01

    Full Text Available Surface proteins play key roles in the interaction between cells and their environment, and in pathogenic microorganisms they are the best targets for drug or vaccine discovery and/or development. In addition, surface proteins can be the basis for serodiagnostic tools aiming at developing more affordable techniques for early diagnosis of infection in patients. We carried out a proteomic analysis of a collection of pediatric clinical isolates of Streptococcus pneumoniae, an important human pathogen responsible for more than 1.5 million child deaths worldwide. For that, cultured live bacterial cells were “shaved” with trypsin, and the recovered peptides were analyzed by LC/MS/MS. We selected 95 proteins to be produced as recombinant polypeptides, and printed them on an array. We probed the protein array with a collection of patient sera to define serodiagnostic antigens. The mass spectrometry proteomics data correspond to those published in [1] and have been deposited to the ProteomeXchange Consortium [2] via the PRIDE partner repository [3] with the dataset identifier http://www.ebi.ac.uk/pride/archive/projects/PXD001740. The protein array raw data are provided as supplemental material in this article. Keywords: Pneumococcus, Protein arrays, Proteomics, Diagnostics

  15. Detailed tail proteomic analysis of axolotl (Ambystoma mexicanum) using an mRNA-seq reference database.

    Science.gov (United States)

    Demircan, Turan; Keskin, Ilknur; Dumlu, Seda Nilgün; Aytürk, Nilüfer; Avşaroğlu, Mahmut Erhan; Akgün, Emel; Öztürk, Gürkan; Baykal, Ahmet Tarık

    2017-01-01

    Salamander axolotl has been emerging as an important model for stem cell research due to its powerful regenerative capacity. Several advantages, such as the high capability of advanced tissue, organ, and appendages regeneration, promote axolotl as an ideal model system to extend our current understanding on the mechanisms of regeneration. Acknowledging the common molecular pathways between amphibians and mammals, there is a great potential to translate the messages from axolotl research to mammalian studies. However, the utilization of axolotl is hindered due to the lack of reference databases of genomic, transcriptomic, and proteomic data. Here, we introduce the proteome analysis of the axolotl tail section searched against an mRNA-seq database. We translated axolotl mRNA sequences to protein sequences and annotated these to process the LC-MS/MS data and identified 1001 nonredundant proteins. Functional classification of identified proteins was performed by gene ontology searches. The presence of some of the identified proteins was validated by in situ antibody labeling. Furthermore, we have analyzed the proteome expressional changes postamputation at three time points to evaluate the underlying mechanisms of the regeneration process. Taken together, this work expands the proteomics data of axolotl to contribute to its establishment as a fully utilized model. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  16. 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.

  17. A proteomic analysis of the chromoplasts isolated from sweet orange fruits [Citrus sinensis (L.) Osbeck].

    Science.gov (United States)

    Zeng, Yunliu; Pan, Zhiyong; Ding, Yuduan; Zhu, Andan; Cao, Hongbo; Xu, Qiang; Deng, Xiuxin

    2011-11-01

    Here, a comprehensive proteomic analysis of the chromoplasts purified from sweet orange using Nycodenz density gradient centrifugation is reported. A GeLC-MS/MS shotgun approach was used to identify the proteins of pooled chromoplast samples. A total of 493 proteins were identified from purified chromoplasts, of which 418 are putative plastid proteins based on in silico sequence homology and functional analyses. Based on the predicted functions of these identified plastid proteins, a large proportion (∼60%) of the chromoplast proteome of sweet orange is constituted by proteins involved in carbohydrate metabolism, amino acid/protein synthesis, and secondary metabolism. Of note, HDS (hydroxymethylbutenyl 4-diphosphate synthase), PAP (plastid-lipid-associated protein), and psHSPs (plastid small heat shock proteins) involved in the synthesis or storage of carotenoid and stress response are among the most abundant proteins identified. A comparison of chromoplast proteomes between sweet orange and tomato suggested a high level of conservation in a broad range of metabolic pathways. However, the citrus chromoplast was characterized by more extensive carotenoid synthesis, extensive amino acid synthesis without nitrogen assimilation, and evidence for lipid metabolism concerning jasmonic acid synthesis. In conclusion, this study provides an insight into the major metabolic pathways as well as some unique characteristics of the sweet orange chromoplasts at the whole proteome level.

  18. A Routine 'Top-Down' Approach to Analysis of the Human Serum Proteome.

    Science.gov (United States)

    D'Silva, Arlene M; Hyett, Jon A; Coorssen, Jens R

    2017-06-06

    Serum provides a rich source of potential biomarker proteoforms. One of the major obstacles in analysing serum proteomes is detecting lower abundance proteins owing to the presence of hyper-abundant species (e.g., serum albumin and immunoglobulins). Although depletion methods have been used to address this, these can lead to the concomitant removal of non-targeted protein species, and thus raise issues of specificity, reproducibility, and the capacity for meaningful quantitative analyses. Altering the native stoichiometry of the proteome components may thus yield a more complex series of issues than dealing directly with the inherent complexity of the sample. Hence, here we targeted method refinements so as to ensure optimum resolution of serum proteomes via a top down two-dimensional gel electrophoresis (2DE) approach that enables the routine assessment of proteoforms and is fully compatible with subsequent mass spectrometric analyses. Testing included various fractionation and non-fractionation approaches. The data show that resolving 500 µg protein on 17 cm 3-10 non-linear immobilised pH gradient strips in the first dimension followed by second dimension resolution on 7-20% gradient gels with a combination of lithium dodecyl sulfate (LDS) and sodium dodecyl sulfate (SDS) detergents markedly improves the resolution and detection of proteoforms in serum. In addition, well established third dimension electrophoretic separations in combination with deep imaging further contributed to the best available resolution, detection, and thus quantitative top-down analysis of serum proteomes.

  19. Application of meta-transcriptomics and –proteomics to analysis of in situ physiological state

    Directory of Open Access Journals (Sweden)

    Allan eKonopka

    2012-05-01

    Full Text Available Analysis of the growth-limiting factor or environmental stressors affecting microbes in situ is of fundamental importance but analytically difficult. Microbes can reduce in situ limiting nutrient concentrations to sub-micromolar levels, and contaminated ecosystems may contain multiple stressors. The patterns of gene or protein expression by microbes in nature can be used to infer growth limitations, because they are regulated in response to environmental conditions. Experimental studies under controlled conditions in the laboratory provide the physiological underpinnings for developing these physiological indicators. Although regulatory networks may differ among specific microbes, there are some broad principles that can be applied, related to limiting nutrient acquisition, resource allocation, and stress responses. As technologies for transcriptomics and proteomics mature, the capacity to apply these approaches to complex microbial communities will accelerate. In particular, global proteomics reflect expressed catalytic activities. Furthermore, the high mass accuracy of some proteomic approaches allows mapping back to specific microbial strains. For example, at the Rifle IFRC field site in Western Colorado, the physiological status of Fe(III-reducing populations has been tracked over time. Members of a subsurface clade within the Geobacter predominated during carbon amendment to the subsurface environment. At the functional level, proteomic identifications produced inferences regarding (i temporal changes in anabolism and catabolism of acetate, (ii the onset of N2 fixation when N became limiting, and (iii expression of phosphate transporters during periods of intense growth. The application of these approaches in situ can lead to discovery of novel physiological adaptations.

  20. Quantitative Proteomic Analysis of Mouse Embryonic Fibroblasts and Induced Pluripotent Stem Cells Using 16O /18O labeling

    Energy Technology Data Exchange (ETDEWEB)

    Huang, Xin; Tian, Changhai; Liu, Miao; Wang, Yongxiang; Tolmachev, Aleksey V.; Sharma, Seema; Yu, Fang; Fu, Kai; Zheng, Jialin; Ding, Shi-Jian

    2012-04-06

    Induced pluripotent stem cells (iPSC) hold great promise for regenerative medicine as well as for investigations into the pathogenesis and treatment of various diseases. Understanding of key intracellular signaling pathways and protein targets that control development of iPSC from somatic cells is essential for designing new approaches to improve reprogramming efficiency. Here we report the development and application of an integrated quantitative proteomics platform for investigating differences in protein expressions between mouse embryonic fibroblasts (MEF) and MEF-derived iPSC. This platform consists of 16O/18O labeling, multidimensional peptide separation coupled with tandem mass spectrometry, and data analysis with UNiquant software. Using this platform a total of 2,481 proteins were identified and quantified from the 16O/18O-labeled MEF-iPSC proteome mixtures with a false discovery rate of 0.01. Among them, 218 proteins were significantly upregulated, while 247 proteins were significantly downregulated in iPSC compared to MEF. Many nuclear proteins, including Hdac1, Dnmt1, Pcna, Ccnd1, Smarcc1, and subunits in DNA replication and RNA polymerase II complex were found to be enhanced in iPSC. Protein network analysis revealed that Pcna functions as a hub orchestrating complicated mechanisms including DNA replication, epigenetic inheritance (Dnmt1) and chromatin remodeling (Smarcc1) to reprogram MEF and maintain stemness of iPSC.

  1. Multiplex Brain Proteomic Analysis Revealed the Molecular Therapeutic Effects of Buyang Huanwu Decoction on Cerebral Ischemic Stroke Mice.

    Directory of Open Access Journals (Sweden)

    Hong-Jhang Chen

    Full Text Available Stroke is the second-leading cause of death worldwide, and tissue plasminogen activator (TPA is the only drug used for a limited group of stroke patients in the acute phase. Buyang Huanwu Decoction (BHD, a traditional Chinese medicine prescription, has long been used for improving neurological functional recovery in stroke. In this study, we characterized the therapeutic effect of TPA and BHD in a cerebral ischemia/reperfusion (CIR injury mouse model using multiplex proteomics approach. After the iTRAQ-based proteomics analysis, 1310 proteins were identified from the mouse brain with <1% false discovery rate. Among them, 877 quantitative proteins, 10.26% (90/877, 1.71% (15/877, and 2.62% (23/877 of the proteins was significantly changed in the CIR, BHD treatment, and TPA treatment, respectively. Functional categorization analysis showed that BHD treatment preserved the integrity of the blood-brain barrier (BBB (Alb, Fga, and Trf, suppressed excitotoxicity (Grm5, Gnai, and Gdi, and enhanced energy metabolism (Bdh, thereby revealing its multiple effects on ischemic stroke mice. Moreover, the neurogenesis marker doublecortin was upregulated, and the activity of glycogen synthase kinase 3 (GSK-3 and Tau was inhibited, which represented the neuroprotective effects. However, TPA treatment deteriorated BBB breakdown. This study highlights the potential of BHD in clinical applications for ischemic stroke.

  2. Analysis of membrane proteome and secretome in cells over-expressing ADAM17 using quantitative proteomics

    International Nuclear Information System (INIS)

    Kawahara, R.; Simabuco, F.M.; Yokoo, S.; Paes Leme, A.F.; Sherman, N.

    2012-01-01

    Full text: A disintegrin and metalloproteinase (ADAM) protease is involved in proteolytic ectodomain shedding of several membrane-associated proteins and modulation of key cell signaling pathways in the tumor microenvironment. In this study, we examined the effect of over-expressing the full length human ADAM17 in membrane and secreted proteins. To this end, we constructed a stable Flp-In T-RExHEK293 cells expressing ADAM17 by tetracycline induction. These cells were grown in Dulbeccos modified Eagles medium containing light lysine, arginine or heavy, L-Arg-13C615N4 and L-Lys -13C615N2 (SILAC: stable isotope labeling with amino acid in cell culture) media and they were treated with an ADAM17 activator, phorbolester (PMA). Controls such as Flp-In T-RExHEK293 cell without PMA treatment and without ADAM17 cloned were cultivated in light medium. The ADAM17 overexpression was induced with tetracycline 500 ng/ml for 24 hours. Cells in a heavy condition were treated with PMA 50 ng/ml for 1 hour and vehicle DMSO was used as control in a light cell condition. The extracellular media were collected, concentrated and used to evaluate the secretome and a cell surface biotinylation-based approach was used to capture cell surface-associated proteins. The biotinylated proteins were eluted with dithiothreitol, alkylated with iodoacetamide and then digested with trypsin. The resulting peptides were subjected to LC-MS/MS analysis on an ETD enabled Orbitrap Velos instrument. The results showed different proteins up or down regulated in membrane and secretome analysis which might represent potential molecules involved in signaling or ADAM17 regulation events. (author)

  3. Analysis of Pacific oyster larval proteome and its response to high-CO2

    KAUST Repository

    Dineshram, R.

    2012-10-01

    Most calcifying organisms show depressed metabolic, growth and calcification rates as symptoms to high-CO2 due to ocean acidification (OA) process. Analysis of the global expression pattern of proteins (proteome analysis) represents a powerful tool to examine these physiological symptoms at molecular level, but its applications are inadequate. To address this knowledge gap, 2-DE coupled with mass spectrophotometer was used to compare the global protein expression pattern of oyster larvae exposed to ambient and to high-CO2. Exposure to OA resulted in marked reduction of global protein expression with a decrease or loss of 71 proteins (18% of the expressed proteins in control), indicating a wide-spread depression of metabolic genes expression in larvae reared under OA. This is, to our knowledge, the first proteome analysis that provides insights into the link between physiological suppression and protein down-regulation under OA in oyster larvae. © 2012 Elsevier Ltd.

  4. Integrative biological analysis for neuropsychopharmacology.

    Science.gov (United States)

    Emmett, Mark R; Kroes, Roger A; Moskal, Joseph R; Conrad, Charles A; Priebe, Waldemar; Laezza, Fernanda; Meyer-Baese, Anke; Nilsson, Carol L

    2014-01-01

    Although advances in psychotherapy have been made in recent years, drug discovery for brain diseases such as schizophrenia and mood disorders has stagnated. The need for new biomarkers and validated therapeutic targets in the field of neuropsychopharmacology is widely unmet. The brain is the most complex part of human anatomy from the standpoint of number and types of cells, their interconnections, and circuitry. To better meet patient needs, improved methods to approach brain studies by understanding functional networks that interact with the genome are being developed. The integrated biological approaches--proteomics, transcriptomics, metabolomics, and glycomics--have a strong record in several areas of biomedicine, including neurochemistry and neuro-oncology. Published applications of an integrated approach to projects of neurological, psychiatric, and pharmacological natures are still few but show promise to provide deep biological knowledge derived from cells, animal models, and clinical materials. Future studies that yield insights based on integrated analyses promise to deliver new therapeutic targets and biomarkers for personalized medicine.

  5. Spatial and molecular resolution of diffuse malignant mesothelioma heterogeneity by integrating label-free FTIR imaging, laser capture microdissection and proteomics

    Science.gov (United States)

    Großerueschkamp, Frederik; Bracht, Thilo; Diehl, Hanna C.; Kuepper, Claus; Ahrens, Maike; Kallenbach-Thieltges, Angela; Mosig, Axel; Eisenacher, Martin; Marcus, Katrin; Behrens, Thomas; Brüning, Thomas; Theegarten, Dirk; Sitek, Barbara; Gerwert, Klaus

    2017-03-01

    Diffuse malignant mesothelioma (DMM) is a heterogeneous malignant neoplasia manifesting with three subtypes: epithelioid, sarcomatoid and biphasic. DMM exhibit a high degree of spatial heterogeneity that complicates a thorough understanding of the underlying different molecular processes in each subtype. We present a novel approach to spatially resolve the heterogeneity of a tumour in a label-free manner by integrating FTIR imaging and laser capture microdissection (LCM). Subsequent proteome analysis of the dissected homogenous samples provides in addition molecular resolution. FTIR imaging resolves tumour subtypes within tissue thin-sections in an automated and label-free manner with accuracy of about 85% for DMM subtypes. Even in highly heterogeneous tissue structures, our label-free approach can identify small regions of interest, which can be dissected as homogeneous samples using LCM. Subsequent proteome analysis provides a location specific molecular characterization. Applied to DMM subtypes, we identify 142 differentially expressed proteins, including five protein biomarkers commonly used in DMM immunohistochemistry panels. Thus, FTIR imaging resolves not only morphological alteration within tissue but it resolves even alterations at the level of single proteins in tumour subtypes. Our fully automated workflow FTIR-guided LCM opens new avenues collecting homogeneous samples for precise and predictive biomarkers from omics studies.

  6. An integrated study of natural hydroxyapatite-induced osteogenic differentiation of mesenchymal stem cells using transcriptomics, proteomics and microRNA analyses

    International Nuclear Information System (INIS)

    Zhang, Zhiwei; Wang, Jiandan; Lü, Xiaoying

    2014-01-01

    This work combined transcriptomics, proteomics, and microRNA (miRNA) analyses to elucidate the mechanism of natural hydroxyapatite (NHA)-induced osteogenic differentiation of mesenchymal stem cells (MSCs). First, NHA powder was obtained from pig bones and fabricated into disc-shaped samples. Subsequently, the proliferation and osteogenic differentiation of MSCs cultured on NHA were investigated. Then, proteomics was employed to detect the protein expression profiles of MSCs cultured on NHA, and the effect of NHA on MSCs was analyzed through an integrated pathway analysis (including proteomics and previous transcriptomics data) in which specific NHA-induced differentiation pathways were analyzed. The pathway nodes with expression data at both the mRNA and protein levels (mRNA–protein pairs) were filtered in differentiation-related pathways. miRNAs corresponding to these target mRNA–protein pairs were predicted, screened and tested, and the regulatory effects of miRNAs on mRNA–protein pairs were analyzed. Finally, the NHA-induced osteogenic pathways were verified. The results of an MTT assay and alkaline phosphatase (ALP) staining showed that the cell proliferation rate decreased and the osteogenic performance improved in the presence of NHA. By integrating transcriptomics and proteomics, the genes and proteins involved in 89 pathways were shown to be differentially expressed. Among them, 5 differentiation-associated pathways, in which 9 miRNAs and 8 regulated-target mRNA–protein zby inhibiting the target mRNA–protein pair HSPA8 in the MAPK signaling pathway, and miR-26a and miR-26b might inhibit adipogenic differentiation by repressing the target mRNA–protein pair HMGA1 in the adipogenesis pathway. A verification experiment for the osteogenic pathway indicated that the ERK1/2 or JNK MAPK pathways might play an important role in NHA-induced osteogenic differentiation. In conclusion, NHA affected MSCs at both the transcriptional and translational levels

  7. Proteomic analysis of cell walls of two developmental stages of alfalfa stems

    Directory of Open Access Journals (Sweden)

    Julian C Verdonk

    2012-12-01

    Full Text Available Cell walls are important for the growth and development of all plants. They are also valuable resources for feed and fiber, and more recently as a potential feedstock for bioenergy production. Cell wall proteins comprise only a fraction of the cell wall, but play important roles in establishing the walls and in the chemical interactions (e.g. crosslinking of cell wall components. This crosslinking provides structure, but restricts digestibility of cell wall complex carbohydrates, limiting available energy in animal and bioenergy production systems. Manipulation of cell wall proteins could be a strategy to improve digestibility. An analysis of the cell wall proteome of apical alfalfa stems (less mature, more digestible and basal alfalfa stems (more mature, less digestible was conducted using a recently developed low-salt/density gradient method for the isolation of cell walls. Walls were subsequently subjected to a modified extraction utilizing EGTA to remove pectins, followed by a LiCl extraction to isolate more tightly bound proteins. Recovered proteins were identified using shotgun proteomics. We identified 272 proteins in the alfalfa stem cell wall proteome, 153 of which had not previously been identified in cell wall proteomic analyses. Nearly 70% percent of the identified proteins were predicted to be secreted, as would be expected for most cell wall proteins, an improvement over previously published studies using traditional cell wall isolation methods. A comparison of our and several other cell wall proteomic studies indicates little overlap in identified proteins among them, which may be largely due to differences in the tissues used as well as differences in experimental approach.

  8. Proteomic analysis of chinook salmon (Oncorhynchus tshawytscha ovarian fluid.

    Directory of Open Access Journals (Sweden)

    Sheri L Johnson

    Full Text Available The ovarian, or coelomic, fluid that is released with the egg mass of many fishes is increasingly found to play an important role in several biological processes crucial for reproductive success. These include maintenance of oocyte fertility and developmental competence, prolonging of sperm motility, and enhancing sperm swimming speed. Here we examined if and how the proteome of chinook salmon (Oncorhynchus tshawytscha ovarian fluid varied among females and then sought to examine the composition of this fluid. Ovarian fluid in chinook salmon was analyzed using 1D SDS PAGE and LC-MS/MS tryptic digest screened against Mascot and Sequest databases. We found marked differences in the number and concentrations of proteins in salmon ovarian fluid across different females. A total of 174 proteins were identified in ovarian fluid, 47 of which were represented by six or more peptides, belonging to one of six Gene Ontology pathways. The response to chemical stimulus and response to hypoxia pathways were best represented, accounting for 26 of the 174 proteins. The current data set provides a resource that furthers our understanding of those factors that influence successful egg production and fertilisation in salmonids and other species.

  9. Proteomics Analysis Reveals Previously Uncharacterized Virulence Factors in Vibrio proteolyticus

    Directory of Open Access Journals (Sweden)

    Ann Ray

    2016-07-01

    Full Text Available Members of the genus Vibrio include many pathogens of humans and marine animals that share genetic information via horizontal gene transfer. Hence, the Vibrio pan-genome carries the potential to establish new pathogenic strains by sharing virulence determinants, many of which have yet to be characterized. Here, we investigated the virulence properties of Vibrio proteolyticus, a Gram-negative marine bacterium previously identified as part of the Vibrio consortium isolated from diseased corals. We found that V. proteolyticus causes actin cytoskeleton rearrangements followed by cell lysis in HeLa cells in a contact-independent manner. In search of the responsible virulence factor involved, we determined the V. proteolyticus secretome. This proteomics approach revealed various putative virulence factors, including active type VI secretion systems and effectors with virulence toxin domains; however, these type VI secretion systems were not responsible for the observed cytotoxic effects. Further examination of the V. proteolyticus secretome led us to hypothesize and subsequently demonstrate that a secreted hemolysin, belonging to a previously uncharacterized clan of the leukocidin superfamily, was the toxin responsible for the V. proteolyticus-mediated cytotoxicity in both HeLa cells and macrophages. Clearly, there remains an armory of yet-to-be-discovered virulence factors in the Vibrio pan-genome that will undoubtedly provide a wealth of knowledge on how a pathogen can manipulate host cells.

  10. Proteomics analysis of Bacillus licheniformis in response to oligosaccharides elicitors.

    Science.gov (United States)

    Reffatti, Patricia Fernanda; Roy, Ipsita; Odell, Mark; Keshavarz, Tajalli

    2014-01-01

    The role of oligosaccharides as biotic elicitors has been recognised in the enhanced production of antibiotics from fungal and bacterial cultures. The yield of bacitracin A in cultures of Bacillus licheniformis was increased after supplementation with oligoguluronate (OG), and mannan oligosaccharides (MO) and its mechanism at transcription level been established already. However, the elicitation mechanism at post transcriptional level has not been reported so far. In this paper we investigate changes in proteomics of B. licheniformis in presence of the oligosaccharide elicitors OG and MO. Differentially expressed proteins were examined using 2D-PAGE stained with colloidal Coomassie and were further identified by LC-MS/MS. We identified 19 differentially expressed proteins including those involved in carbon metabolism, energy generation, amino acid biosynthesis, oxidative and general stress response. The novel findings of this work, together with previous reports, contribute to the unravelling of the overall mechanism of elicitation in B. licheniformis cultures and reliability of the use of these elicitors for potential industrial application. Copyright © 2014 Elsevier Inc. All rights reserved.

  11. Proteomic analysis of rat cerebral cortex following subchronic acrolein toxicity.

    Science.gov (United States)

    Rashedinia, Marzieh; Lari, Parisa; Abnous, Khalil; Hosseinzadeh, Hossein

    2013-10-01

    Acrolein, a member of reactive α,β-unsaturated aldehydes, is a major environmental pollutant. Acrolein is also produced endogenously as a toxic by-product of lipid peroxidation. Because of high reactivity, acrolein may mediate oxidative damages to cells and tissues. It has been shown to be involved in a wide variety of pathological states including pulmonary, atherosclerosis and neurodegenerative diseases. In this study we employed proteomics approach to investigate the effects of subchronic oral exposures to 3mg/kg of acrolein on protein expression profile in the brain of rats. Moreover effects of acrolein on malondialdehyde (MDA) levels and reduced glutathione (GSH) content were investigated. Our results revealed that treatment with acrolein changed levels of several proteins in diverse physiological process including energy metabolism, cell communication and transport, response to stimulus and metabolic process. Interestingly, several differentially over-expressed proteins, including β-synuclein, enolase and calcineurin, are known to be associated with human neurodegenerative diseases. Changes in the levels of some proteins were confirmed by Western blot. Moreover, acrolein increases the level of MDA, as a lipid peroxidation biomarker and decreased GSH concentrations, as a non-enzyme antioxidant in the brain of acrolein treated rats. These findings suggested that acrolein induces the oxidative stress and lipid peroxidation in the brain, and so that may contribute to the pathophysiology of neurological disorders. Copyright © 2013 Elsevier Inc. All rights reserved.

  12. A Proteomic Analysis of Sarcoptes scabiei (Acari: Sarcoptidae)

    Science.gov (United States)

    Morgan, Marjorie S.; Arlian, Larry G.; Rider, S. Dean; Grunwald, William C.; Cool, David R.

    2016-01-01

    The pruritic skin disease scabies is caused by the burrowing of the itch mite Sarcoptes scabiei (De Geer). It is difficult to diagnose this disease because its symptoms often resemble those of other skin diseases. No reliable blood or molecular diagnostic test is available. The aim of this project was to begin to characterize the scabies proteome to identify scabies mite proteins, including those that may be useful in the development of a diagnostic test or vaccine. Various scabies mite extracts were separated by two-dimensional electrophoresis, and 844 Coomassie Blue-stained protein spots were excised, subjected to trypsin digestion, and analyzed by Matrix Assisted Laser Desorption/Ionization Time-Of-Flight/Time-Of-Flight (MALDI-TOF/TOF) mass spectrometry (MS). Tryptic fragment sequences determined by MS were searched against the recently completed S. scabiei annotated genome, leading to the identification of >150 proteins. Only 10 proteins hit to previously identified scabies proteins including actin, tropomyosin, and several ABC transporters. Thirteen proteins had homology to dust mite allergens (members of groups 8, 10, 13, 17, 20, 25, and 28). Most other sequences showed some homology to proteins in other mites and ticks including homologs of calmodulin, calreticulin, lipocalin, and glutathione-S-transferase. These data will now allow the identification of the proteins to which scabies patients produce antibodies, including those that may be good candidates for inclusion in a diagnostic test and vaccine. PMID:26792847

  13. Proteomic Analysis of Hylocereus polyrhizus Reveals Metabolic Pathway Changes

    Directory of Open Access Journals (Sweden)

    Qingzhu Hua

    2016-09-01

    Full Text Available Red dragon fruit or red pitaya (Hylocereus polyrhizus is the only edible fruit that contains betalains. The color of betalains ranges from red and violet to yellow in plants. Betalains may also serve as an important component of health-promoting and disease-preventing functional food. Currently, the biosynthetic and regulatory pathways for betalain production remain to be fully deciphered. In this study, isobaric tags for relative and absolute quantitation (iTRAQ-based proteomic analyses were used to reveal the molecular mechanism of betalain biosynthesis in H. polyrhizus fruits at white and red pulp stages, respectively. A total of 1946 proteins were identified as the differentially expressed between the two samples, and 936 of them were significantly highly expressed at the red pulp stage of H. polyrhizus. RNA-seq and iTRAQ analyses showed that some transcripts and proteins were positively correlated; they belonged to “phenylpropanoid biosynthesis”, “tyrosine metabolism”, “flavonoid biosynthesis”, “ascorbate and aldarate metabolism”, “betalains biosynthesis” and “anthocyanin biosynthesis”. In betalains biosynthesis pathway, several proteins/enzymes such as polyphenol oxidase, CYP76AD3 and 4,5-dihydroxy-phenylalanine (DOPA dioxygenase extradiol-like protein were identified. The present study provides a new insight into the molecular mechanism of the betalain biosynthesis at the posttranscriptional level.

  14. Proteomic analysis of albumin and globulin fractions of pea (Pisum sativum L.) seeds.

    Science.gov (United States)

    Dziuba, Jerzy; Szerszunowicz, Iwona; Nałęcz, Dorota; Dziuba, Marta

    2014-01-01

    Proteomic analysis is emerging as a highly useful tool in food research, including studies of food allergies. Two-dimensional gel electrophoresis involving isoelectric focusing and sodium dodecyl sulfate polyacrylamide gel electrophoresis is the most effective method of separating hundreds or even thousands of proteins. In this study, albumin and globulin tractions of pea seeds cv. Ramrod were subjected to proteomic analysis. Selected potentially alergenic proteins were identified based on their molecular weights and isoelectric points. Pea seeds (Pisum sativum L.) cv. Ramrod harvested over a period of two years (Plant Breeding Station in Piaski-Szelejewo) were used in the experiment. The isolated albumins, globulins and legumin and vicilin fractions of globulins were separated by two-dimensional gel electrophoresis. Proteomic images were analysed in the ImageMaster 2D Platinum program with the use of algorithms from the Melanie application. The relative content, isoelectric points and molecular weights were computed for all identified proteins. Electrophoregrams were analysed by matching spot positions from three independent replications. The proteomes of albumins, globulins and legumin and vicilin fractions of globulins produced up to several hundred spots (proteins). Spots most characteristic of a given fraction were identified by computer analysis and spot matching. The albumin proteome accumulated spots of relatively high intensity over a broad range of pi values of ~4.2-8.1 in 3 molecular weight (MW) ranges: I - high molecular-weight albumins with MW of ~50-110 kDa, II - average molecular-weight albumins with MW of ~20-35 kDa, and III - low molecular-weight albumins with MW of ~13-17 kDa. 2D gel electrophoregrams revealed the presence of 81 characteristic spots, including 24 characteristic of legumin and 14 - of vicilin. Two-dimensional gel electrophoresis proved to be a useful tool for identifying pea proteins. Patterns of spots with similar isoelectric

  15. Proteome stability analysis of snap frozen, RNAlater preserved, and formalin-fixed paraffin-embedded human colon mucosal biopsies

    Directory of Open Access Journals (Sweden)

    Tue Bjerg Bennike

    2016-03-01

    Full Text Available Large repositories of well characterized RNAlater preserved samples and formalin-fixed, paraffin-embedded samples have been generated worldwide. However, the impact on the proteome of the preservation methods remain poorly described. Therefore, we analyzed the impact on the proteome of preserving samples in RNAlater, and by formalin-fixation, paraffin-embedding on human soft tissue, using directly frozen samples as a control (“Comparing the proteome of snap frozen, RNAlater preserved, and formalin-fixed paraffin-embedded human tissue samples” [1]. We here report the data from the analysis. The comparative analysis was performed on 24 colon mucosa biopsies, extracted from the sigmoideum of two gastroenterologically healthy participants for the purpose of this study. A set of biopsies were additionally stored for 30 min at room temperature prior to formalin-fixation. The samples were analyzed by high throughput gel free quantitative proteomics. The MS proteomics data have been deposited to the ProteomeXchange Consortium via the PRIDE partner repository with the dataset identifier PRIDE: http://www.ebi.ac.uk/pride/archive/projects/PXD002029. Keywords: Human, Colon, Mucosa, RNAlater, FFPE, Snap-frozen, Stability, LC–MS, Proteomics

  16. Data-Independent Acquisition-Based Quantitative Proteomic Analysis Reveals Potential Biomarkers of Kidney Cancer.

    Science.gov (United States)

    Song, Yimeng; Zhong, Lijun; Zhou, Juntuo; Lu, Min; Xing, Tianying; Ma, Lulin; Shen, Jing

    2017-12-01

    Renal cell carcinoma (RCC) is a malignant and metastatic cancer with 95% mortality, and clear cell RCC (ccRCC) is the most observed among the five major subtypes of RCC. Specific biomarkers that can distinguish cancer tissues from adjacent normal tissues should be developed to diagnose this disease in early stages and conduct a reliable prognostic evaluation. Data-independent acquisition (DIA) strategy has been widely employed in proteomic analysis because of various advantages, including enhanced protein coverage and reliable data acquisition. In this study, a DIA workflow is constructed on a quadrupole-Orbitrap LC-MS platform to reveal dysregulated proteins between ccRCC and adjacent normal tissues. More than 4000 proteins are identified, 436 of these proteins are dysregulated in ccRCC tissues. Bioinformatic analysis reveals that multiple pathways and Gene Ontology items are strongly associated with ccRCC. The expression levels of L-lactate dehydrogenase A chain, annexin A4, nicotinamide N-methyltransferase, and perilipin-2 examined through RT-qPCR, Western blot, and immunohistochemistry confirm the validity of the proteomic analysis results. The proposed DIA workflow yields optimum time efficiency and data reliability and provides a good choice for proteomic analysis in biological and clinical studies, and these dysregulated proteins might be potential biomarkers for ccRCC diagnosis. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  17. HiQuant: Rapid Postquantification Analysis of Large-Scale MS-Generated Proteomics Data.

    Science.gov (United States)

    Bryan, Kenneth; Jarboui, Mohamed-Ali; Raso, Cinzia; Bernal-Llinares, Manuel; McCann, Brendan; Rauch, Jens; Boldt, Karsten; Lynn, David J

    2016-06-03

    Recent advances in mass-spectrometry-based proteomics are now facilitating ambitious large-scale investigations of the spatial and temporal dynamics of the proteome; however, the increasing size and complexity of these data sets is overwhelming current downstream computational methods, specifically those that support the postquantification analysis pipeline. Here we present HiQuant, a novel application that enables the design and execution of a postquantification workflow, including common data-processing steps, such as assay normalization and grouping, and experimental replicate quality control and statistical analysis. HiQuant also enables the interpretation of results generated from large-scale data sets by supporting interactive heatmap analysis and also the direct export to Cytoscape and Gephi, two leading network analysis platforms. HiQuant may be run via a user-friendly graphical interface and also supports complete one-touch automation via a command-line mode. We evaluate HiQuant's performance by analyzing a large-scale, complex interactome mapping data set and demonstrate a 200-fold improvement in the execution time over current methods. We also demonstrate HiQuant's general utility by analyzing proteome-wide quantification data generated from both a large-scale public tyrosine kinase siRNA knock-down study and an in-house investigation into the temporal dynamics of the KSR1 and KSR2 interactomes. Download HiQuant, sample data sets, and supporting documentation at http://hiquant.primesdb.eu .

  18. Direct digestion of proteins in living cells into peptides for proteomic analysis.

    Science.gov (United States)

    Chen, Qi; Yan, Guoquan; Gao, Mingxia; Zhang, Xiangmin

    2015-01-01

    To analyze the proteome of an extremely low number of cells or even a single cell, we established a new method of digesting whole cells into mass-spectrometry-identifiable peptides in a single step within 2 h. Our sampling method greatly simplified the processes of cell lysis, protein extraction, protein purification, and overnight digestion, without compromising efficiency. We used our method to digest hundred-scale cells. As far as we know, there is no report of proteome analysis starting directly with as few as 100 cells. We identified an average of 109 proteins from 100 cells, and with three replicates, the number of proteins rose to 204. Good reproducibility was achieved, showing stability and reliability of the method. Gene Ontology analysis revealed that proteins in different cellular compartments were well represented.

  19. Proteomic analysis of rutin-induced secreted proteins from Aspergillus flavus.

    Science.gov (United States)

    Medina, Martha L; Kiernan, Urban A; Francisco, Wilson A

    2004-03-01

    Few studies have been conducted to identify the extracellular proteins and enzymes secreted by filamentous fungi, particularly with respect to dispensable metabolic pathways. Proteomic analysis has proven to be the most powerful method for identification of proteins in complex mixtures and is suitable for the study of the alteration of protein expression under different environmental conditions. The filamentous fungus Aspergillus flavus can degrade the flavonoid rutin as the only source of carbon via an extracellular enzyme system. In this study, a proteomic analysis was used to differentiate and identify the extracellular rutin-induced and non-induced proteins secreted by A. flavus. The secreted proteins were analyzed by two-dimensional electrophoresis and MALDI-TOF mass spectrometry. While 15 rutin-induced proteins and 7 non-induced proteins were identified, more than 90 protein spots remain unidentified, indicating that these proteins are either novel proteins or proteins that have not yet been sequenced.

  20. Proteomic analysis of MG132-treated germinating pollen reveals expression signatures associated with proteasome inhibition.

    Directory of Open Access Journals (Sweden)

    Candida Vannini

    Full Text Available Chemical inhibition of the proteasome has been previously found to effectively impair pollen germination and tube growth in vitro. However, the mediators of these effects at the molecular level are unknown. By performing 2DE proteomic analysis, 24 differentially expressed protein spots, representing 14 unique candidate proteins, were identified in the pollen of kiwifruit (Actinidia deliciosa germinated in the presence of the MG132 proteasome inhibitor. qPCR analysis revealed that 11 of these proteins are not up-regulated at the mRNA level, but are most likely stabilized by proteasome inhibition. These differentially expressed proteins are predicted to function in various pathways including energy and lipid metabolism, cell wall synthesis, protein synthesis/degradation and stress responses. In line with this evidence, the MG132-induced changes in the proteome were accompanied by an increase in ATP and ROS content and by an alteration in fatty acid composition.

  1. Proteome stability analysis of snap frozen, RNAlater preserved, and formalin-fixed paraffin-embedded human colon mucosal biopsies

    DEFF Research Database (Denmark)

    Bjerg Bennike, Tue; Kastaniegaard, Kenneth; Padurariu, Simona

    2016-01-01

    Large repositories of well characterized RNAlater preserved samples and formalin-fixed, paraffin-embedded samples have been generated worldwide. However, the impact on the proteome of the preservation methods remain poorly described. Therefore, we analyzed the impact on the proteome of preserving...... samples in RNAlater, and by formalin-fixation, paraffin-embedding on human soft tissue, using directly frozen samples as a control ("Comparing the proteome of snap frozen, RNAlater preserved, and formalin-fixed paraffin-embedded human tissue samples" [1]). We here report the data from the analysis...

  2. Proteomic and Microscopic Strategies towards the Analysis of the Cytoskeletal Networks in Major Neuropsychiatric Disorders

    Directory of Open Access Journals (Sweden)

    Joëlle V. F. Coumans

    2016-04-01

    Full Text Available Mental health disorders have become worldwide health priorities. It is estimated that in the next 20 years they will account for a 16 trillion United State dollars (US$ loss. Up to now, the underlying pathophysiology of psychiatric disorders remains elusive. Altered cytoskeleton proteins expression that may influence the assembly, organization and maintenance of cytoskeletal integrity has been reported in major depressive disorders, schizophrenia and to some extent bipolar disorders. The use of quantitative proteomics, dynamic microscopy and super-resolution microscopy to investigate disease-specific protein signatures holds great promise to improve our understanding of these disorders. In this review, we present the currently available quantitative proteomic approaches use in neurology, gel-based, stable isotope-labelling and label-free methodologies and evaluate their strengths and limitations. We also reported on enrichment/subfractionation methods that target the cytoskeleton associated proteins and discuss the need of alternative methods for further characterization of the neurocytoskeletal proteome. Finally, we present live cell imaging approaches and emerging dynamic microscopy technology that will provide the tools necessary to investigate protein interactions and their dynamics in the whole cells. While these areas of research are still in their infancy, they offer huge potential towards the understanding of the neuronal network stability and its modification across neuropsychiatric disorders.

  3. Comparative Proteomics Analysis of Placenta from Pregnant Women with Intrahepatic Cholestasis of Pregnancy

    OpenAIRE

    Zhang, Ting; Guo, Yueshuai; Guo, Xuejiang; Zhou, Tao; Chen, Daozhen; Xiang, Jingying; Zhou, Zuomin

    2013-01-01

    INTRODUCTION: Intrahepatic cholestasis of pregnancy (ICP) usually occurs in the third trimester and associated with increased risks in fetal complications. Currently, the exact cause of this disease is unknown. In this study we aim to investigate the potential proteins in placenta, which may participate in the molecular mechanisms of ICP-related fetal complications using iTRAQ-based proteomics approach. METHODS: The iTRAQ analysis combined with liquid chromatography-tandem mass spectrometry (...

  4. A proteomic analysis of the chromoplasts isolated from sweet orange fruits [Citrus sinensis (L.) Osbeck

    OpenAIRE

    Zeng, Yunliu; Pan, Zhiyong; Ding, Yuduan; Zhu, Andan; Cao, Hongbo; Xu, Qiang; Deng, Xiuxin

    2011-01-01

    Here, a comprehensive proteomic analysis of the chromoplasts purified from sweet orange using Nycodenz density gradient centrifugation is reported. A GeLC-MS/MS shotgun approach was used to identify the proteins of pooled chromoplast samples. A total of 493 proteins were identified from purified chromoplasts, of which 418 are putative plastid proteins based on in silico sequence homology and functional analyses. Based on the predicted functions of these identified plastid proteins, a large pr...

  5. Proteomic analysis of chromoplasts from six crop species reveals insights into chromoplast function and development

    OpenAIRE

    Wang, Yong-Qiang; Yang, Yong; Fei, Zhangjun; Yuan, Hui; Fish, Tara; Thannhauser, Theodore W.; Mazourek, Michael; Kochian, Leon V.; Wang, Xiaowu; Li, Li

    2013-01-01

    Chromoplasts are unique plastids that accumulate massive amounts of carotenoids. To gain a general and comparative characterization of chromoplast proteins, this study performed proteomic analysis of chromoplasts from six carotenoid-rich crops: watermelon, tomato, carrot, orange cauliflower, red papaya, and red bell pepper. Stromal and membrane proteins of chromoplasts were separated by 1D gel electrophoresis and analysed using nLC-MS/MS. A total of 953?2262 proteins from chromoplasts of diff...

  6. Towards the Development of Proteomics Workflows for the Analysis of Samples Derived from Refractory Plant Tissues

    OpenAIRE

    Thannhauser, T.W.

    2011-01-01

    Carrying out proteomic analyses in plant tissues involves dealing with a number of specialized challenges that can make protein extraction and quantification significantly more difficult than in other organisms. In addition to having relatively low protein concentrations, plant tissues are often rich in proteases, protease inhibitors and other materials that impede protein analysis. These compounds include lipids, tannins, polysaccharides, and a large variety of secondary metabolites. The ext...

  7. Proteomic analysis of venom variability and ontogeny across the arboreal palm-pitvipers (genus Bothriechis).

    Science.gov (United States)

    Pla, Davinia; Sanz, Libia; Sasa, Mahmood; Acevedo, Manuel E; Dwyer, Quetzal; Durban, Jordi; Pérez, Alicia; Rodriguez, Yania; Lomonte, Bruno; Calvete, Juan J

    2017-01-30

    -rich venom, whereas in B. schlegelii the age-dependent compositional shift generates a PLA 2 -rich venom. Overall, genus-wide venomics illustrate the high evolvability of palm-pitviper venoms. The integration of the pattern of venom variation across Bothriechis into a phylogenetic and biogeographic framework may lay the foundation for assessing, in future studies, the evolutionary path that led to the present-day variability of the venoms of palm-pitvipers. Bothriechis represents a monophyletic basal genus of eleven arboreal palm-pitvipers that range from southern Mexico to northern South America. Despite palm-pitvipers' putative status as diet generalists, previous proteomic analyses have revealed remarkable divergence between the venoms of Costa Rican species, B. lateralis, B. schlegelii, B. supraciliaris, and B. nigroviridis. Our current proteomic study of Guatemalan species, B. thalassinus, B. aurifer, and B. bicolor, Honduran B. marchi, and neonate B. lateralis and B. schlegelii from Costa Rica was undertaken to deepen our understanding of the evolutionary pattern of venom proteome diversity across Bothriechis. Ancestral characters are often, but not always, preserved in an organism's development. Venoms of neonate B. lateralis and B. schlegelii are more similar to each other than to adults of their respective species, suggesting that the high evolvability of palm-pitviper venoms may represent an inherent feature of Bothriechis common ancestor. Our genus-wide data identified four nodes of venom phenotype differentiation across the phylogeny of Bothriechis. Integrated into a phylogenetic and biogeographic framework, the pattern of venom variation across Bothriechis may lay the groundwork to establish whether divergence was driven by selection for efficient resource exploitation in arboreal 'islands', thereby contributing to the ecological speciation of the genus. Copyright © 2016 Elsevier B.V. All rights reserved.

  8. Proteomic analysis of rat cerebral cortex following subchronic acrolein toxicity

    Energy Technology Data Exchange (ETDEWEB)

    Rashedinia, Marzieh; Lari, Parisa [Department of Pharmacodynamics and Toxicology, School of Pharmacy, Mashhad University of Medical Sciences, Mashhad (Iran, Islamic Republic of); Abnous, Khalil, E-mail: Abnouskh@mums.ac.r [Pharmaceutical Research Center, Department of Medicinal Chemistry, Mashhad University of Medical Sciences, Mashhad (Iran, Islamic Republic of); Hosseinzadeh, Hossein, E-mail: Hosseinzadehh@mums.ac.ir [Pharmaceutical Research Center, Department of Pharmacodynamics and Toxicology, School of Pharmacy, Mashhad University of Medical Sciences, Mashhad (Iran, Islamic Republic of)

    2013-10-01

    Acrolein, a member of reactive α,β-unsaturated aldehydes, is a major environmental pollutant. Acrolein is also produced endogenously as a toxic by-product of lipid peroxidation. Because of high reactivity, acrolein may mediate oxidative damages to cells and tissues. It has been shown to be involved in a wide variety of pathological states including pulmonary, atherosclerosis and neurodegenerative diseases. In this study we employed proteomics approach to investigate the effects of subchronic oral exposures to 3 mg/kg of acrolein on protein expression profile in the brain of rats. Moreover effects of acrolein on malondialdehyde (MDA) levels and reduced glutathione (GSH) content were investigated. Our results revealed that treatment with acrolein changed levels of several proteins in diverse physiological process including energy metabolism, cell communication and transport, response to stimulus and metabolic process. Interestingly, several differentially over-expressed proteins, including β-synuclein, enolase and calcineurin, are known to be associated with human neurodegenerative diseases. Changes in the levels of some proteins were confirmed by Western blot. Moreover, acrolein increases the level of MDA, as a lipid peroxidation biomarker and decreased GSH concentrations, as a non-enzyme antioxidant in the brain of acrolein treated rats. These findings suggested that acrolein induces the oxidative stress and lipid peroxidation in the brain, and so that may contribute to the pathophysiology of neurological disorders. - Highlights: • Acrolein intoxication increased lipid peroxidation and deplete GSH in rat brain. • Effect of acrolein on protein levels of cerebral cortex was analyzed by 2DE-PAGE. • Levels of a number of proteins with different biological functions were increased.

  9. Proteomic analysis of rat cerebral cortex following subchronic acrolein toxicity

    International Nuclear Information System (INIS)

    Rashedinia, Marzieh; Lari, Parisa; Abnous, Khalil; Hosseinzadeh, Hossein

    2013-01-01

    Acrolein, a member of reactive α,β-unsaturated aldehydes, is a major environmental pollutant. Acrolein is also produced endogenously as a toxic by-product of lipid peroxidation. Because of high reactivity, acrolein may mediate oxidative damages to cells and tissues. It has been shown to be involved in a wide variety of pathological states including pulmonary, atherosclerosis and neurodegenerative diseases. In this study we employed proteomics approach to investigate the effects of subchronic oral exposures to 3 mg/kg of acrolein on protein expression profile in the brain of rats. Moreover effects of acrolein on malondialdehyde (MDA) levels and reduced glutathione (GSH) content were investigated. Our results revealed that treatment with acrolein changed levels of several proteins in diverse physiological process including energy metabolism, cell communication and transport, response to stimulus and metabolic process. Interestingly, several differentially over-expressed proteins, including β-synuclein, enolase and calcineurin, are known to be associated with human neurodegenerative diseases. Changes in the levels of some proteins were confirmed by Western blot. Moreover, acrolein increases the level of MDA, as a lipid peroxidation biomarker and decreased GSH concentrations, as a non-enzyme antioxidant in the brain of acrolein treated rats. These findings suggested that acrolein induces the oxidative stress and lipid peroxidation in the brain, and so that may contribute to the pathophysiology of neurological disorders. - Highlights: • Acrolein intoxication increased lipid peroxidation and deplete GSH in rat brain. • Effect of acrolein on protein levels of cerebral cortex was analyzed by 2DE-PAGE. • Levels of a number of proteins with different biological functions were increased

  10. Proteomic analysis of tyrosine phosphorylation during human liver transplantation

    Directory of Open Access Journals (Sweden)

    Boutros Tarek

    2007-01-01

    Full Text Available Abstract Background Ischemia-reperfusion (I/R causes a dramatic reprogramming of cell metabolism during liver transplantation and can be linked to an alteration of the phosphorylation level of several cellular proteins. Over the past two decades, it became clear that tyrosine phosphorylation plays a pivotal role in a variety of important signalling pathways and was linked to a wide spectrum of diseases. Functional profiling of the tyrosine phosphoproteome during liver transplantation is therefore of great biological significance and is likely to lead to the identification of novel targets for drug discovery and provide a basis for novel therapeutic strategies. Results Using liver biopsies collected during the early phases of organ procurement and transplantation, we aimed at characterizing the global patterns of tyrosine phosphorylation during hepatic I/R. A proteomic approach, based on the purification of tyrosine phosphorylated proteins followed by their identification using mass spectrometry, allowed us to identify Nck-1, a SH2/SH3 adaptor, as a potential regulator of I/R injury. Using immunoblot, cell fractionation and immunohistochemistry, we demonstrate that Nck-1 phosphorylation, expression and localization were affected in liver tissue upon I/R. In addition, mass spectrometry identification of Nck-1 binding partners during the course of the transplantation also suggested a dynamic interaction between Nck-1 and actin during I/R. Conclusion Taken together, our data suggest that Nck-1 may play a role in I/R-induced actin reorganization, which was previously reported to be detrimental for the hepatocytes of the transplanted graft. Nck-1 could therefore represent a target of choice for the design of new organ preservation strategies, which could consequently help to reduce post-reperfusion liver damages and improve transplantation outcomes.

  11. A DIGE proteomic analysis for high-intensity exercise-trained rat skeletal muscle.

    Science.gov (United States)

    Yamaguchi, Wataru; Fujimoto, Eri; Higuchi, Mitsuru; Tabata, Izumi

    2010-09-01

    Exercise training induces various adaptations in skeletal muscles. However, the mechanisms remain unclear. In this study, we conducted 2D-DIGE proteomic analysis, which has not yet been used for elucidating adaptations of skeletal muscle after high-intensity exercise training (HIT). For 5 days, rats performed HIT, which consisted of 14 20-s swimming exercise bouts carrying a weight (14% of the body weight), and 10-s pause between bouts. The 2D-DIGE analysis was conducted on epitrochlearis muscles excised 18 h after the final training exercise. Proteomic profiling revealed that out of 800 detected and matched spots, 13 proteins exhibited changed expression by HIT compared with sedentary rats. All proteins were identified by MALDI-TOF/MS. Furthermore, using western immunoblot analyses, significantly changed expressions of NDUFS1 and parvalbumin (PV) were validated in relation to HIT. In conclusion, the proteomic 2D-DIGE analysis following HIT-identified expressions of NDUFS1 and PV, previously unknown to have functions related to exercise-training adaptations.

  12. Quantitative Proteomics for the Comprehensive Analysis of Stress Responses of Lactobacillus paracasei subsp. paracasei F19.

    Science.gov (United States)

    Schott, Ann-Sophie; Behr, Jürgen; Geißler, Andreas J; Kuster, Bernhard; Hahne, Hannes; Vogel, Rudi F

    2017-10-06

    Lactic acid bacteria are broadly employed as starter cultures in the manufacture of foods. Upon technological preparation, they are confronted with drying stress that amalgamates numerous stress conditions resulting in losses of fitness and survival. To better understand and differentiate physiological stress responses, discover general and specific markers for the investigated stress conditions, and predict optimal preconditioning for starter cultures, we performed a comprehensive genomic and quantitative proteomic analysis of a commonly used model system, Lactobacillus paracasei subsp. paracasei TMW 1.1434 (isogenic with F19) under 11 typical stress conditions, including among others oxidative, osmotic, pH, and pressure stress. We identified and quantified >1900 proteins in triplicate analyses, representing 65% of all genes encoded in the genome. The identified genes were thoroughly annotated in terms of subcellular localization prediction and biological functions, suggesting unbiased and comprehensive proteome coverage. In total, 427 proteins were significantly differentially expressed in at least one condition. Most notably, our analysis suggests that optimal preconditioning toward drying was predicted to be alkaline and high-pressure stress preconditioning. Taken together, we believe the presented strategy may serve as a prototypic example for the analysis and utility of employing quantitative-mass-spectrometry-based proteomics to study bacterial physiology.

  13. Relative Quantitative Proteomic Analysis of Brucella abortus Reveals Metabolic Adaptation to Multiple Environmental Stresses.

    Science.gov (United States)

    Zai, Xiaodong; Yang, Qiaoling; Yin, Ying; Li, Ruihua; Qian, Mengying; Zhao, Taoran; Li, Yaohui; Zhang, Jun; Fu, Ling; Xu, Junjie; Chen, Wei

    2017-01-01

    Brucella spp. are facultative intracellular pathogens that cause chronic brucellosis in humans and animals. The virulence of Brucella primarily depends on its successful survival and replication in host cells. During invasion of the host tissue, Brucella is simultaneously subjected to a variety of harsh conditions, including nutrient limitation, low pH, antimicrobial defenses, and extreme levels of reactive oxygen species (ROS) via the host immune response. This suggests that Brucella may be able to regulate its metabolic adaptation in response to the distinct stresses encountered during its intracellular infection of the host. An investigation into the differential proteome expression patterns of Brucella grown under the relevant stress conditions may contribute toward a better understanding of its pathogenesis and adaptive response. Here, we utilized a mass spectrometry-based label-free relative quantitative proteomics approach to investigate and compare global proteomic changes in B. abortus in response to eight different stress treatments. The 3 h short-term in vitro single-stress and multi-stress conditions mimicked the in vivo conditions of B. abortus under intracellular infection, with survival rates ranging from 3.17 to 73.17%. The proteomic analysis identified and quantified a total of 2,272 proteins and 74% of the theoretical proteome, thereby providing wide coverage of the B. abortus proteome. By including eight distinct growth conditions and comparing these with a control condition, we identified a total of 1,221 differentially expressed proteins (DEPs) that were significantly changed under the stress treatments. Pathway analysis revealed that most of the proteins were involved in oxidative phosphorylation, ABC transporters, two-component systems, biosynthesis of secondary metabolites, the citrate cycle, thiamine metabolism, and nitrogen metabolism; constituting major response mechanisms toward the reconstruction of cellular homeostasis and metabolic

  14. Relative Quantitative Proteomic Analysis of Brucella abortus Reveals Metabolic Adaptation to Multiple Environmental Stresses

    Directory of Open Access Journals (Sweden)

    Xiaodong Zai

    2017-11-01

    Full Text Available Brucella spp. are facultative intracellular pathogens that cause chronic brucellosis in humans and animals. The virulence of Brucella primarily depends on its successful survival and replication in host cells. During invasion of the host tissue, Brucella is simultaneously subjected to a variety of harsh conditions, including nutrient limitation, low pH, antimicrobial defenses, and extreme levels of reactive oxygen species (ROS via the host immune response. This suggests that Brucella may be able to regulate its metabolic adaptation in response to the distinct stresses encountered during its intracellular infection of the host. An investigation into the differential proteome expression patterns of Brucella grown under the relevant stress conditions may contribute toward a better understanding of its pathogenesis and adaptive response. Here, we utilized a mass spectrometry-based label-free relative quantitative proteomics approach to investigate and compare global proteomic changes in B. abortus in response to eight different stress treatments. The 3 h short-term in vitro single-stress and multi-stress conditions mimicked the in vivo conditions of B. abortus under intracellular infection, with survival rates ranging from 3.17 to 73.17%. The proteomic analysis identified and quantified a total of 2,272 proteins and 74% of the theoretical proteome, thereby providing wide coverage of the B. abortus proteome. By including eight distinct growth conditions and comparing these with a control condition, we identified a total of 1,221 differentially expressed proteins (DEPs that were significantly changed under the stress treatments. Pathway analysis revealed that most of the proteins were involved in oxidative phosphorylation, ABC transporters, two-component systems, biosynthesis of secondary metabolites, the citrate cycle, thiamine metabolism, and nitrogen metabolism; constituting major response mechanisms toward the reconstruction of cellular

  15. Semen proteomics and male infertility.

    Science.gov (United States)

    Jodar, Meritxell; Soler-Ventura, Ada; Oliva, Rafael

    2017-06-06

    Semen is a complex body fluid containing an admixture of spermatozoa suspended in secretions from the testes and epididymis which are mixed at the time of ejaculation with secretions from other accessory sex glands such as the prostate and seminal vesicles. High-throughput technologies have revealed that, contrary to the idea that sperm cells are simply a silent delivery vehicle of the male genome to the oocyte, the sperm cells in fact provide both a specific epigenetically marked DNA together with a complex population of proteins and RNAs crucial for embryogenesis. Similarly, -omic technologies have also enlightened that seminal fluid seems to play a much greater role than simply being a medium to carry the spermatozoa through the female reproductive tract. In the present review, we briefly overview the sperm cell biology, consider the key issues in sperm and seminal fluid sample preparation for high-throughput proteomic studies, describe the current state of the sperm and seminal fluid proteomes generated by high-throughput proteomic technologies and provide new insights into the potential communication between sperm and seminal fluid. In addition, comparative proteomic studies open a window to explore the potential pathogenic mechanisms of infertility and the discovery of potential biomarkers with clinical significance. The review updates the numerous proteomics studies performed on semen, including spermatozoa and seminal fluid. In addition, an integrative analysis of the testes, sperm and seminal fluid proteomes is also included providing insights into the molecular mechanisms that regulate the generation, maturation and transit of spermatozoa. Furthermore, the compilation of several differential proteomic studies focused on male infertility reveals potential pathways disturbed in specific subtypes of male infertility and points out towards future research directions in the field. Copyright © 2016 Elsevier B.V. All rights reserved.

  16. The Human Proteome Organization Chromosome 6 Consortium: integrating chromosome-centric and biology/disease driven strategies.

    Science.gov (United States)

    Borchers, C H; Kast, J; Foster, L J; Siu, K W M; Overall, C M; Binkowski, T A; Hildebrand, W H; Scherer, A; Mansoor, M; Keown, P A

    2014-04-04

    The Human Proteome Project (HPP) is designed to generate a comprehensive map of the protein-based molecular architecture of the human body, to provide a resource to help elucidate biological and molecular function, and to advance diagnosis and treatment of diseases. Within this framework, the chromosome-based HPP (C-HPP) has allocated responsibility for mapping individual chromosomes by country or region, while the biology/disease HPP (B/D-HPP) coordinates these teams in cross-functional disease-based groups. Chromosome 6 (Ch6) provides an excellent model for integration of these two tasks. This metacentric chromosome has a complement of 1002-1034 genes that code for known, novel or putative proteins. Ch6 is functionally associated with more than 120 major human diseases, many with high population prevalence, devastating clinical impact and profound societal consequences. The unique combination of genomic, proteomic, metabolomic, phenomic and health services data being drawn together within the Ch6 program has enormous potential to advance personalized medicine by promoting robust biomarkers, subunit vaccines and new drug targets. The strong liaison between the clinical and laboratory teams, and the structured framework for technology transfer and health policy decisions within Canada will increase the speed and efficacy of this transition, and the value of this translational research. Canada has been selected to play a leading role in the international Human Proteome Project, the global counterpart of the Human Genome Project designed to understand the structure and function of the human proteome in health and disease. Canada will lead an international team focusing on chromosome 6, which is functionally associated with more than 120 major human diseases, including immune and inflammatory disorders affecting the brain, skeletal system, heart and blood vessels, lungs, kidney, liver, gastrointestinal tract and endocrine system. Many of these chronic and persistent

  17. Deep proteome analysis of gerontoplasts from the inner integument of developing seeds of Jatropha curcas.

    Science.gov (United States)

    Shah, Mohibullah; Soares, Emanoella L; Lima, Magda L B; Pinheiro, Camila B; Soares, Arlete A; Domont, Gilberto B; Nogueira, Fabio C S; Campos, Francisco A P

    2016-06-30

    The inner integument of Jatropha curcas seeds is a non-photosynthetic tissue that acts primarily as a conduit for the delivery of nutrients to the embryo and endosperm. In this study we performed a histological and transmission electron microscopy analysis of the inner integument in stages prior to fertilization to 25days after pollination, to establish the structural changes associated with the plastid to gerontoplast transition. This study showed that plastids are subjected to progressive changes, which include the dismantling of the internal membrane system, matrix degradation and the formation of stromule-derived vesicles. A proteome analysis of gerontoplasts isolated from the inner integument at 25days after pollination, resulted in the identification of 1923 proteins, which were involved in a myriad of metabolic functions, such as synthesis of amino acids and fatty acids. Among the identified proteins, were also a number of hydrolases (peptidases, lipases and carbohydrases), which presumably are involved in the ordered dismantling of this organelle to provide additional sources of nutrients for the growing embryo and endosperm. The dataset we provide here may provide a foundation for the study of the proteome changes associated with the plastid to gerontoplast transition in non-photosynthetic tissues. We describe ultrastructural features of gerontoplasts isolated from the inner integument of developing seeds of Jatropha curcas, together with a deep proteome analysis of these gerontoplasts. This article explores a new aspect of the biology of plastids, namely the ultrastructural and proteome changes associated with the transition plastid to gerontoplast in a non-photosynthetic tissue. Copyright © 2016 Elsevier B.V. All rights reserved.

  18. Marine proteomics: a critical assessment of an emerging technology.

    Science.gov (United States)

    Slattery, Marc; Ankisetty, Sridevi; Corrales, Jone; Marsh-Hunkin, K Erica; Gochfeld, Deborah J; Willett, Kristine L; Rimoldi, John M

    2012-10-26

    The application of proteomics to marine sciences has increased in recent years because the proteome represents the interface between genotypic and phenotypic variability and, thus, corresponds to the broadest possible biomarker for eco-physiological responses and adaptations. Likewise, proteomics can provide important functional information regarding biosynthetic pathways, as well as insights into mechanism of action, of novel marine natural products. The goal of this review is to (1) explore the application of proteomics methodologies to marine systems, (2) assess the technical approaches that have been used, and (3) evaluate the pros and cons of this proteomic research, with the intent of providing a critical analysis of its future roles in marine sciences. To date, proteomics techniques have been utilized to investigate marine microbe, plant, invertebrate, and vertebrate physiology, developmental biology, seafood safety, susceptibility to disease, and responses to environmental change. However, marine proteomics studies often suffer from poor experimental design, sample processing/optimization difficulties, and data analysis/interpretation issues. Moreover, a major limitation is the lack of available annotated genomes and proteomes for most marine organisms, including several "model species". Even with these challenges in mind, there is no doubt that marine proteomics is a rapidly expanding and powerful integrative molecular research tool from which our knowledge of the marine environment, and the natural products from this resource, will be significantly expanded.

  19. Integrative proteomics and tissue microarray profiling indicate the association between overexpressed serum proteins and non-small cell lung cancer.

    Directory of Open Access Journals (Sweden)

    Yansheng Liu

    Full Text Available Lung cancer is the leading cause of cancer deaths worldwide. Clinically, the treatment of non-small cell lung cancer (NSCLC can be improved by the early detection and risk screening among population. To meet this need, here we describe the application of extensive peptide level fractionation coupled with label free quantitative proteomics for the discovery of potential serum biomarkers for lung cancer, and the usage of Tissue microarray analysis (TMA and Multiple reaction monitoring (MRM assays for the following up validations in the verification phase. Using these state-of-art, currently available clinical proteomic approaches, in the discovery phase we confidently identified 647 serum proteins, and 101 proteins showed a statistically significant association with NSCLC in our 18 discovery samples. This serum proteomic dataset allowed us to discern the differential patterns and abnormal biological processes in the lung cancer blood. Of these proteins, Alpha-1B-glycoprotein (A1BG and Leucine-rich alpha-2-glycoprotein (LRG1, two plasma glycoproteins with previously unknown function were selected as examples for which TMA and MRM verification were performed in a large sample set consisting about 100 patients. We revealed that A1BG and LRG1 were overexpressed in both the blood level and tumor sections, which can be referred to separate lung cancer patients from healthy cases.

  20. Translational plant proteomics: A perspective

    NARCIS (Netherlands)

    Agrawal, G.K.; Pedreschi, R.; Barkla, B.J.; Bindschedler, L.V.; Cramer, R.; Sarkar, A.; Renaut, J.; Job, D.; Rakwal, R.

    2012-01-01

    Translational proteomics is an emerging sub-discipline of the proteomics field in the biological sciences. Translational plant proteomics aims to integrate knowledge from basic sciences to translate it into field applications to solve issues related but not limited to the recreational and economic

  1. A label-free quantitative shotgun proteomics analysis of rice grain development

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    Koh Hee-Jong

    2011-09-01

    Full Text Available Abstract Background Although a great deal of rice proteomic research has been conducted, there are relatively few studies specifically addressing the rice grain proteome. The existing rice grain proteomic researches have focused on the identification of differentially expressed proteins or monitoring protein expression patterns during grain filling stages. Results Proteins were extracted from rice grains 10, 20, and 30 days after flowering, as well as from fully mature grains. By merging all of the identified proteins in this study, we identified 4,172 non-redundant proteins with a wide range of molecular weights (from 5.2 kDa to 611 kDa and pI values (from pH 2.9 to pH 12.6. A Genome Ontology category enrichment analysis for the 4,172 proteins revealed that 52 categories were enriched, including the carbohydrate metabolic process, transport, localization, lipid metabolic process, and secondary metabolic process. The relative abundances of the 1,784 reproducibly identified proteins were compared to detect 484 differentially expressed proteins during rice grain development. Clustering analysis and Genome Ontology category enrichment analysis revealed that proteins involved in the metabolic process were enriched through all stages of development, suggesting that proteome changes occurred even in the desiccation phase. Interestingly, enrichments of proteins involved in protein folding were detected in the desiccation phase and in fully mature grain. Conclusion This is the first report conducting comprehensive identification of rice grain proteins. With a label free shotgun proteomic approach, we identified large number of rice grain proteins and compared the expression patterns of reproducibly identified proteins during rice grain development. Clustering analysis, Genome Ontology category enrichment analysis, and the analysis of composite expression profiles revealed dynamic changes of metabolisms during rice grain development. Interestingly, we

  2. Proteomic analysis of human tooth pulp proteomes – Comparison of caries-resistant and caries-susceptible

    Czech Academy of Sciences Publication Activity Database

    Jágr, Michal; Eckhardt, Adam; Pataridis, Statis; Foltán, R.; Myšák, J.; Mikšík, Ivan

    2016-01-01

    Roč. 145, Aug 11 (2016), s. 127-136 ISSN 1874-3919 R&D Projects: GA MZd(CZ) NT14324 Institutional support: RVO:67985823 Keywords : human tooth pulp * DIGE * proteome * caries * resistance Subject RIV: FF - HEENT, Dentistry Impact factor: 3.914, year: 2016

  3. Comprehensive proteome analysis of the response of Pseudomonas putida KT2440 to the flavor compound vanillin.

    Science.gov (United States)

    Simon, Oliver; Klaiber, Iris; Huber, Armin; Pfannstiel, Jens

    2014-09-23

    Understanding of the molecular response of bacteria to precursors, products and environmental conditions applied in bioconversions is essential for optimizing whole-cell biocatalysis. To investigate the molecular response of the potential biocatalyst Pseudomonas putida KT2440 to the flavor compound vanillin we applied complementary gel- and LC-MS-based quantitative proteomics approaches. Our comprehensive proteomics survey included cytoplasmic and membrane proteins and led to the identification and quantification of 1614 proteins, corresponding to 30% of the total KT2440 proteome. 662 proteins were altered in abundance during growth on vanillin as sole carbon source as compared to growth on glucose. The proteome response entailed an increased abundance of enzymes involved in vanillin degradation, significant changes in central energy metabolism and an activation of solvent tolerance mechanisms. With respect to vanillin metabolism, particularly enzymes belonging to the β-ketoadipate pathway including a transcriptional regulator and porins specific for vanillin uptake increased in abundance. However, catabolism of vanillin was not dependent on vanillin dehydrogenase (Vdh), as shown by quantitative proteome analysis of a Vdh-deficient KT2440 mutant (GN235). Other aldehyde dehydrogenases that were significantly increased in abundance in response to vanillin may replace Vdh and thus may represent interesting targets for improving vanillin production in P. putida KT2440. The high demand for the flavor compound vanillin by the food and fragrance industry makes natural vanillin from vanilla pods a scarce and expensive resource rendering its biotechnological production economically attractive. Pseudomonas bacteria are metabolically very versatile and accept a broad range of hydrocarbons as carbon source making them suitable candidates for bioconversion processes. This work describes the impact of vanillin on the metabolism of the reference strain P. putida KT2440 on a

  4. Comparative proteomic analysis reveals heart toxicity induced by chronic arsenic exposure in rats

    DEFF Research Database (Denmark)

    Huang, Qingyu; Xi, Guochen; Alamdar, Ambreen

    2017-01-01

    Arsenic is a widespread metalloid in the environment, which poses a broad spectrum of adverse effects on human health. However, a global view of arsenic-induced heart toxicity is still lacking, and the underlying molecular mechanisms remain unclear. By performing a comparative quantitative...... proteomic analysis, the present study aims to investigate the alterations of proteome profile in rat heart after long-term exposure to arsenic. As a result, we found that the abundance of 81 proteins were significantly altered by arsenic treatment (35 up-regulated and 46 down-regulated). Among these, 33...... proteins were specifically associated with cardiovascular system development and function, including heart development, heart morphology, cardiac contraction and dilation, and other cardiovascular functions. It is further proposed that the aberrant regulation of 14 proteins induced by arsenic would disturb...

  5. How to use and integrate bioinformatics tools to compare proteomic data from distinct conditions? A tutorial using the pathological similarities between Aortic Valve Stenosis and Coronary Artery Disease as a case-study.

    Science.gov (United States)

    Trindade, Fábio; Ferreira, Rita; Magalhães, Beatriz; Leite-Moreira, Adelino; Falcão-Pires, Inês; Vitorino, Rui

    2018-01-16

    Nowadays we are surrounded by a plethora of bioinformatics tools, powerful enough to deal with the large amounts of data arising from proteomic studies, but whose application is sometimes hard to find. Therefore, we used a specific clinical problem - to discriminate pathophysiology and potential biomarkers between two similar cardiovascular diseases, aortic valve stenosis (AVS) and coronary artery disease (CAD) - to make a step-by-step guide through four bioinformatics tools: STRING, DisGeNET, Cytoscape and ClueGO. Proteome data was collected from articles available on PubMed centered on proteomic studies enrolling subjects with AVS or CAD. Through the analysis of gene ontology provided by STRING and ClueGO we could find specific biological phenomena associated with AVS, such as down-regulation of elastic fiber assembly, and with CAD, such as up-regulation of plasminogen activation. Moreover, through Cytoscape and DisGeNET we could pinpoint surrogate markers either for AVS (e.g. popeye domain containing protein 2 and 28S ribosomal protein S36, mitochondrial) or for CAD (e.g. ankyrin repeat and SOCS box protein 7) which deserve future validation. Data recycling and integration as well as research orientation are among the main advantages of resorting to bioinformatics analysis, hence these tutorials can be of great convenience for proteomics investigators. As we saw for aortic valve stenosis and coronary artery disease, it can be of great relevance to perform preliminary bioinformatics analysis with already published proteomics data. It not only saves us time in the lab (avoiding work duplication) as it points out new hypothesis to explain the phenotypical presentation of the diseases as well as new surrogate markers with clinical relevance, deserving future scrutiny. These essential steps can be easily overcome if one follows the steps proposed in our tutorial for STRING, DisGeNET, Cytoscape and ClueGO utilization. Copyright © 2017 Elsevier B.V. All rights

  6. Proteome analysis of human substantia nigra in Parkinson's disease

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    Werner Cornelius J

    2008-02-01

    Full Text Available Abstract Background Parkinson's disease (PD is the most common neurodegenerative disorder involving the motor system. Although not being the only region involved in PD, affection of the substantia nigra and its projections is responsible for some of the most debilitating features of the disease. To further advance a comprehensive understanding of nigral pathology, we conducted a tissue based comparative proteome study of healthy and diseased human substantia nigra. Results The gross number of differentially regulated proteins in PD was 221. In total, we identified 37 proteins, of which 16 were differentially expressed. Identified differential proteins comprised elements of iron metabolism (H-ferritin and glutathione-related redox metabolism (GST M3, GST P1, GST O1, including novel redox proteins (SH3BGRL. Additionally, many glial or related proteins were found to be differentially regulated in PD (GFAP, GMFB, galectin-1, sorcin, as well as proteins belonging to metabolic pathways sparsely described in PD, such as adenosyl homocysteinase (methylation, aldehyde dehydrogenase 1 and cellular retinol-binding protein 1 (aldehyde metabolism. Further differentially regulated proteins included annexin V, beta-tubulin cofactor A, coactosin-like protein and V-type ATPase subunit 1. Proteins that were similarly expressed in healthy or diseased substantia nigra comprised housekeeping proteins such as COX5A, Rho GDI alpha, actin gamma 1, creatin-kinase B, lactate dehydrogenase B, disulfide isomerase ER-60, Rab GDI beta, methyl glyoxalase 1 (AGE metabolism and glutamine synthetase. Interestingly, also DJ-1 and UCH-L1 were expressed similarly. Furthermore, proteins believed to serve as internal standards were found to be expressed in a constant manner, such as 14-3-3 epsilon and hCRMP-2, thus lending further validity to our results. Conclusion Using an approach encompassing high sensitivity and high resolution, we show that alterations of SN in PD include many

  7. Quantitative proteomic analysis of human lung tumor xenografts treated with the ectopic ATP synthase inhibitor citreoviridin.

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    Yi-Hsuan Wu

    Full Text Available ATP synthase is present on the plasma membrane of several types of cancer cells. Citreoviridin, an ATP synthase inhibitor, selectively suppresses the proliferation and growth of lung cancer without affecting normal cells. However, the global effects of targeting ectopic ATP synthase in vivo have not been well defined. In this study, we performed quantitative proteomic analysis using isobaric tags for relative and absolute quantitation (iTRAQ and provided a comprehensive insight into the complicated regulation by citreoviridin in a lung cancer xenograft model. With high reproducibility of the quantitation, we obtained quantitative proteomic profiling with 2,659 proteins identified. Bioinformatics analysis of the 141 differentially expressed proteins selected by their relative abundance revealed that citreoviridin induces alterations in the expression of glucose metabolism-related enzymes in lung cancer. The up-regulation of enzymes involved in gluconeogenesis and storage of glucose indicated that citreoviridin may reduce the glycolytic intermediates for macromolecule synthesis and inhibit cell proliferation. Using comprehensive proteomics, the results identify metabolic aspects that help explain the antitumorigenic effect of citreoviridin in lung cancer, which may lead to a better understanding of the links between metabolism and tumorigenesis in cancer therapy.

  8. Data set for the proteomics analysis of the endomembrane system from the unicellular Entamoeba histolytica

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    Doranda Perdomo

    2014-12-01

    Full Text Available Entamoeba histolytica is the protozoan parasite agent of amebiasis, an infectious disease of the human intestine and liver. This parasite contact and kills human cells by an active process involving pathogenic factors. Cellular traffic and secretion activities are poorly characterized in E. histolytica. In this work, we took advantage of a wide proteomic analysis to search for principal components of the endomembrane system in E. histolytica. A total of 5683 peptides matching with 1531 proteins (FDR of 1% were identified which corresponds to roughly 20% of the total amebic proteome. Bioinformatics investigations searching for domain homologies (Smart and InterProScan programs and functional descriptions (KEGG and GO terms allowed this data to be organized into distinct categories. This data represents the first in-depth proteomics analysis of subcellular compartments in E. histolytica and allows a detailed map of vesicle traffic components in an ancient single-cell organism that lacks a stereotypical ER and Golgi apparatus to be established. The data are related to [1].

  9. Proteomic analysis and food-grade enzymes of Moringa oleifer Lam. a Lam. flower.

    Science.gov (United States)

    Shi, Yanan; Wang, Xuefeng; Huang, Aixiang

    2018-08-01

    Moringa oleifer Lam. flower contain high-proteins and function nutrients. Many advances have been made to it, but there is still no proteomic information of this species. Total protein from the flowers applied shotgun 2DLC-MS/MS proteomic identified 9443 peptides corresponding to 4004 high-confidence proteins by Proteome Discoverer™ Software 2.1. These proteins were mostly distributed ranging between 40 and 70 kDa. Gene Ontology (GO) analysis indicated that the largest of the proteins were cytoplasm 72.7%, catalytic activity 61.5% and macromolecule metabolism 43.7%, and KEGG analysis revealed that the largest group of 129 proteins was involved in Ribosome to directing protein synthesis (translation). Moreover, a number of commercially important food-grade enzymes were commented, 261 proteins were annotated as carbohydrate-active enzymes, 16 protease, 22 proteins are assigned to the citrate cycle, which the top proteins were assigned to GH family, cysteine synthase and serine/threonine-protein phosphatase. These enzymes indicated that is a new source with potential use for fermentation and brewing industry, fruit and vegetable storage and the development of function peptides. Copyright © 2018 Elsevier B.V. All rights reserved.

  10. Muscle Tissue Damage Induced by the Venom of Bothrops asper: Identification of Early and Late Pathological Events through Proteomic Analysis.

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    Cristina Herrera

    2016-04-01

    Full Text Available The time-course of the pathological effects induced by the venom of the snake Bothrops asper in muscle tissue was investigated by a combination of histology, proteomic analysis of exudates collected in the vicinity of damaged muscle, and immunodetection of extracellular matrix proteins in exudates. Proteomic assay of exudates has become an excellent new methodological tool to detect key biomarkers of tissue alterations for a more integrative perspective of snake venom-induced pathology. The time-course analysis of the intracellular proteins showed an early presence of cytosolic and mitochondrial proteins in exudates, while cytoskeletal proteins increased later on. This underscores the rapid cytotoxic effect of venom, especially in muscle fibers, due to the action of myotoxic phospholipases A2, followed by the action of proteinases in the cytoskeleton of damaged muscle fibers. Similarly, the early presence of basement membrane (BM and other extracellular matrix (ECM proteins in exudates reflects the rapid microvascular damage and hemorrhage induced by snake venom metalloproteinases. The presence of fragments of type IV collagen and perlecan one hour after envenoming suggests that hydrolysis of these mechanically/structurally-relevant BM components plays a key role in the genesis of hemorrhage. On the other hand, the increment of some ECM proteins in the exudate at later time intervals is likely a consequence of the action of endogenous matrix metalloproteinases (MMPs or of de novo synthesis of ECM proteins during tissue remodeling as part of the inflammatory reaction. Our results offer relevant insights for a more integrative and systematic understanding of the time-course dynamics of muscle tissue damage induced by B. asper venom and possibly other viperid venoms.

  11. Identification of azurocidin as a potential periodontitis biomarker by a proteomic analysis of gingival crevicular fluid

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    Lee Jae-Mok

    2011-07-01

    Full Text Available Abstract Background The inflammatory disease periodontitis results in tooth loss and can even lead to diseases of the whole body if not treated. Gingival crevicular fluid (GCF reflects the condition of the gingiva and contains proteins transuded from serum or cells at inflamed sites. In this study, we aimed to discover potential protein biomarkers for periodontitis in GCF proteome using LC-MS/MS. Results We identified 305 proteins from GCF of healthy individuals and periodontitis patients collected using a sterile gel loading tip by ESI-MS/MS coupled to nano-LC. Among these proteins, about 45 proteins were differentially expressed in the GCF proteome of moderate periodontitis patients when compared to the healthy individuals. We first identified azurocidin in the GCF, but not the saliva, as an upregulated protein in the periodontitis patients and verified its increased expression during periodontitis by ELISA using the GCF of the classified periodontitis patients compared to the healthy individuals. In addition, we found that azurocidin inhibited the differentiation of bone marrow-derived macrophages to osteoclasts. Conclusions Our results show that GCF collection using a gel loading tip and subsequent LC-MS/MS analysis following 1D-PAGE proteomic separation are effective for the analysis of the GCF proteome. Our current results also suggest that azurocidin could be a potential biomarker candidate for the early detection of inflammatory periodontal destruction by gingivitis and some chronic periodontitis. Our data also suggest that azurocidin may have an inhibitory role in osteoclast differentiation and, thus, a protective role in alveolar bone loss during the early stages of periodontitis.

  12. Proteomic Analysis Shows Constitutive Secretion of MIF and p53-associated Activity of COX-2−/− Lung Fibroblasts

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    Mandar Dave

    2017-12-01

    Full Text Available The differential expression of two closelyassociated cyclooxygenase isozymes, COX-1 and COX-2, exhibited functions beyond eicosanoid metabolism. We hypothesized that COX-1 or COX-2 knockout lung fibroblasts may display altered protein profiles which may allow us to further differentiate the functional roles of these isozymes at the molecular level. Proteomic analysis shows constitutive production of macrophage migration inhibitory factor (MIF in lung fibroblasts derived from COX-2−/− but not wild-type (WT or COX-1−/− mice. MIF was spontaneously released in high levels into the extracellular milieu of COX2−/− fibroblasts seemingly from the preformed intracellular stores, with no change in the basal gene expression of MIF. The secretion and regulation of MIF in COX-2−/− was “prostaglandin-independent.” GO analysis showed that concurrent with upregulation of MIF, there is a significant surge in expression of genes related to fibroblast growth, FK506 binding proteins, and isomerase activity in COX-2−/− cells. Furthermore, COX-2−/− fibroblasts also exhibit a significant increase in transcriptional activity of various regulators, antagonists, and co-modulators of p53, as well as in the expression of oncogenes and related transcripts. Integrative Oncogenomics Cancer Browser (IntroGen analysis shows downregulation of COX-2 and amplification of MIF and/or p53 activity during development of glioblastomas, ependymoma, and colon adenomas. These data indicate the functional role of the MIF-COX-p53 axis in inflammation and cancer at the genomic and proteomic levels in COX-2-ablated cells. This systematic analysis not only shows the proinflammatory state but also unveils a molecular signature of a pro-oncogenic state of COX-1 in COX-2 ablated cells.

  13. Proteomic analysis of the metabolic adaptation of the biocontrol agent Pseudozyma flocculosa leading to glycolipid production

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    Bélanger Richard R

    2010-02-01

    Full Text Available Abstract The yeast-like epiphytic fungus Pseudozyma flocculosa is known to antagonize powdery mildew fungi through proliferation on colonies presumably preceded by the release of an antifungal glycolipid (flocculosin. In culture conditions, P. flocculosa can be induced to produce or not flocculosin through manipulation of the culture medium nutrients. In order to characterize and understand the metabolic changes in P. flocculosa linked to glycolipid production, we conducted a 2-DE proteomic analysis and compared the proteomic profile of P. flocculosa growing under conditions favoring the development of the fungus (control or conducive to flocculosin synthesis (stress. A large number of protein spots (771 were detected in protein extracts of the control treatment compared to only 435 matched protein spots in extracts of the stress cultures, which clearly suggests an important metabolic reorganization in slow-growing cells producing flocculosin. From the latter treatment, we were able to identify 21 protein spots that were either specific to the treatment or up-regulated significantly (2-fold increase. All of them were identified based on similarity between predicted ORF of the newly sequenced genome of P. flocculosa with Ustilago maydis' available annotated sequences. These proteins were associated with the carbon and fatty acid metabolism, and also with the filamentous change of the fungus leading to flocculosin production. This first look into the proteome of P. flocculosa suggests that flocculosin synthesis is elicited in response to specific stress or limiting conditions.

  14. In-depth analysis of the chicken egg white proteome using an LTQ Orbitrap Velos

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    Mann Matthias

    2011-02-01

    Full Text Available Abstract Background Hen's egg white has been the subject of intensive chemical, biochemical and food technological research for many decades, because of its importance in human nutrition, its importance as a source of easily accessible model proteins, and its potential use in biotechnological processes. Recently the arsenal of tools used to study the protein components of egg white has been complemented by mass spectrometry-based proteomic technologies. Application of these fast and sensitive methods has already enabled the identification of a large number of new egg white proteins. Recent technological advances may be expected to further expand the egg white protein inventory. Results Using a dual pressure linear ion trap Orbitrap instrument, the LTQ Orbitrap Velos, in conjunction with data analysis in the MaxQuant software package, we identified 158 proteins in chicken egg white with two or more sequence unique peptides. This group of proteins identified with very high confidence included 79 proteins identified in egg white for the first time. In addition, 44 proteins were identified tentatively. Conclusions Our results, apart from identifying many new egg white components, indicate that current mass spectrometry technology is sufficiently advanced to permit direct identification of minor components of proteomes dominated by a few major proteins without resorting to indirect techniques, such as chromatographic depletion or peptide library binding, which change the composition of the proteome.

  15. Shotgun proteomic analysis of Emiliania huxleyi, a marine phytoplankton species of major biogeochemical importance.

    Science.gov (United States)

    Jones, Bethan M; Edwards, Richard J; Skipp, Paul J; O'Connor, C David; Iglesias-Rodriguez, M Debora

    2011-06-01

    Emiliania huxleyi is a unicellular marine phytoplankton species known to play a significant role in global biogeochemistry. Through the dual roles of photosynthesis and production of calcium carbonate (calcification), carbon is transferred from the atmosphere to ocean sediments. Almost nothing is known about the molecular mechanisms that control calcification, a process that is tightly regulated within the cell. To initiate proteomic studies on this important and phylogenetically remote organism, we have devised efficient protein extraction protocols and developed a bioinformatics pipeline that allows the statistically robust assignment of proteins from MS/MS data using preexisting EST sequences. The bioinformatics tool, termed BUDAPEST (Bioinformatics Utility for Data Analysis of Proteomics using ESTs), is fully automated and was used to search against data generated from three strains. BUDAPEST increased the number of identifications over standard protein database searches from 37 to 99 proteins when data were amalgamated. Proteins involved in diverse cellular processes were uncovered. For example, experimental evidence was obtained for a novel type I polyketide synthase and for various photosystem components. The proteomic and bioinformatic approaches developed in this study are of wider applicability, particularly to the oceanographic community where genomic sequence data for species of interest are currently scarce.

  16. A Hybrid Feature Subset Selection Algorithm for Analysis of High Correlation Proteomic Data

    Science.gov (United States)

    Kordy, Hussain Montazery; Baygi, Mohammad Hossein Miran; Moradi, Mohammad Hassan

    2012-01-01

    Pathological changes within an organ can be reflected as proteomic patterns in biological fluids such as plasma, serum, and urine. The surface-enhanced laser desorption and ionization time-of-flight mass spectrometry (SELDI-TOF MS) has been used to generate proteomic profiles from biological fluids. Mass spectrometry yields redundant noisy data that the most data points are irrelevant features for differentiating between cancer and normal cases. In this paper, we have proposed a hybrid feature subset selection algorithm based on maximum-discrimination and minimum-correlation coupled with peak scoring criteria. Our algorithm has been applied to two independent SELDI-TOF MS datasets of ovarian cancer obtained from the NCI-FDA clinical proteomics databank. The proposed algorithm has used to extract a set of proteins as potential biomarkers in each dataset. We applied the linear discriminate analysis to identify the important biomarkers. The selected biomarkers have been able to successfully diagnose the ovarian cancer patients from the noncancer control group with an accuracy of 100%, a sensitivity of 100%, and a specificity of 100% in the two datasets. The hybrid algorithm has the advantage that increases reproducibility of selected biomarkers and able to find a small set of proteins with high discrimination power. PMID:23717808

  17. Paving a Path to Understanding Metabolic Responses to Iron Bioavailability: Global Proteomic Analysis of Crocosphaera watsonii

    Science.gov (United States)

    Gauglitz, J.; McIlvin, M. R.; Moran, D. M.; Waterbury, J. B.; Saito, M. A.

    2016-02-01

    Marine diazotrophic cyanobacteria provide a key source of new nitrogen into the oceans and are important contributors to primary production. The geographic distribution of these cyanobacteria is impacted by available iron and phosphorus as well as environmental conditions such as temperature, however available iron concentrations are thought to be particularly critical due to the high demand for iron in cellular processes. Iron bioavailability and microorganismal adaptations to low iron environments may thus play a key role in dictating community structure, however the mechanisms by which cyanobacteria acquire iron and regulate its uptake are not well defined. In this study, the unicellular diazotroph, Crocosphaera watsonii WH8501, was acclimated to a range of bioavailable iron concentrations (from 0.001nM to 8.13nM Fe') using trace metal clean culturing techniques and the proteomes were analyzed by LC/MS-MS. Physiological and proteomic data indicate three distinct phenotypic ranges: iron-replete, iron-limited, and iron-starved. Trends in photosynthetic, carbon fixation and iron storage proteins across the iron gradient indicate that the C. watsonii proteome responds directly to iron availability. Further analysis of relative protein expression, which describes the physiological state of the cell, will lead to insights into how C. watsonii is able to adapt to iron-limited conditions and the resulting biogeochemical implications will be discussed.

  18. Quantitative proteomic analysis of microdissected oral epithelium for cancer biomarker discovery.

    Science.gov (United States)

    Xiao, Hua; Langerman, Alexander; Zhang, Yan; Khalid, Omar; Hu, Shen; Cao, Cheng-Xi; Lingen, Mark W; Wong, David T W

    2015-11-01

    Specific biomarkers are urgently needed for the detection and progression of oral cancer. The objective of this study was to discover cancer biomarkers from oral epithelium through utilizing high throughput quantitative proteomics approaches. Morphologically malignant, epithelial dysplasia, and adjacent normal epithelial tissues were laser capture microdissected (LCM) from 19 patients and used for proteomics analysis. Total proteins from each group were extracted, digested and then labelled with corresponding isobaric tags for relative and absolute quantitation (iTRAQ). Labelled peptides from each sample were combined and analyzed by liquid chromatography-mass spectrometry (LC-MS/MS) for protein identification and quantification. In total, 500 proteins were identified and 425 of them were quantified. When compared with adjacent normal oral epithelium, 17 and 15 proteins were consistently up-regulated or down-regulated in malignant and epithelial dysplasia, respectively. Half of these candidate biomarkers were discovered for oral cancer for the first time. Cornulin was initially confirmed in tissue protein extracts and was further validated in tissue microarray. Its presence in the saliva of oral cancer patients was also explored. Myoglobin and S100A8 were pre-validated by tissue microarray. These data demonstrated that the proteomic biomarkers discovered through this strategy are potential targets for oral cancer detection and salivary diagnostics. Copyright © 2015 Elsevier Ltd. All rights reserved.

  19. Comparative proteomic analysis of differentially expressed proteins in shoots of Salicornia europaea under different salinity.

    Science.gov (United States)

    Wang, Xuchu; Fan, Pengxiang; Song, Hongmiao; Chen, Xianyang; Li, Xiaofang; Li, Yinxin

    2009-07-01

    Soil salinity is a major abiotic stress that limits agriculture productivity worldwide. Salicornia europaea is a succulent annual euhalophyte and one of the most salt tolerant plant species. The elucidation of its salt tolerance mechanism is of significance for generating salt-tolerant crops. In this study, we provided high resolution of proteome reference maps of S. europaea shoot and obtained evidence on the salt tolerance mechanism by analyzing the proteomic responses of this plant to high salinity. Our results demonstrated significant variations existed in 196 out of 1880 protein spots detected on CBB stained 2-DE gels. Of these, 111 proteins were identified by mass spectrometry. Among them, the majority was energy production and conversion related proteins, followed by photosynthesis and carbohydrate metabolism associated enzymes. Analysis of protein expression patters revealed that energy production and ion homeostasis associated proteins played important roles for this plant salt tolerance ability. Hierarchical clustering results revealed many proteins were involved in S. europaea salt tolerance mechanism as a dynamic network. Finally, based on our proteomic results, we brought forward a possible schematic representation of mechanism associated with the systematic salt tolerance phenotype in S. europaea.

  20. Novel proteins from proteomic analysis of the trunk disease fungus Lasiodiplodia theobromae (Botryosphaeriaceae

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    Carla C. Uranga

    2017-06-01

    Full Text Available Many basic science questions remain regarding protein functions in the pathogen: host interaction, especially in the trunk disease fungi family, the Botryosphaeriaceae, which are a global problem for economically important plants, especially fruiting trees. Proteomics is a highly useful technology for studying protein expression and for discovering novel proteins in unsequenced and poorly annotated organisms. Current fungal proteomics approaches involve 2D SDS-PAGE and extensive, complex, protein extraction methodologies. In this work, a modified Folch extraction was applied to protein extraction to perform both de novo peptide sequencing and peptide fragmentation analysis/protein identification of the plant and human fungal pathogen Lasiodiplodia theobromae. Both bioinformatics approaches yielded novel peptide sequences from proteins produced by L. theobromae in the presence of exogenous triglycerides and glucose. These proteins and the functions they may possess could be targeted for further functional characterization and validation efforts, due to their potential uses in biotechnology and as new paradigms for understanding fungal biochemistry, such as the finding of allergenic enolases, as well as various novel proteases, including zinc metalloproteinases homologous to those found in snake venom. This work contributes to genomic annotation efforts, which, hand in hand with genomic sequencing, will help improve fungal bioinformatics databases for future studies of Botryosphaeriaceae. All data, including raw data, are available via the ProteomeXchange data repository with identifier PXD005283. This is the first study of its kind in Botryosphaeriaceae.

  1. Comparative proteomics analysis of sheep sperm under two doses of heavy ion to irradiation

    International Nuclear Information System (INIS)

    Li Hongyan; Zhao Xingxu; He Yuxuan; Zhang Yong; Zhang Hong; Wang Yanling; Li Fadi; Ma Youji

    2011-01-01

    The object of this study was to investigate differential proteomic expressions in sheep sperm protein under two doses (0.5 and 0.3 kGy) heavy ion radiation. The current research presented the protein changes using two-dimensional gel electrophoresis (2-DE) after staining with silver nitrate, differential expression proteins were detected by PDQuest 8.0 software and subjected to ion trap mass spectrometer equipped with a Surveyor HPLC system, and differential spots of protein were identified. Results showed that eight common different expressed protein spots in two doses 2D gels were identified to be three up-regulated proteins (glutaredoxin -1, transcription factor AP -2-alpha and enolase). It was concluded that there was significant difference at protein level in sheep sperm after heavy ion radiation and differential proteome expression analysis may be useful to clarify the physiology state of sheep sperm in heavy ion radiation, which laid a foundation for the further studies on heavy ion radiation of sheep sperm proteomics. (authors)

  2. Comparison of Different Protein Extraction Methods for Gel-Based Proteomic Analysis of Ganoderma spp.

    Science.gov (United States)

    Al-Obaidi, Jameel R; Saidi, Noor Baity; Usuldin, Siti Rokhiyah Ahmad; Hussin, Siti Nahdatul Isnaini Said; Yusoff, Noornabeela Md; Idris, Abu Seman

    2016-04-01

    Ganoderma species are a group of fungi that have the ability to degrade lignin polymers and cause severe diseases such as stem and root rot and can infect economically important plants and perennial crops such as oil palm, especially in tropical countries such as Malaysia. Unfortunately, very little is known about the complex interplay between oil palm and Ganoderma in the pathogenesis of the diseases. Proteomic technologies are simple yet powerful tools in comparing protein profile and have been widely used to study plant-fungus interaction. A critical step to perform a good proteome research is to establish a method that gives the best quality and a wide coverage of total proteins. Despite the availability of various protein extraction protocols from pathogenic fungi in the literature, no single extraction method was found suitable for all types of pathogenic fungi. To develop an optimized protein extraction protocol for 2-DE gel analysis of Ganoderma spp., three previously reported protein extraction protocols were compared: trichloroacetic acid, sucrose and phenol/ammonium acetate in methanol. The third method was found to give the most reproducible gels and highest protein concentration. Using the later method, a total of 10 protein spots (5 from each species) were successfully identified. Hence, the results from this study propose phenol/ammonium acetate in methanol as the most effective protein extraction method for 2-DE proteomic studies of Ganoderma spp.

  3. Application of proteomics to ecology and population biology.

    Science.gov (United States)

    Karr, T L

    2008-02-01

    Proteomics is a relatively new scientific discipline that merges protein biochemistry, genome biology and bioinformatics to determine the spatial and temporal expression of proteins in cells, tissues and whole organisms. There has been very little application of proteomics to the fields of behavioral genetics, evolution, ecology and population dynamics, and has only recently been effectively applied to the closely allied fields of molecular evolution and genetics. However, there exists considerable potential for proteomics to impact in areas related to functional ecology; this review will introduce the general concepts and methodologies that define the field of proteomics and compare and contrast the advantages and disadvantages with other methods. Examples of how proteomics can aid, complement and indeed extend the study of functional ecology will be discussed including the main tool of ecological studies, population genetics with an emphasis on metapopulation structure analysis. Because proteomic analyses provide a direct measure of gene expression, it obviates some of the limitations associated with other genomic approaches, such as microarray and EST analyses. Likewise, in conjunction with associated bioinformatics and molecular evolutionary tools, proteomics can provide the foundation of a systems-level integration approach that can enhance ecological studies. It can be envisioned that proteomics will provide important new information on issues specific to metapopulation biology and adaptive processes in nature. A specific example of the application of proteomics to sperm ageing is provided to illustrate the potential utility of the approach.

  4. Proteomics analysis of the DF-1 chicken fibroblasts infected with avian reovirus strain S1133.

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    Wen-Ting Chen

    Full Text Available BACKGROUND: Avian reovirus (ARV is a member of the Orthoreovirus genus in the Reoviridae family. It is the etiological agent of several diseases, among which viral arthritis and malabsorption syndrome are the most commercially important, causing considerable economic losses in the poultry industry. Although a small but increasing number of reports have characterized some aspects of ARV infection, global changes in protein expression in ARV-infected host cells have not been examined. The current study used a proteomics approach to obtain a comprehensive view of changes in protein levels in host cells upon infection by ARV. METHODOLOGY AND PRINCIPAL FINDINGS: The proteomics profiles of DF-1 chicken fibroblast cells infected with ARV strain S1133 were analyzed by two-dimensional differential-image gel electrophoresis. The majority of protein expression changes (≥ 1.5 fold, p<0.05 occurred at 72 h post-infection. Matrix-assisted laser desorption ionization time-of-flight mass spectrometry identified 51 proteins with differential expression levels, including 25 that were upregulated during ARV infection and 26 that were downregulated. These proteins were divided into eight groups according to biological function: signal transduction, stress response, RNA processing, the ubiquitin-proteasome pathway, lipid metabolism, carbohydrate metabolism, energy metabolism, and cytoskeleton organization. They were further examined by immunoblotting to validate the observed alterations in protein expression. CONCLUSION/SIGNIFICANCE: This is the first report of a time-course proteomic analysis of ARV-infected host cells. Notably, all identified proteins involved in signal transduction, RNA processing, and the ubiquitin-proteasome pathway were downregulated in infected cells, whereas proteins involved in DNA synthesis, apoptosis, and energy production pathways were upregulated. In addition, other differentially expressed proteins were linked with the cytoskeleton

  5. Transcriptomic and proteomic analysis of Oenococcus oeni adaptation to wine stress conditions

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    Mar Margalef-Català

    2016-09-01

    Full Text Available Oenococcus oeni, the main lactic acid bacteria responsible for malolactic fermentation in wine, has to adapt to stressful conditions, such as low pH and high ethanol content. In this study, the changes in the transcriptome and the proteome of O. oeni PSU-1 during the adaptation period before MLF start have been studied. DNA microarrays were used for the transcriptomic analysis and two complementary proteomic techniques, 2-D DIGE and iTRAQ labeling were used to analyze the proteomic response. One of the most influenced functions in PSU-1 due to inoculation into wine-like medium (WLM was translation, showing the over-expression of certain ribosomal genes and the corresponding proteins. Amino acid metabolism and transport was also altered and several peptidases were up regulated both at gene and protein level. Certain proteins involved in glutamine and glutamate metabolism showed an increased abundance revealing the key role of nitrogen uptake under stressful conditions. A strong transcriptional inhibition of carbohydrate metabolism related genes was observed. On the other hand, the transcriptional up-regulation of malate transport and citrate consumption was indicative of the use of L-malate and citrate associated to stress response and as an alternative energy source to sugar metabolism. Regarding the stress mechanisms, our results support the relevance of the thioredoxin and glutathione systems in the adaptation of O. oeni to wine related stress. Genes and proteins related to cell wall showed also significant changes indicating the relevance of the cell envelop as protective barrier to environmental stress. The differences found between transcriptomic and proteomic data suggested the relevance of post-transcriptional mechanisms and the complexity of the stress response in O. oeni adaptation. Further research should deepen into the metabolisms mostly altered due to wine conditions to elucidate the role of each mechanism in the O. oeni ability to

  6. Proteomic analysis of Bombyx mori molting fluid: Insights into the molting process.

    Science.gov (United States)

    Liu, Hua-Wei; Wang, Luo-Ling; Tang, Xin; Dong, Zhao-Ming; Guo, Peng-Chao; Zhao, Dong-Chao; Xia, Qing-You; Zhao, Ping

    2018-02-20

    Molting is an essential biological process occurring multiple times throughout the life cycle of most Ecdysozoa. Molting fluids accumulate and function in the exuvial space during the molting process. In this study, we used liquid chromatography-tandem mass spectrometry to investigate the molting fluids to analyze the molecular mechanisms of molting in the silkworm, Bombyx mori. In total, 375 proteins were identified in molting fluids from the silkworm at 14-16h before pupation and eclosion, including 12 chitin metabolism-related enzymes, 35 serine proteases, 15 peptidases, and 38 protease inhibitors. Gene ontology analysis indicated that "catalytic" constitutes the most enriched function in the molting fluid. Gene expression patterns and bioinformatic analyses suggested that numerous enzymes are involved in the degradation of cuticle proteins and chitin. Protein-protein interaction network and activity analyses showed that protease inhibitors are involved in the regulation of multiple pathways in molting fluid. Additionally, many immune-related proteins may be involved in the immune defense during molting. These results provide a comprehensive proteomic insight into proteolytic enzymes and protease inhibitors in molting fluid, and will likely improve the current understanding of physiological processes in insect molting. Insect molting constitutes a dynamic physiological process. To better understand this process, we used LC-MS/MS to investigate the proteome of silkworm molting fluids and identified key proteins involved in silkworm molting. The biological processes of the old cuticle degradation pathway and immune defense response were analyzed in the proteome of silkworm molting fluid. We report that protease inhibitors serve as key factors in the regulation of the molting process. The proteomic results provide new insight into biological molting processes in insects. Copyright © 2017 Elsevier B.V. All rights reserved.

  7. Comparative proteomic analysis of human malignant ascitic fluids for the development of gastric cancer biomarkers.

    Science.gov (United States)

    Jin, Jonghwa; Son, Minsoo; Kim, Hyeyoon; Kim, Hyeyeon; Kong, Seong-Ho; Kim, Hark Kyun; Kim, Youngsoo; Han, Dohyun

    2018-04-11

    Malignant ascites is a sign of peritoneal seeding, which is one of the most frequent forms of incurable distant metastasis. Because the development of malignant ascites is associated with an extremely poor prognosis, determining whether it resulted from peritoneal seeding has critical clinical implications in diagnosis, choice of treatment, and active surveillance. At present, the molecular characterizations of malignant ascites are especially limited in case of gastric cancer. We aimed to identify malignant ascites-specific proteins that may contribute to the development of alternative methods for diagnosis and therapeutic monitoring and also increase our understanding of the pathophysiology of peritoneal seeding. First, comprehensive proteomic strategies were employed to construct an in-depth proteome of ascitic fluids. Label-free quantitative proteomic analysis was subsequently performed to identify candidates that can differentiate between malignant ascitic fluilds of gastric cancer patients from benign ascitic fluids. Finally, two candidate proteins were verified by ELISA in 84 samples with gastric cancer or liver cirrhosis. Comprehensive proteome profiling resulted in the identification of 5347 ascites proteins. Using label-free quantification, we identified 299 proteins that were differentially expressed in ascitic fluids between liver cirrhosis and stage IV gastric cancer patients. In addition, we identified 645 proteins that were significantly expressed in ascitic fluids between liver cirrhosis and gastric cancer patients with peritoneal seeding. Finally, Gastriscin and Periostin that can distinguish malignant ascites from benign ascites were verified by ELISA. This study identified and verified protein markers that can distinguish malignant ascites with or without peritoneal seeding from benign ascites. Consequently, our results could be a significant resource for gastric cancer research and biomarker discovery in the diagnosis of malignant ascites

  8. Using a novel "Integrated Biomarker Proteomic" index to assess the effects of freshwater pollutants in European eel peripheral blood mononuclear cells.

    Science.gov (United States)

    Roland, Kathleen; Kestemont, Patrick; Dieu, Marc; Raes, Martine; Silvestre, Frédéric

    2016-03-30

    Using proteomic data as biomarkers of environmental pollution has the potential to be of a great interest in ecological risk assessment as they constitute early warning indicators of ecologically relevant effects on biological systems. To develop such specific and sensitive biomarkers, the use of a set of proteins is required and the identification of protein expression signatures (PES) may reflect the exposure to specific classes of pollutants. Using 2D-DIGE (Differential in Gel Electrophoresis) methodology, this study aimed at identifying specific PES on European eel (Anguilla anguilla) peripheral blood mononuclear cells (PBMC) after 48 h in vitro exposure to two sublethal concentrations of dichlorodiphenyltrichloroethane (p,p'-DDT) (10 μg/L and 1mg/L) or cadmium (Cd) (1 μg/L and 100 μg/L). The present results have been supplemented with data of a first in vitro study on cells exposed to perfluorooctane sulfonate (PFOS) (10 μg/L and 1mg/L). A total of thirty-four protein spots, belonging to 18 different identified proteins found in all conditions, have been selected as possible biomarkers to develop a synthetic Integrated Biomarker Proteomic (IBP) index. IBP follows a dose-response relationship with higher values at the highest tested concentration for each pollutant (Cd: 9.96; DDT: 7.44; PFOS: 7.94) compared to the lowest tested concentration (Cd: 3.81; DDT: 2.91; PFOS: 2.06). In a second step, star plot graphs have been applied to proteomic data in order to allow visual integration of a set of early warning responses measured with protein biomarkers. Such star plots permit to discriminate the type of pollutant inducing a proteomic response. We conclude that using IBP is relevant in environmental risk assessment, giving to this index the potential to be applied as a global index of proteome alteration in endangered species such as the European eel. In this study, 34 protein spots have been selected as possible biomarkers to develop a synthetic Integrated

  9. Challenges and Strategies for Proteome Analysis of the Interaction of Human Pathogenic Fungi with Host Immune Cells.

    Science.gov (United States)

    Krüger, Thomas; Luo, Ting; Schmidt, Hella; Shopova, Iordana; Kniemeyer, Olaf

    2015-12-14

    Opportunistic human pathogenic fungi including the saprotrophic mold Aspergillus fumigatus and the human commensal Candida albicans can cause severe fungal infections in immunocompromised or critically ill patients. The first line of defense against opportunistic fungal pathogens is the innate immune system. Phagocytes such as macrophages, neutrophils and dendritic cells are an important pillar of the innate immune response and have evolved versatile defense strategies against microbial pathogens. On the other hand, human-pathogenic fungi have sophisticated virulence strategies to counteract the innate immune defense. In this context, proteomic approaches can provide deeper insights into the molecular mechanisms of the interaction of host immune cells with fungal pathogens. This is crucial for the identification of both diagnostic biomarkers for fungal infections and therapeutic targets. Studying host-fungal interactions at the protein level is a challenging endeavor, yet there are few studies that have been undertaken. This review draws attention to proteomic techniques and their application to fungal pathogens and to challenges, difficulties, and limitations that may arise in the course of simultaneous dual proteome analysis of host immune cells interacting with diverse morphotypes of fungal pathogens. On this basis, we discuss strategies to overcome these multifaceted experimental and analytical challenges including the viability of immune cells during co-cultivation, the increased and heterogeneous protein complexity of the host proteome dynamically interacting with the fungal proteome, and the demands on normalization strategies in terms of relative quantitative proteome analysis.

  10. [Methods of quantitative proteomics].

    Science.gov (United States)

    Kopylov, A T; Zgoda, V G

    2007-01-01

    In modern science proteomic analysis is inseparable from other fields of systemic biology. Possessing huge resources quantitative proteomics operates colossal information on molecular mechanisms of life. Advances in proteomics help researchers to solve complex problems of cell signaling, posttranslational modification, structure and functional homology of proteins, molecular diagnostics etc. More than 40 various methods have been developed in proteomics for quantitative analysis of proteins. Although each method is unique and has certain advantages and disadvantages all these use various isotope labels (tags). In this review we will consider the most popular and effective methods employing both chemical modifications of proteins and also metabolic and enzymatic methods of isotope labeling.

  11. Proteomic analysis of the seed development in Jatropha curcas: from carbon flux to the lipid accumulation.

    Science.gov (United States)

    Liu, Hui; Wang, Cuiping; Komatsu, Setsuko; He, Mingxia; Liu, Gongshe; Shen, Shihua

    2013-10-08

    To characterize the metabolic signatures of lipid accumulation in Jatropha curcas seeds, comparative proteomic technique was employed to profile protein changes during the seed development. Temporal changes in comparative proteome were examined using gels-based proteomic technique at six developmental stages for lipid accumulation. And 104 differentially expressed proteins were identified by MALDI-TOF/TOF tandem mass spectrometry. These protein species were classified into 10 functional categories, and the results demonstrated that protein species related to energy and metabolism were notably accumulated and involved in the carbon flux to lipid accumulation that occurs primarily from early to late stage in seed development. Glycolysis and oxidative pentose phosphate pathways were the major pathways of producing carbon flux, and the glucose-6-phosphate and triose-phosphate are the major carbon source for fatty acid synthesis. Lipid analysis revealed that fatty acid accumulation initiated 25days after flowering at the late stage of seed development of J. curcas. Furthermore, C16:0 was initially synthesized as the precursor for the elongation to C18:1 and C18:2 in the developing seeds of J. curcas. Together, the metabolic signatures on protein changes in seed development provide profound knowledge and perspective insights into understanding lipid network in J. curcas. Due to the abundant oil content in seeds, Jatropha curcas seeds are being considered as the ideal materials for biodiesel. Although several studies had carried out the transcriptomic project to study the genes expression profiles in seed development of J. curcas, these ESTs hadn't been confirmed by qRT-PCR. Yet, the seed development of J. curcas had been described for a pool of developing seeds instead of being characterized systematically. Moreover, cellular metabolic events are also controlled by protein-protein interactions, posttranslational protein modifications, and enzymatic activities which

  12. Growth Phase-Dependent Proteomes of the Malaysian Isolated Lactococcus lactis Dairy Strain M4 Using Label-Free Qualitative Shotgun Proteomics Analysis

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    Theresa Wan Chen Yap

    2014-01-01

    Full Text Available Lactococcus lactis is the most studied mesophilic fermentative lactic acid bacterium. It is used extensively in the food industry and plays a pivotal role as a cell factory and also as vaccine delivery platforms. The proteome of the Malaysian isolated L. lactis M4 dairy strain, obtained from the milk of locally bred cows, was studied to elucidate the physiological changes occurring between the growth phases of this bacterium. In this study, ultraperformance liquid chromatography nanoflow electrospray ionization tandem mass spectrometry (UPLC- nano-ESI-MSE approach was used for qualitative proteomic analysis. A total of 100 and 121 proteins were identified from the midexponential and early stationary growth phases, respectively, of the L. lactis strain M4. During the exponential phase, the most important reaction was the generation of sufficient energy, whereas, in the early stationary phase, the metabolic energy pathways decreased and the biosynthesis of proteins became more important. Thus, the metabolism of the cells shifted from energy production in the exponential phase to the synthesis of macromolecules in the stationary phase. The resultant proteomes are essential in providing an improved view of the cellular machinery of L. lactis during the transition of growth phases and hence provide insight into various biotechnological applications.

  13. Growth phase-dependent proteomes of the Malaysian isolated Lactococcus lactis dairy strain M4 using label-free qualitative shotgun proteomics analysis.

    Science.gov (United States)

    Yap, Theresa Wan Chen; Rabu, Amir; Abu Bakar, Farah Diba; Rahim, Raha Abdul; Mahadi, Nor Muhammad; Illias, Rosli Md; Murad, Abdul Munir Abdul

    2014-01-01

    Lactococcus lactis is the most studied mesophilic fermentative lactic acid bacterium. It is used extensively in the food industry and plays a pivotal role as a cell factory and also as vaccine delivery platforms. The proteome of the Malaysian isolated L. lactis M4 dairy strain, obtained from the milk of locally bred cows, was studied to elucidate the physiological changes occurring between the growth phases of this bacterium. In this study, ultraperformance liquid chromatography nanoflow electrospray ionization tandem mass spectrometry (UPLC- nano-ESI-MS(E)) approach was used for qualitative proteomic analysis. A total of 100 and 121 proteins were identified from the midexponential and early stationary growth phases, respectively, of the L. lactis strain M4. During the exponential phase, the most important reaction was the generation of sufficient energy, whereas, in the early stationary phase, the metabolic energy pathways decreased and the biosynthesis of proteins became more important. Thus, the metabolism of the cells shifted from energy production in the exponential phase to the synthesis of macromolecules in the stationary phase. The resultant proteomes are essential in providing an improved view of the cellular machinery of L. lactis during the transition of growth phases and hence provide insight into various biotechnological applications.

  14. Growth Phase-Dependent Proteomes of the Malaysian Isolated Lactococcus lactis Dairy Strain M4 Using Label-Free Qualitative Shotgun Proteomics Analysis

    Science.gov (United States)

    Yap, Theresa Wan Chen; Rabu, Amir; Abu Bakar, Farah Diba; Abdul Rahim, Raha; Mahadi, Nor Muhammad; Illias, Rosli Md.

    2014-01-01

    Lactococcus lactis is the most studied mesophilic fermentative lactic acid bacterium. It is used extensively in the food industry and plays a pivotal role as a cell factory and also as vaccine delivery platforms. The proteome of the Malaysian isolated L. lactis M4 dairy strain, obtained from the milk of locally bred cows, was studied to elucidate the physiological changes occurring between the growth phases of this bacterium. In this study, ultraperformance liquid chromatography nanoflow electrospray ionization tandem mass spectrometry (UPLC- nano-ESI-MSE) approach was used for qualitative proteomic analysis. A total of 100 and 121 proteins were identified from the midexponential and early stationary growth phases, respectively, of the L. lactis strain M4. During the exponential phase, the most important reaction was the generation of sufficient energy, whereas, in the early stationary phase, the metabolic energy pathways decreased and the biosynthesis of proteins became more important. Thus, the metabolism of the cells shifted from energy production in the exponential phase to the synthesis of macromolecules in the stationary phase. The resultant proteomes are essential in providing an improved view of the cellular machinery of L. lactis during the transition of growth phases and hence provide insight into various biotechnological applications. PMID:24982972

  15. Proteome analysis of human colorectal cancer tissue using 2-D ...

    African Journals Online (AJOL)

    Jane

    2010-10-11

    Oct 11, 2010 ... protein spots were identified by mass spectrometric analysis. The cDNA of the ..... sensitivity, dynamic range and reproducibility vs the conventional 2-D ... linkage, and also has molecular chaperones activity for inhibiting the ...

  16. Integrative microRNA and proteomic approaches identify novel osteoarthritis genes and their collaborative metabolic and inflammatory networks.

    Directory of Open Access Journals (Sweden)

    Dimitrios Iliopoulos

    Full Text Available BACKGROUND: Osteoarthritis is a multifactorial disease characterized by destruction of the articular cartilage due to genetic, mechanical and environmental components affecting more than 100 million individuals all over the world. Despite the high prevalence of the disease, the absence of large-scale molecular studies limits our ability to understand the molecular pathobiology of osteoathritis and identify targets for drug development. METHODOLOGY/PRINCIPAL FINDINGS: In this study we integrated genetic, bioinformatic and proteomic approaches in order to identify new genes and their collaborative networks involved in osteoarthritis pathogenesis. MicroRNA profiling of patient-derived osteoarthritic cartilage in comparison to normal cartilage, revealed a 16 microRNA osteoarthritis gene signature. Using reverse-phase protein arrays in the same tissues we detected 76 differentially expressed proteins between osteoarthritic and normal chondrocytes. Proteins such as SOX11, FGF23, KLF6, WWOX and GDF15 not implicated previously in the genesis of osteoarthritis were identified. Integration of microRNA and proteomic data with microRNA gene-target prediction algorithms, generated a potential "interactome" network consisting of 11 microRNAs and 58 proteins linked by 414 potential functional associations. Comparison of the molecular and clinical data, revealed specific microRNAs (miR-22, miR-103 and proteins (PPARA, BMP7, IL1B to be highly correlated with Body Mass Index (BMI. Experimental validation revealed that miR-22 regulated PPARA and BMP7 expression and its inhibition blocked inflammatory and catabolic changes in osteoarthritic chondrocytes. CONCLUSIONS/SIGNIFICANCE: Our findings indicate that obesity and inflammation are related to osteoarthritis, a metabolic disease affected by microRNA deregulation. Gene network approaches provide new insights for elucidating the complexity of diseases such as osteoarthritis. The integration of microRNA, proteomic

  17. Proteomic and comparative genomic analysis reveals adaptability of Brassica napus to phosphorus-deficient stress.

    Science.gov (United States)

    Chen, Shuisen; Ding, Guangda; Wang, Zhenhua; Cai, Hongmei; Xu, Fangsen

    2015-03-18

    Given low solubility and immobility in many soils of the world, phosphorus (P) may be the most widely studied macronutrient for plants. In an attempt to gain an insight into the adaptability of Brassica napus to P deficiency, proteome alterations of roots and leaves in two B. napus contrasting genotypes, P-efficient 'Eyou Changjia' and P-inefficient 'B104-2', under long-term low P stress and short-term P-free starvation conditions were investigated, and proteomic combined with comparative genomic analyses were conducted to interpret the interrelation of differential abundance protein species (DAPs) responding to P deficiency with quantitative trait loci (QTLs) for P deficiency tolerance. P-efficient 'Eyou Changjia' had higher dry weight and P content, and showed high tolerance to low P stress compared with P-inefficient 'B104-2'. A total of 146 DAPs were successfully identified by MALDI TOF/TOF MS, which were categorized into several groups including defense and stress response, carbohydrate and energy metabolism, signaling and regulation, amino acid and fatty acid metabolism, protein process, biogenesis and cellular component, and function unknown. 94 of 146 DAPs were mapped to a linkage map constructed by a B. napus population derived from a cross between the two genotypes, and 72 DAPs were located in the confidence intervals of QTLs for P efficiency related traits. We conclude that the identification of these DAPs and the co-location of DAPs with QTLs in the B. napus linkage genetic map provide us novel information in understanding the adaptability of B. napus to P deficiency, and helpful to isolate P-efficient genes in B. napus. Low P seriously limits the production and quality of B. napus. Proteomics and genetic linkage map were widely used to study the adaptive strategies of B. napus response to P deficiency, proteomic combined with comparative genetic analysis to investigate the correlations between DAPs and QTLs are scarce. Thus, we herein investigated

  18. Identification of BAG3 target proteins in anaplastic thyroid cancer cells by proteomic analysis.

    Science.gov (United States)

    Galdiero, Francesca; Bello, Anna Maria; Spina, Anna; Capiluongo, Anna; Liuu, Sophie; De Marco, Margot; Rosati, Alessandra; Capunzo, Mario; Napolitano, Maria; Vuttariello, Emilia; Monaco, Mario; Califano, Daniela; Turco, Maria Caterina; Chiappetta, Gennaro; Vinh, Joëlle; Chiappetta, Giovanni

    2018-01-30

    BAG3 protein is an apoptosis inhibitor and is highly expressed in Anaplastic Thyroid Cancer. We investigated the entire set of proteins modulated by BAG3 silencing in the human anaplastic thyroid 8505C cancer cells by using the Stable-Isotope Labeling by Amino acids in Cell culture strategy combined with mass spectrometry analysis. By this approach we identified 37 up-regulated and 54 down-regulated proteins in BAG3-silenced cells. Many of these proteins are reportedly involved in tumor progression, invasiveness and resistance to therapies. We focused our attention on an oncogenic protein, CAV1, and a tumor suppressor protein, SERPINB2, that had not previously been reported to be modulated by BAG3. Their expression levels in BAG3-silenced cells were confirmed by qRT-PCR and western blot analyses, disclosing two novel targets of BAG3 pro-tumor activity. We also examined the dataset of proteins obtained by the quantitative proteomics analysis using two tools, Downstream Effect Analysis and Upstream Regulator Analysis of the Ingenuity Pathways Analysis software. Our analyses confirm the association of the proteome profile observed in BAG3-silenced cells with an increase in cell survival and a decrease in cell proliferation and invasion, and highlight the possible involvement of four tumor suppressor miRNAs and TP53/63 proteins in BAG3 activity.

  19. PETALS: Proteomic Evaluation and Topological Analysis of a mutated Locus' Signaling

    Directory of Open Access Journals (Sweden)

    Patel Vishal

    2010-12-01

    Full Text Available Abstract Background Colon cancer is driven by mutations in a number of genes, the most notorious of which is Apc. Though much of Apc's signaling has been mechanistically identified over the years, it is not always clear which functions or interactions are operative in a particular tumor. This is confounded by the presence of mutations in a number of other putative cancer driver (CAN genes, which often synergize with mutations in Apc. Computational methods are, thus, required to predict which pathways are likely to be operative when a particular mutation in Apc is observed. Results We developed a pipeline, PETALS, to predict and test likely signaling pathways connecting Apc to other CAN-genes, where the interaction network originating at Apc is defined as a "blossom," with each Apc-CAN-gene subnetwork referred to as a "petal." Known and predicted protein interactions are used to identify an Apc blossom with 24 petals. Then, using a novel measure of bimodality, the coexpression of each petal is evaluated against proteomic (2 D differential In Gel Electrophoresis, 2D-DIGE measurements from the Apc1638N+/-mouse to test the network-based hypotheses. Conclusions The predicted pathways linking Apc and Hapln1 exhibited the highest amount of bimodal coexpression with the proteomic targets, prioritizing the Apc-Hapln1 petal over other CAN-gene pairs and suggesting that this petal may be involved in regulating the observed proteome-level effects. These results not only demonstrate how functional 'omics data can be employed to test in silico predictions of CAN-gene pathways, but also reveal an approach to integrate models of upstream genetic interference with measured, downstream effects.

  20. Proteomic Analysis of Early Mid-Trimester Amniotic Fluid Does Not Predict Spontaneous Preterm Delivery

    Science.gov (United States)

    Lenco, Juraj; Vajrychova, Marie; Link, Marek; Tambor, Vojtech; Liman, Victor; Bullarbo, Maria; Nilsson, Staffan; Tsiartas, Panagiotis; Cobo, Teresa; Kacerovsky, Marian; Jacobsson, Bo

    2016-01-01

    Objective The aim of this study was to identify early proteomic biomarkers of spontaneous preterm delivery (PTD) in mid-trimester amniotic fluid from asymptomatic women. Methods This is a case-cohort study. Amniotic fluid from mid-trimester genetic amniocentesis (14–19 weeks of gestation) was collected from 2008 to 2011. The analysis was conducted in 24 healthy women with subsequent spontaneous PTD (cases) and 40 randomly selected healthy women delivering at term (controls). An exploratory phase with proteomics analysis of pooled samples was followed by a verification phase with ELISA of individual case and control samples. Results The median (interquartile range (IQR: 25th; 75th percentiles) gestational age at delivery was 35+5 (33+6–36+6) weeks in women with spontaneous PTD and 40+0 (39+1–40+5) weeks in women who delivered at term. In the exploratory phase, the most pronounced differences were found in C-reactive protein (CRP) levels, that were approximately two-fold higher in the pooled case samples than in the pooled control samples. However, we could not verify these differences with ELISA. The median (25th; 75th IQR) CRP level was 95.2 ng/mL (64.3; 163.5) in women with spontaneous PTD and 86.0 ng/mL (51.2; 145.8) in women delivering at term (p = 0.37; t-test). Conclusions Proteomic analysis with mass spectrometry of mid-trimester amniotic fluid suggests CRP as a potential marker of spontaneous preterm delivery, but this prognostic potential was not verified with ELISA. PMID:27214132

  1. Cereal Crop Proteomics: Systemic Analysis of Crop Drought Stress Responses Towards Marker-Assisted Selection Breeding

    Directory of Open Access Journals (Sweden)

    Arindam Ghatak

    2017-06-01

    Full Text Available Sustainable crop production is the major challenge in the current global climate change scenario. Drought stress is one of the most critical abiotic factors which negatively impact crop productivity. In recent years, knowledge about molecular regulation has been generated to understand drought stress responses. For example, information obtained by transcriptome analysis has enhanced our knowledge and facilitated the identification of candidate genes which can be utilized for plant breeding. On the other hand, it becomes more and more evident that the translational and post-translational machinery plays a major role in stress adaptation, especially for immediate molecular processes during stress adaptation. Therefore, it is essential to measure protein levels and post-translational protein modifications to reveal information about stress inducible signal perception and transduction, translational activity and induced protein levels. This information cannot be revealed by genomic or transcriptomic analysis. Eventually, these processes will provide more direct insight into stress perception then genetic markers and might build a complementary basis for future marker-assisted selection of drought resistance. In this review, we survey the role of proteomic studies to illustrate their applications in crop stress adaptation analysis with respect to productivity. Cereal crops such as wheat, rice, maize, barley, sorghum and pearl millet are discussed in detail. We provide a comprehensive and comparative overview of all detected protein changes involved in drought stress in these crops and have summarized existing knowledge into a proposed scheme of drought response. Based on a recent proteome study of pearl millet under drought stress we compare our findings with wheat proteomes and another recent study which defined genetic marker in pearl millet.

  2. Comparative proteomic analysis provides insight into cadmium stress responses in brown algae Sargassum fusiforme

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Aiqin; Xu, Tao [Alkali Soil Natural Environmental Science Center, Northeast Forestry University, Key Laboratory of Saline–alkali Vegetation Ecology Restoration in Oil Field, Ministry of Education, Harbin 150040 (China); Zou, Huixi [Zhejiang Provincial Key Laboratory for Subtropical Water Environment and Marine Biological Resources Protection, College of Life and Environmental Science, Wenzhou University, Wenzhou 325035 (China); Pang, Qiuying, E-mail: qiuying@nefu.edu.cn [Alkali Soil Natural Environmental Science Center, Northeast Forestry University, Key Laboratory of Saline–alkali Vegetation Ecology Restoration in Oil Field, Ministry of Education, Harbin 150040 (China)

    2015-06-15

    Highlights: • Proteomic analysis of brown algae response different level Cd stress was performed. • Proteins involved in carbohydrate metabolism were reduced under 1 day Cd stress. • 5 days Cd stress induced glycolysis and citrate cycle related proteins. • Graphic depiction of different metabolic pathways response to Cd stress was framed. - Abstract: Sargassum fusiforme is one of the most widely consumed seaweeds in China, Korea and Japan. In this work, we performed growth analysis and comparative proteomics to investigate the molecular mechanisms of the response to 1 day and 5 days Cd stress in S. fusiforme. Our results showed a significant decrease in growth rate and an increase in Cd ion content in S. fusiforme in response to Cd treatment. Comparative proteomic analysis revealed 25 and 51 differentially expressed protein spots in S. fusiforme under 1 day and 5 days Cd stress, respectively. A great number of these proteins was metabolic enzymes involved in carbohydrate metabolism and energy metabolism. Many proteins involved in the processing of genetic information showed a decrease in abundance under 1 day Cd stress. In contrast, 9 of the identified protein spots primarily involved in genetic information processing and carbohydrate metabolism were greatly enriched under 5 days Cd stress. Overall, our investigation indicated that Cd stress negatively affects the metabolic activity of S. fusiforme through the down-regulation of key metabolic enzymes. In addition, S. fusiforme may adapt to 5 days Cd stress by promoting consumption of photoassimilates through the up-regulation of glycolysis and the citrate cycle to supply energy for survival.

  3. Differential proteomic analysis of noncardia gastric cancer from individuals of northern Brazil.

    Science.gov (United States)

    Leal, Mariana Ferreira; Chung, Janete; Calcagno, Danielle Queiroz; Assumpção, Paulo Pimentel; Demachki, Samia; da Silva, Ismael Dale Cotrim Guerreiro; Chammas, Roger; Burbano, Rommel Rodríguez; de Arruda Cardoso Smith, Marília

    2012-01-01

    Gastric cancer is the second leading cause of cancer-related death worldwide. The identification of new cancer biomarkers is necessary to reduce the mortality rates through the development of new screening assays and early diagnosis, as well as new target therapies. In this study, we performed a proteomic analysis of noncardia gastric neoplasias of individuals from Northern Brazil. The proteins were analyzed by two-dimensional electrophoresis and mass spectrometry. For the identification of differentially expressed proteins, we used statistical tests with bootstrapping resampling to control the type I error in the multiple comparison analyses. We identified 111 proteins involved in gastric carcinogenesis. The computational analysis revealed several proteins involved in the energy production processes and reinforced the Warburg effect in gastric cancer. ENO1 and HSPB1 expression were further evaluated. ENO1 was selected due to its role in aerobic glycolysis that may contribute to the Warburg effect. Although we observed two up-regulated spots of ENO1 in the proteomic analysis, the mean expression of ENO1 was reduced in gastric tumors by western blot. However, mean ENO1 expression seems to increase in more invasive tumors. This lack of correlation between proteomic and western blot analyses may be due to the presence of other ENO1 spots that present a slightly reduced expression, but with a high impact in the mean protein expression. In neoplasias, HSPB1 is induced by cellular stress to protect cells against apoptosis. In the present study, HSPB1 presented an elevated protein and mRNA expression in a subset of gastric cancer samples. However, no association was observed between HSPB1 expression and clinicopathological characteristics. Here, we identified several possible biomarkers of gastric cancer in individuals from Northern Brazil. These biomarkers may be useful for the assessment of prognosis and stratification for therapy if validated in larger clinical study

  4. Differential proteomic analysis of noncardia gastric cancer from individuals of northern Brazil.

    Directory of Open Access Journals (Sweden)

    Mariana Ferreira Leal

    Full Text Available Gastric cancer is the second leading cause of cancer-related death worldwide. The identification of new cancer biomarkers is necessary to reduce the mortality rates through the development of new screening assays and early diagnosis, as well as new target therapies. In this study, we performed a proteomic analysis of noncardia gastric neoplasias of individuals from Northern Brazil. The proteins were analyzed by two-dimensional electrophoresis and mass spectrometry. For the identification of differentially expressed proteins, we used statistical tests with bootstrapping resampling to control the type I error in the multiple comparison analyses. We identified 111 proteins involved in gastric carcinogenesis. The computational analysis revealed several proteins involved in the energy production processes and reinforced the Warburg effect in gastric cancer. ENO1 and HSPB1 expression were further evaluated. ENO1 was selected due to its role in aerobic glycolysis that may contribute to the Warburg effect. Although we observed two up-regulated spots of ENO1 in the proteomic analysis, the mean expression of ENO1 was reduced in gastric tumors by western blot. However, mean ENO1 expression seems to increase in more invasive tumors. This lack of correlation between proteomic and western blot analyses may be due to the presence of other ENO1 spots that present a slightly reduced expression, but with a high impact in the mean protein expression. In neoplasias, HSPB1 is induced by cellular stress to protect cells against apoptosis. In the present study, HSPB1 presented an elevated protein and mRNA expression in a subset of gastric cancer samples. However, no association was observed between HSPB1 expression and clinicopathological characteristics. Here, we identified several possible biomarkers of gastric cancer in individuals from Northern Brazil. These biomarkers may be useful for the assessment of prognosis and stratification for therapy if validated in

  5. Comparative proteomic analysis provides insight into cadmium stress responses in brown algae Sargassum fusiforme

    International Nuclear Information System (INIS)

    Zhang, Aiqin; Xu, Tao; Zou, Huixi; Pang, Qiuying

    2015-01-01

    Highlights: • Proteomic analysis of brown algae response different level Cd stress was performed. • Proteins involved in carbohydrate metabolism were reduced under 1 day Cd stress. • 5 days Cd stress induced glycolysis and citrate cycle related proteins. • Graphic depiction of different metabolic pathways response to Cd stress was framed. - Abstract: Sargassum fusiforme is one of the most widely consumed seaweeds in China, Korea and Japan. In this work, we performed growth analysis and comparative proteomics to investigate the molecular mechanisms of the response to 1 day and 5 days Cd stress in S. fusiforme. Our results showed a significant decrease in growth rate and an increase in Cd ion content in S. fusiforme in response to Cd treatment. Comparative proteomic analysis revealed 25 and 51 differentially expressed protein spots in S. fusiforme under 1 day and 5 days Cd stress, respectively. A great number of these proteins was metabolic enzymes involved in carbohydrate metabolism and energy metabolism. Many proteins involved in the processing of genetic information showed a decrease in abundance under 1 day Cd stress. In contrast, 9 of the identified protein spots primarily involved in genetic information processing and carbohydrate metabolism were greatly enriched under 5 days Cd stress. Overall, our investigation indicated that Cd stress negatively affects the metabolic activity of S. fusiforme through the down-regulation of key metabolic enzymes. In addition, S. fusiforme may adapt to 5 days Cd stress by promoting consumption of photoassimilates through the up-regulation of glycolysis and the citrate cycle to supply energy for survival

  6. Proteomic analysis of Rhodotorula mucilaginosa: dealing with the issues of a non-conventional yeast.

    Science.gov (United States)

    Addis, Maria Filippa; Tanca, Alessandro; Landolfo, Sara; Abbondio, Marcello; Cutzu, Raffaela; Biosa, Grazia; Pagnozzi, Daniela; Uzzau, Sergio; Mannazzu, Ilaria

    2016-08-01

    Red yeasts ascribed to the species Rhodotorula mucilaginosa are gaining increasing attention, due to their numerous biotechnological applications, spanning carotenoid production, liquid bioremediation, heavy metal biotransformation and antifungal and plant growth-promoting actions, but also for their role as opportunistic pathogens. Nevertheless, their characterization at the 'omic' level is still scarce. Here, we applied different proteomic workflows to R. mucilaginosa with the aim of assessing their potential in generating information on proteins and functions of biotechnological interest, with a particular focus on the carotenogenic pathway. After optimization of protein extraction, we tested several gel-based (including 2D-DIGE) and gel-free sample preparation techniques, followed by tandem mass spectrometry analysis. Contextually, we evaluated different bioinformatic strategies for protein identification and interpretation of the biological significance of the dataset. When 2D-DIGE analysis was applied, not all spots returned a unambiguous identification and no carotenogenic enzymes were identified, even upon the application of different database search strategies. Then, the application of shotgun proteomic workflows with varying levels of sensitivity provided a picture of the information depth that can be reached with different analytical resources, and resulted in a plethora of information on R. mucilaginosa metabolism. However, also in these cases no proteins related to the carotenogenic pathway were identified, thus indicating that further improvements in sequence databases and functional annotations are strictly needed for increasing the outcome of proteomic analysis of this and other non-conventional yeasts. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

  7. iTRAQ-Based Quantitative Proteomic Analysis of the Initiation of Head Regeneration in Planarians.

    Directory of Open Access Journals (Sweden)

    Xiaofang Geng

    Full Text Available The planarian Dugesia japonica has amazing ability to regenerate a head from the anterior ends of the amputated stump with maintenance of the original anterior-posterior polarity. Although planarians present an attractive system for molecular investigation of regeneration and research has focused on clarifying the molecular mechanism of regeneration initiation in planarians at transcriptional level, but the initiation mechanism of planarian head regeneration (PHR remains unclear at the protein level. Here, a global analysis of proteome dynamics during the early stage of PHR was performed using isobaric tags for relative and absolute quantitation (iTRAQ-based quantitative proteomics strategy, and our data are available via ProteomeXchange with identifier PXD002100. The results showed that 162 proteins were differentially expressed at 2 h and 6 h following amputation. Furthermore, the analysis of expression patterns and functional enrichment of the differentially expressed proteins showed that proteins involved in muscle contraction, oxidation reduction and protein synthesis were up-regulated in the initiation of PHR. Moreover, ingenuity pathway analysis showed that predominant signaling pathways such as ILK, calcium, EIF2 and mTOR signaling which were associated with cell migration, cell proliferation and protein synthesis were likely to be involved in the initiation of PHR. The results for the first time demonstrated that muscle contraction and ILK signaling might played important roles in the initiation of PHR at the global protein level. The findings of this research provide a molecular basis for further unraveling the mechanism of head regeneration initiation in planarians.

  8. Urine proteome analysis in Dent's disease shows high selective changes potentially involved in chronic renal damage.

    Science.gov (United States)

    Santucci, Laura; Candiano, Giovanni; Anglani, Franca; Bruschi, Maurizio; Tosetto, Enrica; Cremasco, Daniela; Murer, Luisa; D'Ambrosio, Chiara; Scaloni, Andrea; Petretto, Andrea; Caridi, Gianluca; Rossi, Roberta; Bonanni, Alice; Ghiggeri, Gian Marco

    2016-01-01

    Definition of the urinary protein composition would represent a potential tool for diagnosis in many clinical conditions. The use of new proteomic technologies allows detection of genetic and post-trasductional variants that increase sensitivity of the approach but complicates comparison within a heterogeneous patient population. Overall, this limits research of urinary biomarkers. Studying monogenic diseases are useful models to address this issue since genetic variability is reduced among first- and second-degree relatives of the same family. We applied this concept to Dent's disease, a monogenic condition characterised by low-molecular-weight proteinuria that is inherited following an X-linked trait. Results are presented here on a combined proteomic approach (LC-mass spectrometry, Western blot and zymograms for proteases and inhibitors) to characterise urine proteins in a large family (18 members, 6 hemizygous patients, 6 carrier females, and 6 normals) with Dent's diseases due to the 1070G>T mutation of the CLCN5. Gene ontology analysis on more than 1000 proteins showed that several clusters of proteins characterised urine of affected patients compared to carrier females and normal subjects: proteins involved in extracellular matrix remodelling were the major group. Specific analysis on metalloproteases and their inhibitors underscored unexpected mechanisms potentially involved in renal fibrosis. Studying with new-generation techniques for proteomic analysis of the members of a large family with Dent's disease sharing the same molecular defect allowed highly repetitive results that justify conclusions. Identification in urine of proteins actively involved in interstitial matrix remodelling poses the question of active anti-fibrotic drugs in Dent's patients. Copyright © 2015 Elsevier B.V. All rights reserved.

  9. Proteome analysis of interaction between rootstocks and scions in ...

    African Journals Online (AJOL)

    The main propagation method of rubber tree (Hevea brasiliensis Muell. Arg.) is by grafting. However, the molecular mechanism underlying rootstock-scion interactions remains poorly understood. Identification and analysis of proteins related to rootstock-scion interactions are the bases of clarifying the molecular mechanism ...

  10. Common Data Analysis Pipeline | Office of Cancer Clinical Proteomics Research

    Science.gov (United States)

    CPTAC supports analyses of the mass spectrometry raw data (mapping of spectra to peptide sequences and protein identification) for the public using a Common Data Analysis Pipeline (CDAP). The data types available on the public portal are described below. A general overview of this pipeline can be downloaded here. Mass Spectrometry Data Formats RAW (Vendor) Format

  11. Proteomic analysis of human oral verrucous carcinoma | Wang ...

    African Journals Online (AJOL)

    Peptide mass fingerprints (PMFs) obtained by the MALDI-TOF analysis were applied to National Center for Biotechnology Information (NCBI), SWISS-PROT and MSDB databases using Mascot software. Then the 2-DE gel imaging showed that 74, 36 and 31 differential protein spots were found between OVC and OSCC, ...

  12. Proteomic analysis of heparin-binding proteins from human seminal ...

    Indian Academy of Sciences (India)

    Prakash

    (MALDI TOF/MS) for protein analysis of human HBPs. We resolved 70 ... Thus, the combined effects of seminal plasma components support the survival of ...... The BBXB motif of RANTES is the principal site for heparin binding and controls ...

  13. Molecular system analysis, multidimensional, dynamic, ultra-sensitive exploration of proteomes

    International Nuclear Information System (INIS)

    Scharattenholz, A.; Soski, V.; Stegmann, W.; Schroer, K.; Godovac-Zimmermann, J.; Cabuk, A.; Pejovi, V.; Wozny, W.; Cahill, M.A.; Drukier, A.K.; Volkovitsky, P.

    2001-01-01

    ProteoSys AG's holistic proteomics strategy extends beyond classical proteome research as a new paradigm. Our concept of multidimensional molecular systems analysis of complex model systems employs the innovative ProteoDyn TM approach. This enables us to correlate dynamic changes of proteomes with their biophysical and biochemical environment. Our supersensitive Multi Photon Detection (MPD) technology enables ultra-sensitive detection of proteins, deep into the low abundance domain. Our technology platform includes the affinity analysis of phospho- and glyco-proteomes, and with our 'fish hook' methods we can capture and fully characterize even serpentine G-coupled receptors and associated proteins, including routine comprehensive post-translational analyses performed by a well equipped mass spectrometry group. Throughput and quality is obtained by automation and high end robotics, with data management handled by a dedicated bioinformatics department. Thus ProteoSys AG has a range of state of the art and proprietary tools at its disposal to analyse even the most difficult complex model systems. MPD is an isotopic detection method proprietary to ProteoSys For MPD analysis we have implemented protocols where over 99% of proteins can be iodinated, and where the iodinated proteins can be identified by mass spectrometry. Because MPD measures the energy of detected particles, it can discriminate between signals originating from different isotopes co-electrophoresed by 2D-PAGE. Thus MPD imagers have a 'multicolour' functionality suitable for differential display and improved throughput, eliminating inter-gel variations. Importantly, MPD opens up not only the world of detection of low abundance proteins, but also identification and characterization. Radioactive low abundance protein spots containing less than one attomole of protein can be excised from a 2D-gel, mixed with unlabelled proteins, and 'tracked' by MPD. The identity of the labeled protein is determined by

  14. Unravelling the differences: comparative proteomic analysis of a clonal virulent and an attenuated Histomonas meleagridis strain.

    Science.gov (United States)

    Monoyios, Andreas; Patzl, Martina; Schlosser, Sarah; Hess, Michael; Bilic, Ivana

    2018-02-01

    The current study focused on Histomonas meleagridis, a unicellular protozoan, responsible for histomonosis in poultry. Recently, the occurrence of the disease increased due to the ban of effective chemotherapeutic drugs. Basic questions regarding the molecular biology, virulence mechanisms or even life cycle of the flagellate are still puzzling. In order to address some of these issues, we conducted a comparative proteomic analysis of a virulent and an attenuated H. meleagridis strain traced back to a single cell and propagated in vitro as monoxenic mono-eukaryotic cultures. Using two-dimensional electrophoresis (2-DE) for proteome visualization with computational 2-DE gel image and statistical analysis, upregulated proteins in either of the two H. meleagridis strains were detected. Statistical analysis fulfilling two criteria (≥threefold upregulation and P singled out 32 spots as specific for the flagellate. These spots were shown to correspond to 24 different proteins that reflect the increased metabolism, in vitro adaptation of the parasite, and amoeboid morphology. In addition to H. meleagridis proteins, the analysis identified differential expression of Escherichia coli DH5α proteins that could have been influenced by the co-cultivated H. meleagridis strain, indicating a reciprocal interaction of these two organisms during monoxenic cultivation. Copyright © 2017 The Author(s). Published by Elsevier Ltd.. All rights reserved.

  15. Nanoscale Proteomic Analysis of Oncoproteins in Hematopoietic Cancers

    Science.gov (United States)

    2012-05-01

    can be measured. Quantitation of AKT2 phosphorylated and unphosphorylated protein peaks was performed using Compass (version 1.8.0) analysis soft...5a SDf → PD 12 112 81F RAEB-2 Highh 5d SDg → PD 14+ 117 75F RAEB-2 Highh 3c,e PD skin (mCR) Yes 5 121 80M RAEB-T Highh 1c PD Yes 3 122 76F RAEB-1

  16. Targeted proteomics guided by label-free global proteome analysis in saliva reveal transition signatures from health to periodontal disease.

    Science.gov (United States)

    Bostanci, Nagihan; Selevsek, Nathalie; Wolski, Witold; Grossmann, Jonas; Bao, Kai; Wahlander, Asa; Trachsel, Christian; Schlapbach, Ralph; Özturk, Veli Özgen; Afacan, Beral; Emingil, Gulnur; Belibasakis, Georgios N

    2018-04-02

    Periodontal diseases are among the most prevalent worldwide, but largely silent, chronic diseases. They affect the tooth-supporting tissues with multiple ramifications on life quality. Their early diagnosis is still challenging, due to lack of appropriate molecular diagnostic methods. Saliva offers a non-invasively collectable reservoir of clinically relevant biomarkers, which, if utilized efficiently, could facilitate early diagnosis and monitoring of ongoing disease. Despite several novel protein markers being recently enlisted by discovery proteomics, their routine diagnostic application is hampered by the lack of validation platforms that allow for rapid, accurate and simultaneous quantification of multiple proteins in large cohorts. We carried out a pipeline of two proteomic platforms; firstly, we applied open ended label-free quantitative (LFQ) proteomics for discovery in saliva (n=67, health, gingivitis, and periodontitis), followed by selected-reaction monitoring (SRM)-targeted proteomics for validation in an independent cohort (n=82). The LFQ platform led to the discovery of 119 proteins with at least two-fold significant difference between health and disease. The 65 proteins chosen for the subsequent SRM platform included 50 related proteins derived from the significantly enriched processes of the LFQ data, 11 from literature-mining, and four house-keeping ones. Among those, 60 were reproducibly quantifiable proteins (92% success rate), represented by a total of 143 peptides. Machine-learning modeling led to a narrowed-down panel of five proteins of high predictive value for periodontal diseases (higher in disease: Matrix metalloproteinase-9, Ras-related protein-1, Actin-related protein 2/3 complex subunit 5; lower in disease: Clusterin, Deleted in Malignant Brain Tumors 1), with maximum area under the receiver operating curve >0.97. This panel enriches the pool of credible clinical biomarker candidates for diagnostic assay development. Yet, the quantum

  17. Proteomic Analysis Reveals the Leaf Color Regulation Mechanism in Chimera Hosta "Gold Standard" Leaves.

    Science.gov (United States)

    Yu, Juanjuan; Zhang, Jinzheng; Zhao, Qi; Liu, Yuelu; Chen, Sixue; Guo, Hongliang; Shi, Lei; Dai, Shaojun

    2016-03-08

    Leaf color change of variegated leaves from chimera species is regulated by fine-tuned molecular mechanisms. Hosta "Gold Standard" is a typical chimera Hosta species with golden-green variegated leaves, which is an ideal material to investigate the molecular mechanisms of leaf variegation. In this study, the margin and center regions of young and mature leaves from Hosta "Gold Standard", as well as the leaves from plants after excess nitrogen fertilization were studied using physiological and comparative proteomic approaches. We identified 31 differentially expressed proteins in various regions and development stages of variegated leaves. Some of them may be related to the leaf color regulation in Hosta "Gold Standard". For example, cytosolic glutamine synthetase (GS1), heat shock protein 70 (Hsp70), and chloroplastic elongation factor G (cpEF-G) were involved in pigment-related nitrogen synthesis as well as protein synthesis and processing. By integrating the proteomics data with physiological results, we revealed the metabolic patterns of nitrogen metabolism, photosynthesis, energy supply, as well as chloroplast protein synthesis, import and processing in various leaf regions at different development stages. Additionally, chloroplast-localized proteoforms involved in nitrogen metabolism, photosynthesis and protein processing implied that post-translational modifications were crucial for leaf color regulation. These results provide new clues toward understanding the mechanisms of leaf color regulation in variegated leaves.

  18. Selecting Sample Preparation Workflows for Mass Spectrometry-Based Proteomic and Phosphoproteomic Analysis of Patient Samples with Acute Myeloid Leukemia.

    Science.gov (United States)

    Hernandez-Valladares, Maria; Aasebø, Elise; Selheim, Frode; Berven, Frode S; Bruserud, Øystein

    2016-08-22

    Global mass spectrometry (MS)-based proteomic and phosphoproteomic studies of acute myeloid leukemia (AML) biomarkers represent a powerful strategy to identify and confirm proteins and their phosphorylated modifications that could be applied in diagnosis and prognosis, as a support for individual treatment regimens and selection of patients for bone marrow transplant. MS-based studies require optimal and reproducible workflows that allow a satisfactory coverage of the proteome and its modifications. Preparation of samples for global MS analysis is a crucial step and it usually requires method testing, tuning and optimization. Different proteomic workflows that have been used to prepare AML patient samples for global MS analysis usually include a standard protein in-solution digestion procedure with a urea-based lysis buffer. The enrichment of phosphopeptides from AML patient samples has previously been carried out either with immobilized metal affinity chromatography (IMAC) or metal oxide affinity chromatography (MOAC). We have recently tested several methods of sample preparation for MS analysis of the AML proteome and phosphoproteome and introduced filter-aided sample preparation (FASP) as a superior methodology for the sensitive and reproducible generation of peptides from patient samples. FASP-prepared peptides can be further fractionated or IMAC-enriched for proteome or phosphoproteome analyses. Herein, we will review both in-solution and FASP-based sample preparation workflows and encourage the use of the latter for the highest protein and phosphorylation coverage and reproducibility.

  19. Selecting Sample Preparation Workflows for Mass Spectrometry-Based Proteomic and Phosphoproteomic Analysis of Patient Samples with Acute Myeloid Leukemia

    Directory of Open Access Journals (Sweden)

    Maria Hernandez-Valladares

    2016-08-01

    Full Text Available Global mass spectrometry (MS-based proteomic and phosphoproteomic studies of acute myeloid leukemia (AML biomarkers represent a powerful strategy to identify and confirm proteins and their phosphorylated modifications that could be applied in diagnosis and prognosis, as a support for individual treatment regimens and selection of patients for bone marrow transplant. MS-based studies require optimal and reproducible workflows that allow a satisfactory coverage of the proteome and its modifications. Preparation of samples for global MS analysis is a crucial step and it usually requires method testing, tuning and optimization. Different proteomic workflows that have been used to prepare AML patient samples for global MS analysis usually include a standard protein in-solution digestion procedure with a urea-based lysis buffer. The enrichment of phosphopeptides from AML patient samples has previously been carried out either with immobilized metal affinity chromatography (IMAC or metal oxide affinity chromatography (MOAC. We have recently tested several methods of sample preparation for MS analysis of the AML proteome and phosphoproteome and introduced filter-aided sample preparation (FASP as a superior methodology for the sensitive and reproducible generation of peptides from patient samples. FASP-prepared peptides can be further fractionated or IMAC-enriched for proteome or phosphoproteome analyses. Herein, we will review both in-solution and FASP-based sample preparation workflows and encourage the use of the latter for the highest protein and phosphorylation coverage and reproducibility.

  20. Proteome profiling analysis of human ovarian cancer serum samples

    International Nuclear Information System (INIS)

    Cognetti, F.; Citro, G.

    2009-01-01

    Mass Spectrometry represents a powerful tool in cancer research to discovery of potential bio markers through peak identification from serum profiling. By using high resolution MALDITOF and bioinformatic analysis almost 400 serum sample homogeneously distributed between biopsy confirmed ovarian cancer and high risk serum samples were analyzed. Each serum sample run in duplicate and whole serum sample preparation procedure has been performed by Hamilton Star Robot in order to reduce bias and the replicates with a low Pearson coefficient are removed. After automated reverse phase magnetic beads separation the samples were tested in MALDI-TOF

  1. Proteomic analysis of isolated chlamydomonas centrioles reveals orthologs of ciliary-disease genes.

    Science.gov (United States)

    Keller, Lani C; Romijn, Edwin P; Zamora, Ivan; Yates, John R; Marshall, Wallace F

    2005-06-21

    The centriole is one of the most enigmatic organelles in the cell. Centrioles are cylindrical, microtubule-based barrels found in the core of the centrosome. Centrioles also act as basal bodies during interphase to nucleate the assembly of cilia and flagella. There are currently only a handful of known centriole proteins. We used mass-spectrometry-based MudPIT (multidimensional protein identification technology) to identify the protein composition of basal bodies (centrioles) isolated from the green alga Chlamydomonas reinhardtii. This analysis detected the majority of known centriole proteins, including centrin, epsilon tubulin, and the cartwheel protein BLD10p. By combining proteomic data with information about gene expression and comparative genomics, we identified 45 cross-validated centriole candidate proteins in two classes. Members of the first class of proteins (BUG1-BUG27) are encoded by genes whose expression correlates with flagellar assembly and which therefore may play a role in ciliogenesis-related functions of basal bodies. Members of the second class (POC1-POC18) are implicated by comparative-genomics and -proteomics studies to be conserved components of the centriole. We confirmed centriolar localization for the human homologs of four candidate proteins. Three of the cross-validated centriole candidate proteins are encoded by orthologs of genes (OFD1, NPHP-4, and PACRG) implicated in mammalian ciliary function and disease, suggesting that oral-facial-digital syndrome and nephronophthisis may involve a dysfunction of centrioles and/or basal bodies. By analyzing isolated Chlamydomonas basal bodies, we have been able to obtain the first reported proteomic analysis of the centriole.

  2. Quantitative Proteomic Analysis of Optimal Cutting Temperature (OCT) Embedded Core-Needle Biopsy of Lung Cancer

    Science.gov (United States)

    Zhao, Xiaozheng; Huffman, Kenneth E.; Fujimoto, Junya; Canales, Jamie Rodriguez; Girard, Luc; Nie, Guangjun; Heymach, John V.; Wistuba, Igacio I.; Minna, John D.; Yu, Yonghao

    2017-10-01

    With recent advances in understanding the genomic underpinnings and oncogenic drivers of pathogenesis in different subtypes, it is increasingly clear that proper pretreatment diagnostics are essential for the choice of appropriate treatment options for non-small cell lung cancer (NSCLC). Tumor tissue preservation in optimal cutting temperature (OCT) compound is commonly used in the surgical suite. However, proteins recovered from OCT-embedded specimens pose a challenge for LC-MS/MS experiments, due to the large amounts of polymers present in OCT. Here we present a simple workflow for whole proteome analysis of OCT-embedded NSCLC tissue samples, which involves a simple trichloroacetic acid precipitation step. Comparisons of protein recovery between frozen versus OCT-embedded tissue showed excellent consistency with more than 9200 proteins identified. Using an isobaric labeling strategy, we quantified more than 5400 proteins in tumor versus normal OCT-embedded core needle biopsy samples. Gene ontology analysis indicated that a number of proliferative as well as squamous cell carcinoma (SqCC) marker proteins were overexpressed in the tumor, consistent with the patient's pathology based diagnosis of "poorly differentiated SqCC". Among the most downregulated proteins in the tumor sample, we noted a number of proteins with potential immunomodulatory functions. Finally, interrogation of the aberrantly expressed proteins using a candidate approach and cross-referencing with publicly available databases led to the identification of potential druggable targets in DNA replication and DNA damage repair pathways. We conclude that our approach allows LC-MS/MS proteomic analyses on OCT-embedded lung cancer specimens, opening the way to bring powerful proteomics into the clinic. [Figure not available: see fulltext.

  3. Comparative proteomic analysis provides new insights into cadmium accumulation in rice grain under cadmium stress

    Energy Technology Data Exchange (ETDEWEB)

    Xue, Dawei, E-mail: dwxue@hznu.edu.cn [College of Life and Environmental Sciences, Hangzhou Normal University, Hangzhou 310036 (China); State Key Laboratory of Rice Biology, China National Rice Research Institute, Hangzhou 310006 (China); Jiang, Hua [State Key Laboratory Breeding Base for Zhejiang Sustainable Pest and Disease Control, Zhejiang Academy of Agricultural Science, Hangzhou 310021 (China); Deng, Xiangxiong; Zhang, Xiaoqin [College of Life and Environmental Sciences, Hangzhou Normal University, Hangzhou 310036 (China); Wang, Hua [State Key Laboratory Breeding Base for Zhejiang Sustainable Pest and Disease Control, Zhejiang Academy of Agricultural Science, Hangzhou 310021 (China); Xu, Xiangbin [College of Life and Environmental Sciences, Hangzhou Normal University, Hangzhou 310036 (China); Hu, Jiang; Zeng, Dali [State Key Laboratory of Rice Biology, China National Rice Research Institute, Hangzhou 310006 (China); Guo, Longbiao, E-mail: guolongbiao@caas.cn [State Key Laboratory of Rice Biology, China National Rice Research Institute, Hangzhou 310006 (China); Qian, Qian, E-mail: qianqian188@hotmail.com [College of Life and Environmental Sciences, Hangzhou Normal University, Hangzhou 310036 (China); State Key Laboratory of Rice Biology, China National Rice Research Institute, Hangzhou 310006 (China)

    2014-09-15

    Graphical abstract: - Highlights: • Cd is the most toxic heavy metal and is a major pollutant in rice grains. • The mechanism of Cd accumulation in rice grains has not been well demonstrated. • Proteomics analysis is carried out and the verification is implemented by QPCR. • Proteins associated with ROS and photosynthesis showed large variation in expression. - Abstract: Rice is one of the most important staple crops. During the growth season, rice plants are inevitably subjected to numerous stresses, among which heavy metal stress represented by cadmium contamination not only hindering the yield of rice but also affecting the food safety by Cd accumulating in rice grains. The mechanism of Cd accumulation in rice grains has not been well elucidated. In this study, we compare the proteomic difference between two genotypes with different Cd accumulation ability in grains. Verification of differentially expressed protein-encoding genes was analyzing by quantitative PCR (QPCR) and reanalysis of microarray expression data. Forty-seven proteins in total were successfully identified through proteomic screening. GO and KEGG enrichment analysis showed Cd accumulation triggered stress-related pathways in the cells, and strongly affecting metabolic pathways. Many proteins associated with nutrient reservoir and starch-related enzyme were identified in this study suggesting that a considerably damage on grain quality was caused. The results also implied stress response was initiated by the abnormal cells and the transmission of signals may mediated by reactive oxygen species (ROS). Our research will provide new insights into Cd accumulation in rice grain under Cd stress.

  4. Cell wall proteome analysis of Mycobacterium smegmatis strain MC2 155

    Directory of Open Access Journals (Sweden)

    De Buck Jeroen

    2010-04-01

    Full Text Available Abstract Background The usually non-pathogenic soil bacterium Mycobacterium smegmatis is commonly used as a model mycobacterial organism because it is fast growing and shares many features with pathogenic mycobacteria. Proteomic studies of M. smegmatis can shed light on mechanisms of mycobacterial growth, complex lipid metabolism, interactions with the bacterial environment and provide a tractable system for antimycobacterial drug development. The cell wall proteins are particularly interesting in this respect. The aim of this study was to construct a reference protein map for these proteins in M. smegmatis. Results A proteomic analysis approach, based on one dimensional polyacrylamide gel electrophoresis and LC-MS/MS, was used to identify and characterize the cell wall associated proteins of M. smegmatis. An enzymatic cell surface shaving method was used to determine the surface-exposed proteins. As a result, a total of 390 cell wall proteins and 63 surface-exposed proteins were identified. Further analysis of the 390 cell wall proteins provided the theoretical molecular mass and pI distributions and determined that 26 proteins are shared with the surface-exposed proteome. Detailed information about functional classification, signal peptides and number of transmembrane domains are given next to discussing the identified transcriptional regulators, transport proteins and the proteins involved in lipid metabolism and cell division. Conclusion In short, a comprehensive profile of the M. smegmatis cell wall subproteome is reported. The current research may help the identification of some valuable vaccine and drug target candidates and provide foundation for the future design of preventive, diagnostic, and therapeutic strategies against mycobacterial diseases.

  5. Comparative Proteomic Analysis of Yak Follicular Fluid during Estrus

    Directory of Open Access Journals (Sweden)

    Xian Guo

    2016-09-01

    Full Text Available The breeding of yaks is highly seasonal, there are many crucial proteins involved in the reproduction control program, especially in follicular development. In order to isolate differential proteins between mature and immature follicular fluid (FF of yak, the FF from yak follicles with different sizes were sampled respectively, and two-dimensional gel electrophoresis (2-DE of the proteins was carried out. After silver staining, the Image Master 2D platinum software was used for protein analysis and matrix-assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF-MS was performed for differential protein identification. The expression level of transferrin and enolase superfamily member 1 (ENOSF1 was determined by Western blotting for verification analysis. The results showed that 2-DE obtained an electrophoresis map of proteins from mature and immature yak FF with high resolution and repeatability. A comparison of protein profiles identified 12 differently expressed proteins, out of which 10 of them were upregulated while 2 were downregulated. Western blotting showed that the expression of transferrin and ENOSF1 was enhanced with follicular development. Both the obtained protein profiles and the differently expressed proteins identified in this study provided experimental data related to follicular development during yak breeding seasons. This study also laid the foundation for understanding the microenvironment during oocyte development.

  6. Proteomic Analysis of the Endosperm Ontogeny of Jatropha curcas L. Seeds.

    Science.gov (United States)

    Shah, Mohibullah; Soares, Emanoella L; Carvalho, Paulo C; Soares, Arlete A; Domont, Gilberto B; Nogueira, Fábio C S; Campos, Francisco A P

    2015-06-05

    Seeds of Jatropha curcas L. represent a potential source of raw material for the production of biodiesel. However, this use is hampered by the lack of basic information on the biosynthetic pathways associated with synthesis of toxic diterpenes, fatty acids, and triacylglycerols, as well as the pattern of deposition of storage proteins during seed development. In this study, we performed an in-depth proteome analysis of the endosperm isolated from five developmental stages which resulted in the identification of 1517, 1256, 1033, 752, and 307 proteins, respectively, summing up 1760 different proteins. Proteins with similar label free quantitation expression pattern were grouped into five clusters. The biological significance of these identifications is discussed with special focus on the analysis of seed storage proteins, proteins involved in the metabolism of fatty acids, carbohydrates, toxic components and proteolytic processing. Although several enzymes belonging to the biosynthesis of diterpenoid precursors were identified, we were unable to find any terpene synthase/cyclase, indicating that the synthesis of phorbol esters, the main toxic diterpenes, does not occur in seeds. The strategy used enabled us to provide a first in depth proteome analysis of the developing endosperm of this biodiesel plant, providing an important glimpse into the enzymatic machinery devoted to the production of C and N sources to sustain seed development.

  7. Enhanced detergent extraction for analysis of membrane proteomes by two-dimensional gel electrophoresis

    Directory of Open Access Journals (Sweden)

    Hsu Kimberly K

    2005-06-01

    Full Text Available Abstract Background The analysis of hydrophobic membrane proteins by two-dimensional gel electrophoresis has long been hampered by the concept of inherent difficulty due to solubility issues. We have optimized extraction protocols by varying the detergent composition of the solubilization buffer with a variety of commercially available non-ionic and zwitterionic detergents and detergent-like phospholipids. Results After initial analyses by one-dimensional SDS-PAGE, quantitative two-dimensional analyses of human erythrocyte membranes, mouse liver membranes, and mouse brain membranes, extracted with buffers that included the zwitterionic detergent MEGA 10 (decanoyl-N-methylglucamide and the zwitterionic lipid LPC (1-lauroyl lysophosphatidylcholine, showed selective improvement over extraction with the common 2-DE detergent CHAPS (3 [(3-cholamidopropyldimethylammonio]-1-propanesulfonate. Mixtures of the three detergents showed additive improvements in spot number, density, and resolution. Substantial improvements in the analysis of a brain membrane proteome were observed. Conclusion This study demonstrates that an optimized detergent mix, coupled with rigorous sample handling and electrophoretic protocols, enables simple and effective analysis of membrane proteomes using two-dimensional electrophoresis.

  8. Comprehensive proteome analysis of lysosomes reveals the diverse function of macrophages in immune responses.

    Science.gov (United States)

    Gao, Yanpan; Chen, Yanyu; Zhan, Shaohua; Zhang, Wenhao; Xiong, Feng; Ge, Wei

    2017-01-31

    Phagocytosis and autophagy in macrophages have been shown to be essential to both innate and adaptive immunity. Lysosomes are the main catabolic subcellular organelles responsible for degradation and recycling of both extracellular and intracellular material, which are the final steps in phagocytosis and autophagy. However, the molecular mechanisms underlying lysosomal functions after infection remain obscure. In this study, we conducted a quantitative proteomics analysis of the changes in constitution and glycosylation of proteins in lysosomes derived from murine RAW 264.7 macrophage cells treated with different types of pathogens comprising examples of bacteria (Listeria monocytogenes, L. m), DNA viruses (herpes simplex virus type-1, HSV-1) and RNA viruses (vesicular stomatitis virus, VSV). In total, 3,704 lysosome-related proteins and 300 potential glycosylation sites on 193 proteins were identified. Comparative analysis showed that the aforementioned pathogens induced distinct alterations in the proteome of the lysosome, which is closely associated with the immune functions of macrophages, such as toll-like receptor activation, inflammation and antigen-presentation. The most significant changes in proteins and fluctuations in glycosylation were also determined. Furthermore, Western blot analysis showed that the changes in expression of these proteins were undetectable at the whole cell level. Thus, our study provides unique insights into the function of lysosomes in macrophage activation and immune responses.

  9. Label free quantitative proteomics analysis on the cisplatin resistance in ovarian cancer cells.

    Science.gov (United States)

    Wang, F; Zhu, Y; Fang, S; Li, S; Liu, S

    2017-05-20

    Quantitative proteomics has been made great progress in recent years. Label free quantitative proteomics analysis based on the mass spectrometry is widely used. Using this technique, we determined the differentially expressed proteins in the cisplatin-sensitive ovarian cancer cells COC1 and cisplatin-resistant cells COC1/DDP before and after the application of cisplatin. Using the GO analysis, we classified those proteins into different subgroups bases on their cellular component, biological process, and molecular function. We also used KEGG pathway analysis to determine the key signal pathways that those proteins were involved in. There are 710 differential proteins between COC1 and COC1/DDP cells, 783 between COC1 and COC1/DDP cells treated with cisplatin, 917 between the COC1/DDP cells and COC1/DDP cells treated with LaCl3, 775 between COC1/DDP cells treated with cisplatin and COC1/DDP cells treated with cisplatin and LaCl3. Among the same 411 differentially expressed proteins in cisplatin-sensitive COC1 cells and cisplain-resistant COC1/DDP cells before and after cisplatin treatment, 14% of them were localized on the cell membrane. According to the KEGG results, differentially expressed proteins were classified into 21 groups. The most abundant proteins were involved in spliceosome. This study lays a foundation for deciphering the mechanism for drug resistance in ovarian tumor.

  10. Comparative Proteomic Analysis of Human Lung Adenocarcinoma Cisplatin-resistant Cell Strain A549/CDDP

    Directory of Open Access Journals (Sweden)

    Sien SHI

    2009-11-01

    Full Text Available Background and objective Chemotherapy plays an important role in the comprehensive therapy of lung cancer. However, the drug-resistance often causes the failure of the chemotherapy. The aim of this study is to identify differently expressed protein before and after cisplatin resistance of human lung adenocarcinoma cell A549 by proteomic analysis. Methods Cisplatin-resistant cell strain A549/CDDP was established by combining gradually increasing concentration of cisplatin with large dosage impact. Comparative proteomic analysis of A549 and A549/CDDP were carried out by means of two-dimensional gel electrophoresis. The differentially expressed proteins were detected and identified by MALDI-TOF mass spectrometry. Results Eighty-two differentially expressed proteins were screened by analysis the electrophoretic maps of A549 and A549/CDDP. Six differential proteins were analyzed by peptide mass fingerprinting. Glucose regulating protein 75, ribosomal protein S4, mitochondrial ATP synthase F1 complex beta subunit and immunoglobulin heavy chain variable region were identified. All four differentially expressed proteins were over-expressed in A549/CDDP, whereas low-expressed or no-expressed in A549. Conclusion These differentially expressed proteins give some clues to elucidate the mechanism of lung cancer cell resistant of cisplatin, providing the basis of searching for potential target of chemotherapy of lung cancer.

  11. Proteomic analysis of osteogenic differentiation of dental follicle precursor cells

    DEFF Research Database (Denmark)

    Morsczeck, Christian; Petersen, Jørgen; Völlner, Florian

    2009-01-01

    of differentiation. In the present study we applied 2-DE combined with capillary-LC-MS/MS analysis to profile differentially regulated proteins upon differentiation of dental follicle precursor cells (DFPCs). Out of 115 differentially regulated proteins, glutamine synthetase, lysosomal proteinase cathepsin B....... The bioinformatic analyses suggest that proteins associated with cell cycle progression and protein metabolism were down-regulated and proteins involved in catabolism, cell motility and biological quality were up-regulated. These results display the general physiological state of DFPCs before and after osteogenic...... proteins, plastin 3 T-isoform, beta-actin, superoxide dismutases, and transgelin were found to be highly up-regulated, whereas cofilin-1, pro-alpha 1 collagen, destrin, prolyl 4-hydrolase and dihydrolipoamide dehydrogenase were found to be highly down-regulated. The group of up-regulated proteins...

  12. Comparative analysis of proteome and transcriptome variation in mouse.

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    Anatole Ghazalpour

    2011-06-01

    Full Text Available The relationships between the levels of transcripts and the levels of the proteins they encode have not been examined comprehensively in mammals, although previous work in plants and yeast suggest a surprisingly modest correlation. We have examined this issue using a genetic approach in which natural variations were used to perturb both transcript levels and protein levels among inbred strains of mice. We quantified over 5,000 peptides and over 22,000 transcripts in livers of 97 inbred and recombinant inbred strains and focused on the 7,185 most heritable transcripts and 486 most reliable proteins. The transcript levels were quantified by microarray analysis in three replicates and the proteins were quantified by Liquid Chromatography-Mass Spectrometry using O(18-reference-based isotope labeling approach. We show that the levels of transcripts and proteins correlate significantly for only about half of the genes tested, with an average correlation of 0.27, and the correlations of transcripts and proteins varied depending on the cellular location and biological function of the gene. We examined technical and biological factors that could contribute to the modest correlation. For example, differential splicing clearly affects the analyses for certain genes; but, based on deep sequencing, this does not substantially contribute to the overall estimate of the correlation. We also employed genome-wide association analyses to map loci controlling both transcript and protein levels. Surprisingly, little overlap was observed between the protein- and transcript-mapped loci. We have typed numerous clinically relevant traits among the strains, including adiposity, lipoprotein levels, and tissue parameters. Using correlation analysis, we found that a low number of clinical trait relationships are preserved between the protein and mRNA gene products and that the majority of such relationships are specific to either the protein levels or transcript levels

  13. Proteomic approaches in brain research and neuropharmacology.

    Science.gov (United States)

    Vercauteren, Freya G G; Bergeron, John J M; Vandesande, Frans; Arckens, Lut; Quirion, Rémi

    2004-10-01

    Numerous applications of genomic technologies have enabled the assembly of unprecedented inventories of genes, expressed in cells under specific physiological and pathophysiological conditions. Complementing the valuable information generated through functional genomics with the integrative knowledge of protein expression and function should enable the development of more efficient diagnostic tools and therapeutic agents. Proteomic analyses are particularly suitable to elucidate posttranslational modifications, expression levels and protein-protein interactions of thousands of proteins at a time. In this review, two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) investigations of brain tissues in neurodegenerative diseases such as Alzheimer's disease, Down syndrome and schizophrenia, and the construction of 2D-PAGE proteome maps of the brain are discussed. The role of the Human Proteome Organization (HUPO) as an international coordinating organization for proteomic efforts, as well as challenges for proteomic technologies and data analysis are also addressed. It is expected that the use of proteomic strategies will have significant impact in neuropharmacology over the coming decade.

  14. Proteomic Analysis of the Mediator Complex Interactome in Saccharomyces cerevisiae.

    Science.gov (United States)

    Uthe, Henriette; Vanselow, Jens T; Schlosser, Andreas

    2017-02-27

    Here we present the most comprehensive analysis of the yeast Mediator complex interactome to date. Particularly gentle cell lysis and co-immunopurification conditions allowed us to preserve even transient protein-protein interactions and to comprehensively probe the molecular environment of the Mediator complex in the cell. Metabolic 15 N-labeling thereby enabled stringent discrimination between bona fide interaction partners and nonspecifically captured proteins. Our data indicates a functional role for Mediator beyond transcription initiation. We identified a large number of Mediator-interacting proteins and protein complexes, such as RNA polymerase II, general transcription factors, a large number of transcriptional activators, the SAGA complex, chromatin remodeling complexes, histone chaperones, highly acetylated histones, as well as proteins playing a role in co-transcriptional processes, such as splicing, mRNA decapping and mRNA decay. Moreover, our data provides clear evidence, that the Mediator complex interacts not only with RNA polymerase II, but also with RNA polymerases I and III, and indicates a functional role of the Mediator complex in rRNA processing and ribosome biogenesis.

  15. PINK1-Interacting Proteins: Proteomic Analysis of Overexpressed PINK1

    Directory of Open Access Journals (Sweden)

    Aleksandar Rakovic

    2011-01-01

    Full Text Available Recent publications suggest that the Parkinson's disease- (PD- related PINK1/Parkin pathway promotes elimination of dysfunctional mitochondria by autophagy. We used tandem affinity purification (TAP, SDS-PAGE, and mass spectrometry as a first step towards identification of possible substrates for PINK1. The cellular abundance of selected identified interactors was investigated by Western blotting. Furthermore, one candidate gene was sequenced in 46 patients with atypical PD. In addition to two known binding partners (HSP90, CDC37, 12 proteins were identified using the TAP assay; four of which are mitochondrially localized (GRP75, HSP60, LRPPRC, and TUFM. Western blot analysis showed no differences in cellular abundance of these proteins comparing PINK1 mutant and control fibroblasts. When sequencing LRPPRC, four exonic synonymous changes and 20 polymorphisms in noncoding regions were detected. Our study provides a list of putative PINK1 binding partners, confirming previously described interactions, but also introducing novel mitochondrial proteins as potential components of the PINK1/Parkin mitophagy pathway.

  16. Comparative Proteomic Analysis of Peritoneal Dialysate from Chronic Glomerulonephritis Patients

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    Hsin-Yi Wu

    2013-01-01

    Full Text Available Peritoneal dialysis (PD frequently contributes to peritoneal damage which cannot be easily identified without invasive techniques, implying the urgent need for biomarkers and revealing mechanisms. Chronic glomerulonephritis (CGN is one of the leading causes of receiving dialysis treatment. Here, we attempted to analyze the peritoneal dialysate collected from CGN patients when they receive continuous ambulatory peritoneal dialysis (CAPD treatment for the first time and after a year to reveal the protein changes that resulted from PD. Proteins were displayed by two-dimensional gel electrophoresis (2DE. Altered gel spots were digested followed by liquid chromatography-tandem mass spectrometry (LC-MS/MS analysis for protein identification. Eight proteins were found to have differential expression levels between two groups. Their differential expressions were validated by Western blots in other sets of peritoneal dialysates. Proteins identified with higher levels in the first-time dialysate suggested their dominant appearance in CGN patients, while those that showed higher levels in peritoneal dialysate collected after one year may result from initial peritoneal inflammation or changes in the permeability of the peritoneum to middle-sized proteins. All the identified proteins may provide a perceptiveness of peritoneal changes caused by PD and may function as potential biomarkers or drug targets.

  17. Protein functional analysis data in support of comparative proteomics of the pathogenic black yeast Exophiala dermatitidis under different temperature conditions

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    Donatella Tesei

    2015-12-01

    Full Text Available In the current study a comparative proteomic approach was used to investigate the response of the human pathogen black yeast Exophiala dermatitidis toward temperature treatment. Protein functional analysis – based on cellular process GO terms – was performed on the 32 temperature-responsive identified proteins. The bioinformatics analyses and data presented here provided novel insights