HypoxiaDB: a database of hypoxia-regulated proteins

PubMed Central

Khurana, Pankaj; Sugadev, Ragumani; Jain, Jaspreet; Singh, Shashi Bala

2013-01-01

There has been intense interest in the cellular response to hypoxia, and a large number of differentially expressed proteins have been identified through various high-throughput experiments. These valuable data are scattered, and there have been no systematic attempts to document the various proteins regulated by hypoxia. Compilation, curation and annotation of these data are important in deciphering their role in hypoxia and hypoxia-related disorders. Therefore, we have compiled HypoxiaDB, a database of hypoxia-regulated proteins. It is a comprehensive, manually-curated, non-redundant catalog of proteins whose expressions are shown experimentally to be altered at different levels and durations of hypoxia. The database currently contains 72 000 manually curated entries taken on 3500 proteins extracted from 73 peer-reviewed publications selected from PubMed. HypoxiaDB is distinctive from other generalized databases: (i) it compiles tissue-specific protein expression changes under different levels and duration of hypoxia. Also, it provides manually curated literature references to support the inclusion of the protein in the database and establish its association with hypoxia. (ii) For each protein, HypoxiaDB integrates data on gene ontology, KEGG (Kyoto Encyclopedia of Genes and Genomes) pathway, protein–protein interactions, protein family (Pfam), OMIM (Online Mendelian Inheritance in Man), PDB (Protein Data Bank) structures and homology to other sequenced genomes. (iii) It also provides pre-compiled information on hypoxia-proteins, which otherwise requires tedious computational analysis. This includes information like chromosomal location, identifiers like Entrez, HGNC, Unigene, Uniprot, Ensembl, Vega, GI numbers and Genbank accession numbers associated with the protein. These are further cross-linked to respective public databases augmenting HypoxiaDB to the external repositories. (iv) In addition, HypoxiaDB provides an online sequence-similarity search tool for users to compare their protein sequences with HypoxiaDB protein database. We hope that HypoxiaDB will enrich our knowledge about hypoxia-related biology and eventually will lead to the development of novel hypothesis and advancements in diagnostic and therapeutic activities. HypoxiaDB is freely accessible for academic and non-profit users via http://www.hypoxiadb.com. Database URL: http://www.hypoxiadb.com PMID:24178989

AN OVERVIEW OF COMPUTATIONAL LIFE SCIENCE DATABASES & EXCHANGE FORMATS OF RELEVANCE TO CHEMICAL BIOLOGY RESEARCH

PubMed Central

Hall, Aaron Smalter; Shan, Yunfeng; Lushington, Gerald; Visvanathan, Mahesh

2016-01-01

Databases and exchange formats describing biological entities such as chemicals and proteins, along with their relationships, are a critical component of research in life sciences disciplines, including chemical biology wherein small information about small molecule properties converges with cellular and molecular biology. Databases for storing biological entities are growing not only in size, but also in type, with many similarities between them and often subtle differences. The data formats available to describe and exchange these entities are numerous as well. In general, each format is optimized for a particular purpose or database, and hence some understanding of these formats is required when choosing one for research purposes. This paper reviews a selection of different databases and data formats with the goal of summarizing their purposes, features, and limitations. Databases are reviewed under the categories of 1) protein interactions, 2) metabolic pathways, 3) chemical interactions, and 4) drug discovery. Representation formats will be discussed according to those describing chemical structures, and those describing genomic/proteomic entities. PMID:22934944

An overview of computational life science databases & exchange formats of relevance to chemical biology research.

PubMed

Smalter Hall, Aaron; Shan, Yunfeng; Lushington, Gerald; Visvanathan, Mahesh

2013-03-01

Databases and exchange formats describing biological entities such as chemicals and proteins, along with their relationships, are a critical component of research in life sciences disciplines, including chemical biology wherein small information about small molecule properties converges with cellular and molecular biology. Databases for storing biological entities are growing not only in size, but also in type, with many similarities between them and often subtle differences. The data formats available to describe and exchange these entities are numerous as well. In general, each format is optimized for a particular purpose or database, and hence some understanding of these formats is required when choosing one for research purposes. This paper reviews a selection of different databases and data formats with the goal of summarizing their purposes, features, and limitations. Databases are reviewed under the categories of 1) protein interactions, 2) metabolic pathways, 3) chemical interactions, and 4) drug discovery. Representation formats will be discussed according to those describing chemical structures, and those describing genomic/proteomic entities.

Genome-Wide Identification of Arabidopsis Coiled-Coil Proteins and Establishment of the ARABI-COIL Database1

PubMed Central

Rose, Annkatrin; Manikantan, Sankaraganesh; Schraegle, Shannon J.; Maloy, Michael A.; Stahlberg, Eric A.; Meier, Iris

2004-01-01

Increasing evidence demonstrates the importance of long coiled-coil proteins for the spatial organization of cellular processes. Although several protein classes with long coiled-coil domains have been studied in animals and yeast, our knowledge about plant long coiled-coil proteins is very limited. The repeat nature of the coiled-coil sequence motif often prevents the simple identification of homologs of animal coiled-coil proteins by generic sequence similarity searches. As a consequence, counterparts of many animal proteins with long coiled-coil domains, like lamins, golgins, or microtubule organization center components, have not been identified yet in plants. Here, all Arabidopsis proteins predicted to contain long stretches of coiled-coil domains were identified by applying the algorithm MultiCoil to a genome-wide screen. A searchable protein database, ARABI-COIL (http://www.coiled-coil.org/arabidopsis), was established that integrates information on number, size, and position of predicted coiled-coil domains with subcellular localization signals, transmembrane domains, and available functional annotations. ARABI-COIL serves as a tool to sort and browse Arabidopsis long coiled-coil proteins to facilitate the identification and selection of candidate proteins of potential interest for specific research areas. Using the database, candidate proteins were identified for Arabidopsis membrane-bound, nuclear, and organellar long coiled-coil proteins. PMID:15020757

Secretome profile analysis of multidrug-resistant, monodrug-resistant and drug-susceptible Mycobacterium tuberculosis.

PubMed

Putim, Chanyanuch; Phaonakrop, Narumon; Jaresitthikunchai, Janthima; Gamngoen, Ratikorn; Tragoolpua, Khajornsak; Intorasoot, Sorasak; Anukool, Usanee; Tharincharoen, Chayada Sitthidet; Phunpae, Ponrut; Tayapiwatana, Chatchai; Kasinrerk, Watchara; Roytrakul, Sittiruk; Butr-Indr, Bordin

2018-03-01

The emergence of drug-resistant tuberculosis has generated great concern in the control of tuberculosis and HIV/TB patients have established severe complications that are difficult to treat. Although, the gold standard of drug-susceptibility testing is highly accurate and efficient, it is time-consuming. Diagnostic biomarkers are, therefore, necessary in discriminating between infection from drug-resistant and drug-susceptible strains. One strategy that aids to effectively control tuberculosis is understanding the function of secreting proteins that mycobacteria use to manipulate the host cellular defenses. In this study, culture filtrate proteins from Mycobacterium tuberculosis H37Rv, isoniazid-resistant, rifampicin-resistant and multidrug-resistant strains were gathered and profiled by shotgun-proteomics technique. Mass spectrometric analysis of the secreted proteome identified several proteins, of which 837, 892, 838 and 850 were found in M. tuberculosis H37Rv, isoniazid-resistant, rifampicin-resistant and multidrug-resistant strains, respectively. These proteins have been implicated in various cellular processes, including biological adhesion, biological regulation, developmental process, immune system process localization, cellular process, cellular component organization or biogenesis, metabolic process, and response to stimulus. Analysis based on STITCH database predicted the interaction of DNA topoisomerase I, 3-oxoacyl-(acyl-carrier protein) reductase, ESAT-6-like protein, putative prophage phiRv2 integrase, and 3-phosphoshikimate 1-carboxyvinyltransferase with isoniazid, rifampicin, pyrazinamide, ethambutol and streptomycin, suggesting putative roles in controlling the anti-tuberculosis ability. However, several proteins with no interaction with all first-line anti-tuberculosis drugs might be used as markers for mycobacterial identification.

RaftProt: mammalian lipid raft proteome database.

PubMed

Shah, Anup; Chen, David; Boda, Akash R; Foster, Leonard J; Davis, Melissa J; Hill, Michelle M

2015-01-01

RaftProt (http://lipid-raft-database.di.uq.edu.au/) is a database of mammalian lipid raft-associated proteins as reported in high-throughput mass spectrometry studies. Lipid rafts are specialized membrane microdomains enriched in cholesterol and sphingolipids thought to act as dynamic signalling and sorting platforms. Given their fundamental roles in cellular regulation, there is a plethora of information on the size, composition and regulation of these membrane microdomains, including a large number of proteomics studies. To facilitate the mining and analysis of published lipid raft proteomics studies, we have developed a searchable database RaftProt. In addition to browsing the studies, performing basic queries by protein and gene names, searching experiments by cell, tissue and organisms; we have implemented several advanced features to facilitate data mining. To address the issue of potential bias due to biochemical preparation procedures used, we have captured the lipid raft preparation methods and implemented advanced search option for methodology and sample treatment conditions, such as cholesterol depletion. Furthermore, we have identified a list of high confidence proteins, and enabled searching only from this list of likely bona fide lipid raft proteins. Given the apparent biological importance of lipid raft and their associated proteins, this database would constitute a key resource for the scientific community. © The Author(s) 2014. Published by Oxford University Press on behalf of Nucleic Acids Research.

mpMoRFsDB: a database of molecular recognition features in membrane proteins.

PubMed

Gypas, Foivos; Tsaousis, Georgios N; Hamodrakas, Stavros J

2013-10-01

Molecular recognition features (MoRFs) are small, intrinsically disordered regions in proteins that undergo a disorder-to-order transition on binding to their partners. MoRFs are involved in protein-protein interactions and may function as the initial step in molecular recognition. The aim of this work was to collect, organize and store all membrane proteins that contain MoRFs. Membrane proteins constitute ∼30% of fully sequenced proteomes and are responsible for a wide variety of cellular functions. MoRFs were classified according to their secondary structure, after interacting with their partners. We identified MoRFs in transmembrane and peripheral membrane proteins. The position of transmembrane protein MoRFs was determined in relation to a protein's topology. All information was stored in a publicly available mySQL database with a user-friendly web interface. A Jmol applet is integrated for visualization of the structures. mpMoRFsDB provides valuable information related to disorder-based protein-protein interactions in membrane proteins. http://bioinformatics.biol.uoa.gr/mpMoRFsDB

Mps1 (Monopolar Spindle 1) Protein Inhibition Affects Cellular Growth and Pro-Embryogenic Masses Morphology in Embryogenic Cultures of Araucaria angustifolia (Araucariaceae).

PubMed

Douétts-Peres, Jackellinne C; Cruz, Marco Antônio L; Reis, Ricardo S; Heringer, Angelo S; de Oliveira, Eduardo A G; Elbl, Paula M; Floh, Eny I S; Silveira, Vanildo; Santa-Catarina, Claudete

2016-01-01

Somatic embryogenesis has been shown to be an efficient tool for studying processes based on cell growth and development. The fine regulation of the cell cycle is essential for proper embryo formation during the process of somatic embryogenesis. The aims of the present work were to identify and perform a structural and functional characterization of Mps1 and to analyze the effects of the inhibition of this protein on cellular growth and pro-embryogenic mass (PEM) morphology in embryogenic cultures of A. angustifolia. A single-copy Mps1 gene named AaMps1 was retrieved from the A. angustifolia transcriptome database, and through a mass spectrometry approach, AaMps1 was identified and quantified in embryogenic cultures. The Mps1 inhibitor SP600125 (10 μM) inhibited cellular growth and changed PEMs, and these effects were accompanied by a reduction in AaMps1 protein levels in embryogenic cultures. Our work has identified the Mps1 protein in a gymnosperm species for the first time, and we have shown that inhibiting Mps1 affects cellular growth and PEM differentiation during A. angustifolia somatic embryogenesis. These data will be useful for better understanding cell cycle control during somatic embryogenesis in plants.

Mps1 (Monopolar Spindle 1) Protein Inhibition Affects Cellular Growth and Pro-Embryogenic Masses Morphology in Embryogenic Cultures of Araucaria angustifolia (Araucariaceae)

PubMed Central

Douétts-Peres, Jackellinne C.; Cruz, Marco Antônio L.; Reis, Ricardo S.; Heringer, Angelo S.; de Oliveira, Eduardo A. G.; Elbl, Paula M.; Floh, Eny I. S.; Silveira, Vanildo

2016-01-01

Somatic embryogenesis has been shown to be an efficient tool for studying processes based on cell growth and development. The fine regulation of the cell cycle is essential for proper embryo formation during the process of somatic embryogenesis. The aims of the present work were to identify and perform a structural and functional characterization of Mps1 and to analyze the effects of the inhibition of this protein on cellular growth and pro-embryogenic mass (PEM) morphology in embryogenic cultures of A. angustifolia. A single-copy Mps1 gene named AaMps1 was retrieved from the A. angustifolia transcriptome database, and through a mass spectrometry approach, AaMps1 was identified and quantified in embryogenic cultures. The Mps1 inhibitor SP600125 (10 μM) inhibited cellular growth and changed PEMs, and these effects were accompanied by a reduction in AaMps1 protein levels in embryogenic cultures. Our work has identified the Mps1 protein in a gymnosperm species for the first time, and we have shown that inhibiting Mps1 affects cellular growth and PEM differentiation during A. angustifolia somatic embryogenesis. These data will be useful for better understanding cell cycle control during somatic embryogenesis in plants. PMID:27064899

Proteomic analysis by iTRAQ in red claw crayfish, Cherax quadricarinatus, hematopoietic tissue cells post white spot syndrome virus infection.

PubMed

Jeswin, Joseph; Xie, Xiao-lu; Ji, Qiao-lin; Wang, Ke-jian; Liu, Hai-peng

2016-03-01

To elucidate proteomic changes of Hpt cells from red claw crayfish, Cherax quadricarinatus, we have carried out isobaric tags for relative and absolute quantitation (iTRAQ) of cellular proteins at both early (1 hpi) and late stage (12 hpi) post white spot syndrome virus (WSSV) infection. Protein database search revealed 594 protein hits by Mascot, in which 17 and 30 proteins were present as differentially expressed proteins at early and late viral infection, respectively. Generally, these differentially expressed proteins include: 1) the metabolic process related proteins in glycolysis and glucogenesis, DNA replication, nucleotide/amino acid/fatty acid metabolism and protein biosynthesis; 2) the signal transduction related proteins like small GTPases, G-protein-alpha stimulatory subunit, proteins bearing PDZ- or 14-3-3-domains that help holding together and organize signaling complexes, casein kinase I and proteins of the MAP-kinase signal transduction pathway; 3) the immune defense related proteins such as α-2 macroglobulin, transglutaminase and trans-activation response RNA-binding protein 1. Taken together, these protein information shed new light on the host cellular response against WSSV infection in a crustacean cell culture. Copyright © 2016 Elsevier Ltd. All rights reserved.

CPLA 1.0: an integrated database of protein lysine acetylation.

PubMed

Liu, Zexian; Cao, Jun; Gao, Xinjiao; Zhou, Yanhong; Wen, Longping; Yang, Xiangjiao; Yao, Xuebiao; Ren, Jian; Xue, Yu

2011-01-01

As a reversible post-translational modification (PTM) discovered decades ago, protein lysine acetylation was known for its regulation of transcription through the modification of histones. Recent studies discovered that lysine acetylation targets broad substrates and especially plays an essential role in cellular metabolic regulation. Although acetylation is comparable with other major PTMs such as phosphorylation, an integrated resource still remains to be developed. In this work, we presented the compendium of protein lysine acetylation (CPLA) database for lysine acetylated substrates with their sites. From the scientific literature, we manually collected 7151 experimentally identified acetylation sites in 3311 targets. We statistically studied the regulatory roles of lysine acetylation by analyzing the Gene Ontology (GO) and InterPro annotations. Combined with protein-protein interaction information, we systematically discovered a potential human lysine acetylation network (HLAN) among histone acetyltransferases (HATs), substrates and histone deacetylases (HDACs). In particular, there are 1862 triplet relationships of HAT-substrate-HDAC retrieved from the HLAN, at least 13 of which were previously experimentally verified. The online services of CPLA database was implemented in PHP + MySQL + JavaScript, while the local packages were developed in JAVA 1.5 (J2SE 5.0). The CPLA database is freely available for all users at: http://cpla.biocuckoo.org.

The systematic annotation of the three main GPCR families in Reactome.

PubMed

Jassal, Bijay; Jupe, Steven; Caudy, Michael; Birney, Ewan; Stein, Lincoln; Hermjakob, Henning; D'Eustachio, Peter

2010-07-29

Reactome is an open-source, freely available database of human biological pathways and processes. A major goal of our work is to provide an integrated view of cellular signalling processes that spans from ligand-receptor interactions to molecular readouts at the level of metabolic and transcriptional events. To this end, we have built the first catalogue of all human G protein-coupled receptors (GPCRs) known to bind endogenous or natural ligands. The UniProt database has records for 797 proteins classified as GPCRs and sorted into families A/1, B/2 and C/3 on the basis of amino acid sequence. To these records we have added details from the IUPHAR database and our own manual curation of relevant literature to create reactions in which 563 GPCRs bind ligands and also interact with specific G-proteins to initiate signalling cascades. We believe the remaining 234 GPCRs are true orphans. The Reactome GPCR pathway can be viewed as a detailed interactive diagram and can be exported in many forms. It provides a template for the orthology-based inference of GPCR reactions for diverse model organism species, and can be overlaid with protein-protein interaction and gene expression datasets to facilitate overrepresentation studies and other forms of pathway analysis. Database URL: http://www.reactome.org.

MAPU: Max-Planck Unified database of organellar, cellular, tissue and body fluid proteomes

PubMed Central

Zhang, Yanling; Zhang, Yong; Adachi, Jun; Olsen, Jesper V.; Shi, Rong; de Souza, Gustavo; Pasini, Erica; Foster, Leonard J.; Macek, Boris; Zougman, Alexandre; Kumar, Chanchal; Wiśniewski, Jacek R.; Jun, Wang; Mann, Matthias

2007-01-01

Mass spectrometry (MS)-based proteomics has become a powerful technology to map the protein composition of organelles, cell types and tissues. In our department, a large-scale effort to map these proteomes is complemented by the Max-Planck Unified (MAPU) proteome database. MAPU contains several body fluid proteomes; including plasma, urine, and cerebrospinal fluid. Cell lines have been mapped to a depth of several thousand proteins and the red blood cell proteome has also been analyzed in depth. The liver proteome is represented with 3200 proteins. By employing high resolution MS and stringent validation criteria, false positive identification rates in MAPU are lower than 1:1000. Thus MAPU datasets can serve as reference proteomes in biomarker discovery. MAPU contains the peptides identifying each protein, measured masses, scores and intensities and is freely available at using a clickable interface of cell or body parts. Proteome data can be queried across proteomes by protein name, accession number, sequence similarity, peptide sequence and annotation information. More than 4500 mouse and 2500 human proteins have already been identified in at least one proteome. Basic annotation information and links to other public databases are provided in MAPU and we plan to add further analysis tools. PMID:17090601

FPD: A comprehensive phosphorylation database in fungi.

PubMed

Bai, Youhuang; Chen, Bin; Li, Mingzhu; Zhou, Yincong; Ren, Silin; Xu, Qin; Chen, Ming; Wang, Shihua

2017-10-01

Protein phosphorylation, one of the most classic post-translational modification, plays a critical role in diverse cellular processes including cell cycle, growth, and signal transduction pathways. However, the available information about phosphorylation in fungi is limited. Here, we provided a Fungi Phosphorylation Database (FPD) that comprises high-confidence in vivo phosphosites identified by MS-based proteomics in various fungal species. This comprehensive phosphorylation database contains 62 272 non-redundant phosphorylation sites in 11 222 proteins across eight organisms, including Aspergillus flavus, Aspergillus nidulans, Fusarium graminearum, Magnaporthe oryzae, Neurospora crassa, Saccharomyces cerevisiae, Schizosaccharomyces pombe, and Cryptococcus neoformans. A fungi-specific phosphothreonine motif and several conserved phosphorylation motifs were discovered by comparatively analysing the pattern of phosphorylation sites in plants, animals, and fungi. Copyright © 2017 British Mycological Society. Published by Elsevier Ltd. All rights reserved.

Identification of Fur, Aconitase, and Other Proteins Expressed by Mycobacterium tuberculosis under Conditions of Low and High Concentrations of Iron by Combined Two-Dimensional Gel Electrophoresis and Mass Spectrometry

PubMed Central

Wong, Diane K.; Lee, Bai-Yu; Horwitz, Marcus A.; Gibson, Bradford W.

1999-01-01

Iron plays a critical role in the pathophysiology of Mycobacterium tuberculosis. To gain a better understanding of iron regulation by this organism, we have used two-dimensional (2-D) gel electrophoresis, mass spectrometry, and database searching to study protein expression in M. tuberculosis under conditions of high and low iron concentration. Proteins in cellular extracts from M. tuberculosis Erdman strain grown under low-iron (1 μM) and high-iron (70 μM) conditions were separated by 2-D polyacrylamide gel electrophoresis, which allowed high-resolution separation of several hundred proteins, as visualized by Coomassie staining. The expression of at least 15 proteins was induced, and the expression of at least 12 proteins was decreased under low-iron conditions. In-gel trypsin digestion was performed on these differentially expressed proteins, and the digestion mixtures were analyzed by matrix-assisted laser desorption ionization time-of-flight mass spectrometry to determine the molecular masses of the resulting tryptic peptides. Partial sequence data on some of the peptides were obtained by using after source decay and/or collision-induced dissociation. The fragmentation data were used to search computerized peptide mass and protein sequence databases for known proteins. Ten iron-regulated proteins were identified, including Fur and aconitase proteins, both of which are known to be regulated by iron in other bacterial systems. Our study shows that, where large protein sequence databases are available from genomic studies, the combined use of 2-D gel electrophoresis, mass spectrometry, and database searching to analyze proteins expressed under defined environmental conditions is a powerful tool for identifying expressed proteins and their physiologic relevance. PMID:9864233

The Molecule Pages database

PubMed Central

Saunders, Brian; Lyon, Stephen; Day, Matthew; Riley, Brenda; Chenette, Emily; Subramaniam, Shankar

2008-01-01

The UCSD-Nature Signaling Gateway Molecule Pages (http://www.signaling-gateway.org/molecule) provides essential information on more than 3800 mammalian proteins involved in cellular signaling. The Molecule Pages contain expert-authored and peer-reviewed information based on the published literature, complemented by regularly updated information derived from public data source references and sequence analysis. The expert-authored data includes both a full-text review about the molecule, with citations, and highly structured data for bioinformatics interrogation, including information on protein interactions and states, transitions between states and protein function. The expert-authored pages are anonymously peer reviewed by the Nature Publishing Group. The Molecule Pages data is present in an object-relational database format and is freely accessible to the authors, the reviewers and the public from a web browser that serves as a presentation layer. The Molecule Pages are supported by several applications that along with the database and the interfaces form a multi-tier architecture. The Molecule Pages and the Signaling Gateway are routinely accessed by a very large research community. PMID:17965093

The Molecule Pages database.

PubMed

Saunders, Brian; Lyon, Stephen; Day, Matthew; Riley, Brenda; Chenette, Emily; Subramaniam, Shankar; Vadivelu, Ilango

2008-01-01

The UCSD-Nature Signaling Gateway Molecule Pages (http://www.signaling-gateway.org/molecule) provides essential information on more than 3800 mammalian proteins involved in cellular signaling. The Molecule Pages contain expert-authored and peer-reviewed information based on the published literature, complemented by regularly updated information derived from public data source references and sequence analysis. The expert-authored data includes both a full-text review about the molecule, with citations, and highly structured data for bioinformatics interrogation, including information on protein interactions and states, transitions between states and protein function. The expert-authored pages are anonymously peer reviewed by the Nature Publishing Group. The Molecule Pages data is present in an object-relational database format and is freely accessible to the authors, the reviewers and the public from a web browser that serves as a presentation layer. The Molecule Pages are supported by several applications that along with the database and the interfaces form a multi-tier architecture. The Molecule Pages and the Signaling Gateway are routinely accessed by a very large research community.

Suspended marine particulate proteins in coastal and oligotrophic waters

NASA Astrophysics Data System (ADS)

Bridoux, Maxime C.; Neibauer, Jaqui; Ingalls, Anitra E.; Nunn, Brook L.; Keil, Richard G.

2015-03-01

Metaproteomic analyses were performed on suspended sediments collected in one coastal environment (Washington margin, Pacific Ocean, n = 5) and two oligotrophic environments (Atlantic Ocean near BATS, n = 5, and Pacific Ocean near HOTS, n = 5). Using a database of 2.3 million marine proteins developed using the NCBI database, 443 unique peptides were detected from which 363 unique proteins were identified. Samples from the euphotic zone contained on average 2-3x more identifiable proteins than deeper waters (150-1500 m) and these proteins were predominately from photosynthetic organisms. Diatom peptides dominate the spectra of the Washington margin while peptides from cyanobacteria, such as Synechococcus sp. dominated the spectra of both oligotrophic sites. Despite differences in the exact proteins identified at each location, there is good agreement for protein function and cellular location. Proteins in surface waters code for a variety of cellular functions including photosynthesis (24% of detected proteins), energy production (10%), membrane production (9%) and genetic coding and reading (9%), and are split 60-40 between membrane proteins and intracellular cytoplasmic proteins. Sargasso Sea surface waters contain a suite of peptides consistent with proteins involved in circadian rhythms that promote both C and N fixation at night. At depth in the Sargasso Sea, both muscle-derived myosin protein and the muscle-hydrolyzing proteases deseasin MCP-01 and metalloprotease Mcp02 from γ-proteobacteria were observed. Deeper waters contain peptides predominately sourced from γ-proteobacteria (37% of detected proteins) and α-proteobacteria (26%), although peptides from membrane and photosynthetic proteins attributable to phytoplankton were still observed (13%). Relative to surface values, detection frequencies for bacterial membrane proteins and extracellular enzymes rose from 9 to 16 and 2 to 4% respectively below the thermocline and the overall balance between membrane proteins and intracellular proteins grows to an approximate 75-25 split. Unlike the phytoplankton membrane proteins, which are detrital in nature, the bacterial protein suite at depth is consistent with living biomass.

Proteomic analysis of bovine nucleolus.

PubMed

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

2010-09-01

Nucleolus is the most prominent subnuclear structure, which performs a wide variety of functions in the eukaryotic cellular processes. In order to understand the structural and functional role of the nucleoli in bovine cells, we analyzed the proteomic composition of the bovine nucleoli. The nucleoli were isolated from Madin Darby bovine kidney cells and subjected to proteomic analysis by LC-MS/MS after fractionation by SDS-PAGE and strong cation exchange chromatography. Analysis of the data using the Mascot database search and the GPM database search identified 311 proteins in the bovine nucleoli, which contained 22 proteins previously not identified in the proteomic analysis of human nucleoli. Analysis of the identified proteins using the GoMiner software suggested that the bovine nucleoli contained proteins involved in ribosomal biogenesis, cell cycle control, transcriptional, translational and post-translational regulation, transport, and structural organization. Copyright © 2010 Beijing Genomics Institute. Published by Elsevier Ltd. All rights reserved.

ANCAC: amino acid, nucleotide, and codon analysis of COGs--a tool for sequence bias analysis in microbial orthologs.

PubMed

Meiler, Arno; Klinger, Claudia; Kaufmann, Michael

2012-09-08

The COG database is the most popular collection of orthologous proteins from many different completely sequenced microbial genomes. Per definition, a cluster of orthologous groups (COG) within this database exclusively contains proteins that most likely achieve the same cellular function. Recently, the COG database was extended by assigning to every protein both the corresponding amino acid and its encoding nucleotide sequence resulting in the NUCOCOG database. This extended version of the COG database is a valuable resource connecting sequence features with the functionality of the respective proteins. Here we present ANCAC, a web tool and MySQL database for the analysis of amino acid, nucleotide, and codon frequencies in COGs on the basis of freely definable phylogenetic patterns. We demonstrate the usefulness of ANCAC by analyzing amino acid frequencies, codon usage, and GC-content in a species- or function-specific context. With respect to amino acids we, at least in part, confirm the cognate bias hypothesis by using ANCAC's NUCOCOG dataset as the largest one available for that purpose thus far. Using the NUCOCOG datasets, ANCAC connects taxonomic, amino acid, and nucleotide sequence information with the functional classification via COGs and provides a GUI for flexible mining for sequence-bias. Thereby, to our knowledge, it is the only tool for the analysis of sequence composition in the light of physiological roles and phylogenetic context without requirement of substantial programming-skills.

ANCAC: amino acid, nucleotide, and codon analysis of COGs – a tool for sequence bias analysis in microbial orthologs

PubMed Central

2012-01-01

Background The COG database is the most popular collection of orthologous proteins from many different completely sequenced microbial genomes. Per definition, a cluster of orthologous groups (COG) within this database exclusively contains proteins that most likely achieve the same cellular function. Recently, the COG database was extended by assigning to every protein both the corresponding amino acid and its encoding nucleotide sequence resulting in the NUCOCOG database. This extended version of the COG database is a valuable resource connecting sequence features with the functionality of the respective proteins. Results Here we present ANCAC, a web tool and MySQL database for the analysis of amino acid, nucleotide, and codon frequencies in COGs on the basis of freely definable phylogenetic patterns. We demonstrate the usefulness of ANCAC by analyzing amino acid frequencies, codon usage, and GC-content in a species- or function-specific context. With respect to amino acids we, at least in part, confirm the cognate bias hypothesis by using ANCAC’s NUCOCOG dataset as the largest one available for that purpose thus far. Conclusions Using the NUCOCOG datasets, ANCAC connects taxonomic, amino acid, and nucleotide sequence information with the functional classification via COGs and provides a GUI for flexible mining for sequence-bias. Thereby, to our knowledge, it is the only tool for the analysis of sequence composition in the light of physiological roles and phylogenetic context without requirement of substantial programming-skills. PMID:22958836

Differential Proteome Analysis of a Flor Yeast Strain under Biofilm Formation.

PubMed

Moreno-García, Jaime; Mauricio, Juan Carlos; Moreno, Juan; García-Martínez, Teresa

2017-03-28

Several Saccharomyces cerevisiae strains (flor yeasts) form a biofilm (flor velum) on the surface of Sherry wines after fermentation, when glucose is depleted. This flor velum is fundamental to biological aging of these particular wines. In this study, we identify abundant proteins in the formation of the biofilm of an industrial flor yeast strain. A database search to enrich flor yeast "biological process" and "cellular component" according to Gene Ontology Terminology (GO Terms) and, "pathways" was carried out. The most abundant proteins detected were largely involved in respiration, translation, stress damage prevention and repair, amino acid metabolism (glycine, isoleucine, leucine and arginine), glycolysis/gluconeogenesis and biosynthesis of vitamin B9 (folate). These proteins were located in cellular components as in the peroxisome, mitochondria, vacuole, cell wall and extracellular region; being these two last directly related with the flor formation. Proteins like Bgl2p, Gcv3p, Hyp2p, Mdh1p, Suc2p and Ygp1p were quantified in very high levels. This study reveals some expected processes and provides new and important information for the design of conditions and genetic constructions of flor yeasts for improving the cellular survival and, thus, to optimize biological aging of Sherry wine production.

CellMap visualizes protein-protein interactions and subcellular localization

PubMed Central

Dallago, Christian; Goldberg, Tatyana; Andrade-Navarro, Miguel Angel; Alanis-Lobato, Gregorio; Rost, Burkhard

2018-01-01

Many tools visualize protein-protein interaction (PPI) networks. The tool introduced here, CellMap, adds one crucial novelty by visualizing PPI networks in the context of subcellular localization, i.e. the location in the cell or cellular component in which a PPI happens. Users can upload images of cells and define areas of interest against which PPIs for selected proteins are displayed (by default on a cartoon of a cell). Annotations of localization are provided by the user or through our in-house database. The visualizer and server are written in JavaScript, making CellMap easy to customize and to extend by researchers and developers. PMID:29497493

Exploring Short Linear Motifs Using the ELM Database and Tools.

PubMed

Gouw, Marc; Sámano-Sánchez, Hugo; Van Roey, Kim; Diella, Francesca; Gibson, Toby J; Dinkel, Holger

2017-06-27

The Eukaryotic Linear Motif (ELM) resource is dedicated to the characterization and prediction of short linear motifs (SLiMs). SLiMs are compact, degenerate peptide segments found in many proteins and essential to almost all cellular processes. However, despite their abundance, SLiMs remain largely uncharacterized. The ELM database is a collection of manually annotated SLiM instances curated from experimental literature. In this article we illustrate how to browse and search the database for curated SLiM data, and cover the different types of data integrated in the resource. We also cover how to use this resource in order to predict SLiMs in known as well as novel proteins, and how to interpret the results generated by the ELM prediction pipeline. The ELM database is a very rich resource, and in the following protocols we give helpful examples to demonstrate how this knowledge can be used to improve your own research. © 2017 by John Wiley & Sons, Inc. Copyright © 2017 John Wiley & Sons, Inc.

CPLA 1.0: an integrated database of protein lysine acetylation

PubMed Central

Liu, Zexian; Cao, Jun; Gao, Xinjiao; Zhou, Yanhong; Wen, Longping; Yang, Xiangjiao; Yao, Xuebiao; Ren, Jian; Xue, Yu

2011-01-01

As a reversible post-translational modification (PTM) discovered decades ago, protein lysine acetylation was known for its regulation of transcription through the modification of histones. Recent studies discovered that lysine acetylation targets broad substrates and especially plays an essential role in cellular metabolic regulation. Although acetylation is comparable with other major PTMs such as phosphorylation, an integrated resource still remains to be developed. In this work, we presented the compendium of protein lysine acetylation (CPLA) database for lysine acetylated substrates with their sites. From the scientific literature, we manually collected 7151 experimentally identified acetylation sites in 3311 targets. We statistically studied the regulatory roles of lysine acetylation by analyzing the Gene Ontology (GO) and InterPro annotations. Combined with protein–protein interaction information, we systematically discovered a potential human lysine acetylation network (HLAN) among histone acetyltransferases (HATs), substrates and histone deacetylases (HDACs). In particular, there are 1862 triplet relationships of HAT-substrate-HDAC retrieved from the HLAN, at least 13 of which were previously experimentally verified. The online services of CPLA database was implemented in PHP + MySQL + JavaScript, while the local packages were developed in JAVA 1.5 (J2SE 5.0). The CPLA database is freely available for all users at: http://cpla.biocuckoo.org. PMID:21059677

Comparative bioinformatics analyses and profiling of lysosome-related organelle proteomes

NASA Astrophysics Data System (ADS)

Hu, Zhang-Zhi; Valencia, Julio C.; Huang, Hongzhan; Chi, An; Shabanowitz, Jeffrey; Hearing, Vincent J.; Appella, Ettore; Wu, Cathy

2007-01-01

Complete and accurate profiling of cellular organelle proteomes, while challenging, is important for the understanding of detailed cellular processes at the organelle level. Mass spectrometry technologies coupled with bioinformatics analysis provide an effective approach for protein identification and functional interpretation of organelle proteomes. In this study, we have compiled human organelle reference datasets from large-scale proteomic studies and protein databases for seven lysosome-related organelles (LROs), as well as the endoplasmic reticulum and mitochondria, for comparative organelle proteome analysis. Heterogeneous sources of human organelle proteins and rodent homologs are mapped to human UniProtKB protein entries based on ID and/or peptide mappings, followed by functional annotation and categorization using the iProXpress proteomic expression analysis system. Cataloging organelle proteomes allows close examination of both shared and unique proteins among various LROs and reveals their functional relevance. The proteomic comparisons show that LROs are a closely related family of organelles. The shared proteins indicate the dynamic and hybrid nature of LROs, while the unique transmembrane proteins may represent additional candidate marker proteins for LROs. This comparative analysis, therefore, provides a basis for hypothesis formulation and experimental validation of organelle proteins and their functional roles.

MAPU: Max-Planck Unified database of organellar, cellular, tissue and body fluid proteomes.

PubMed

Zhang, Yanling; Zhang, Yong; Adachi, Jun; Olsen, Jesper V; Shi, Rong; de Souza, Gustavo; Pasini, Erica; Foster, Leonard J; Macek, Boris; Zougman, Alexandre; Kumar, Chanchal; Wisniewski, Jacek R; Jun, Wang; Mann, Matthias

2007-01-01

Mass spectrometry (MS)-based proteomics has become a powerful technology to map the protein composition of organelles, cell types and tissues. In our department, a large-scale effort to map these proteomes is complemented by the Max-Planck Unified (MAPU) proteome database. MAPU contains several body fluid proteomes; including plasma, urine, and cerebrospinal fluid. Cell lines have been mapped to a depth of several thousand proteins and the red blood cell proteome has also been analyzed in depth. The liver proteome is represented with 3200 proteins. By employing high resolution MS and stringent validation criteria, false positive identification rates in MAPU are lower than 1:1000. Thus MAPU datasets can serve as reference proteomes in biomarker discovery. MAPU contains the peptides identifying each protein, measured masses, scores and intensities and is freely available at http://www.mapuproteome.com using a clickable interface of cell or body parts. Proteome data can be queried across proteomes by protein name, accession number, sequence similarity, peptide sequence and annotation information. More than 4500 mouse and 2500 human proteins have already been identified in at least one proteome. Basic annotation information and links to other public databases are provided in MAPU and we plan to add further analysis tools.

Constructing Proteome Reference Map of the Porcine Jejunal Cell Line (IPEC-J2) by Label-Free Mass Spectrometry.

PubMed

Kim, Sang Hoon; Pajarillo, Edward Alain B; Balolong, Marilen P; Lee, Ji Yoon; Kang, Dae-Kyung

2016-06-28

In this study, the global proteome of the IPEC-J2 cell line was evaluated using ultra-high performance liquid chromatography coupled to a quadrupole Q Exactive™ Orbitrap mass spectrometer. Proteins were isolated from highly confluent IPEC-J2 cells in biological replicates and analyzed by label-free mass spectrometry prior to matching against a porcine genomic dataset. The results identified 1,517 proteins, accounting for 7.35% of all genes in the porcine genome. The highly abundant proteins detected, such as actin, annexin A2, and AHNAK nucleoprotein, are involved in structural integrity, signaling mechanisms, and cellular homeostasis. The high abundance of heat shock proteins indicated their significance in cellular defenses, barrier function, and gut homeostasis. Pathway analysis and annotation using the Kyoto Encyclopedia of Genes and Genomes database resulted in a putative protein network map of the regulation of immunological responses and structural integrity in the cell line. The comprehensive proteome analysis of IPEC-J2 cells provides fundamental insights into overall protein expression and pathway dynamics that might be useful in cell adhesion studies and immunological applications.

RAID: a comprehensive resource for human RNA-associated (RNA–RNA/RNA–protein) interaction

PubMed Central

Zhang, Xiaomeng; Wu, Deng; Chen, Liqun; Li, Xiang; Yang, Jinxurong; Fan, Dandan; Dong, Tingting; Liu, Mingyue; Tan, Puwen; Xu, Jintian; Yi, Ying; Wang, Yuting; Zou, Hua; Hu, Yongfei; Fan, Kaili; Kang, Juanjuan; Huang, Yan; Miao, Zhengqiang; Bi, Miaoman; Jin, Nana; Li, Kongning; Li, Xia; Xu, Jianzhen; Wang, Dong

2014-01-01

Transcriptomic analyses have revealed an unexpected complexity in the eukaryote transcriptome, which includes not only protein-coding transcripts but also an expanding catalog of noncoding RNAs (ncRNAs). Diverse coding and noncoding RNAs (ncRNAs) perform functions through interaction with each other in various cellular processes. In this project, we have developed RAID (http://www.rna-society.org/raid), an RNA-associated (RNA–RNA/RNA–protein) interaction database. RAID intends to provide the scientific community with all-in-one resources for efficient browsing and extraction of the RNA-associated interactions in human. This version of RAID contains more than 6100 RNA-associated interactions obtained by manually reviewing more than 2100 published papers, including 4493 RNA–RNA interactions and 1619 RNA–protein interactions. Each entry contains detailed information on an RNA-associated interaction, including RAID ID, RNA/protein symbol, RNA/protein categories, validated method, expressing tissue, literature references (Pubmed IDs), and detailed functional description. Users can query, browse, analyze, and manipulate RNA-associated (RNA–RNA/RNA–protein) interaction. RAID provides a comprehensive resource of human RNA-associated (RNA–RNA/RNA–protein) interaction network. Furthermore, this resource will help in uncovering the generic organizing principles of cellular function network. PMID:24803509

PCoM-DB Update: A Protein Co-Migration Database for Photosynthetic Organisms.

PubMed

Takabayashi, Atsushi; Takabayashi, Saeka; Takahashi, Kaori; Watanabe, Mai; Uchida, Hiroko; Murakami, Akio; Fujita, Tomomichi; Ikeuchi, Masahiko; Tanaka, Ayumi

2017-01-01

The identification of protein complexes is important for the understanding of protein structure and function and the regulation of cellular processes. We used blue-native PAGE and tandem mass spectrometry to identify protein complexes systematically, and built a web database, the protein co-migration database (PCoM-DB, http://pcomdb.lowtem.hokudai.ac.jp/proteins/top), to provide prediction tools for protein complexes. PCoM-DB provides migration profiles for any given protein of interest, and allows users to compare them with migration profiles of other proteins, showing the oligomeric states of proteins and thus identifying potential interaction partners. The initial version of PCoM-DB (launched in January 2013) included protein complex data for Synechocystis whole cells and Arabidopsis thaliana thylakoid membranes. Here we report PCoM-DB version 2.0, which includes new data sets and analytical tools. Additional data are included from whole cells of the pelagic marine picocyanobacterium Prochlorococcus marinus, the thermophilic cyanobacterium Thermosynechococcus elongatus, the unicellular green alga Chlamydomonas reinhardtii and the bryophyte Physcomitrella patens. The Arabidopsis protein data now include data for intact mitochondria, intact chloroplasts, chloroplast stroma and chloroplast envelopes. The new tools comprise a multiple-protein search form and a heat map viewer for protein migration profiles. Users can compare migration profiles of a protein of interest among different organelles or compare migration profiles among different proteins within the same sample. For Arabidopsis proteins, users can compare migration profiles of a protein of interest with putative homologous proteins from non-Arabidopsis organisms. The updated PCoM-DB will help researchers find novel protein complexes and estimate their evolutionary changes in the green lineage. © The Author 2017. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.

Analysis of Sir2E in the cellular slime mold Dictyostelium discoideum: cellular localization, spatial expression and overexpression.

PubMed

Katayama, Takahiro; Yasukawa, Hiro

2008-10-01

It has been reported that Dictyostelium discoideum encodes four silent information regulator 2 (Sir2) proteins (Sir2A-D) showing sequence similarity to human homologues of Sir2 (SIRT1-3). Further screening in a database revealed that D. discoideum encodes an additional Sir2 homologue (Sir2E). The amino acid sequence of Sir2E is not similar to those of SIRTs but is similar to those of proteins encoded by Giardia lamblia, Cryptosporidium hominis and Cryptosporidium parvum. Fluorescence of Sir2E-green fluorescent protein fusion protein was detected in the D. discoideum nucleus, indicating that Sir2E is a nuclear localizing protein. Reverse transcription-polymerase chain reaction and whole-mount in situ hybridization analyses showed that D. discoideum expressed sir2E in amoebae in the growth phase and in prestalk cells in the developmental phase. D. discoideum overexpressing sir2E grew faster than the wild type. These results indicate that Sir2E plays important roles both in the growth phase and developmental phase of D. discoideum.

Cytoscape: a software environment for integrated models of biomolecular interaction networks.

PubMed

Shannon, Paul; Markiel, Andrew; Ozier, Owen; Baliga, Nitin S; Wang, Jonathan T; Ramage, Daniel; Amin, Nada; Schwikowski, Benno; Ideker, Trey

2003-11-01

Cytoscape is an open source software project for integrating biomolecular interaction networks with high-throughput expression data and other molecular states into a unified conceptual framework. Although applicable to any system of molecular components and interactions, Cytoscape is most powerful when used in conjunction with large databases of protein-protein, protein-DNA, and genetic interactions that are increasingly available for humans and model organisms. Cytoscape's software Core provides basic functionality to layout and query the network; to visually integrate the network with expression profiles, phenotypes, and other molecular states; and to link the network to databases of functional annotations. The Core is extensible through a straightforward plug-in architecture, allowing rapid development of additional computational analyses and features. Several case studies of Cytoscape plug-ins are surveyed, including a search for interaction pathways correlating with changes in gene expression, a study of protein complexes involved in cellular recovery to DNA damage, inference of a combined physical/functional interaction network for Halobacterium, and an interface to detailed stochastic/kinetic gene regulatory models.

Proteomic analysis of tardigrades: towards a better understanding of molecular mechanisms by anhydrobiotic organisms.

PubMed

Schokraie, Elham; Hotz-Wagenblatt, Agnes; Warnken, Uwe; Mali, Brahim; Frohme, Marcus; Förster, Frank; Dandekar, Thomas; Hengherr, Steffen; Schill, Ralph O; Schnölzer, Martina

2010-03-03

Tardigrades are small, multicellular invertebrates which are able to survive times of unfavourable environmental conditions using their well-known capability to undergo cryptobiosis at any stage of their life cycle. Milnesium tardigradum has become a powerful model system for the analysis of cryptobiosis. While some genetic information is already available for Milnesium tardigradum the proteome is still to be discovered. Here we present to the best of our knowledge the first comprehensive study of Milnesium tardigradum on the protein level. To establish a proteome reference map we developed optimized protocols for protein extraction from tardigrades in the active state and for separation of proteins by high resolution two-dimensional gel electrophoresis. Since only limited sequence information of M. tardigradum on the genome and gene expression level is available to date in public databases we initiated in parallel a tardigrade EST sequencing project to allow for protein identification by electrospray ionization tandem mass spectrometry. 271 out of 606 analyzed protein spots could be identified by searching against the publicly available NCBInr database as well as our newly established tardigrade protein database corresponding to 144 unique proteins. Another 150 spots could be identified in the tardigrade clustered EST database corresponding to 36 unique contigs and ESTs. Proteins with annotated function were further categorized in more detail by their molecular function, biological process and cellular component. For the proteins of unknown function more information could be obtained by performing a protein domain annotation analysis. Our results include proteins like protein member of different heat shock protein families and LEA group 3, which might play important roles in surviving extreme conditions. The proteome reference map of Milnesium tardigradum provides the basis for further studies in order to identify and characterize the biochemical mechanisms of tolerance to extreme desiccation. The optimized proteomics workflow will enable application of sensitive quantification techniques to detect differences in protein expression, which are characteristic of the active and anhydrobiotic states of tardigrades.

Proteomic Analysis of Tardigrades: Towards a Better Understanding of Molecular Mechanisms by Anhydrobiotic Organisms

PubMed Central

Schokraie, Elham; Hotz-Wagenblatt, Agnes; Warnken, Uwe; Mali, Brahim; Frohme, Marcus; Förster, Frank; Dandekar, Thomas; Hengherr, Steffen; Schill, Ralph O.; Schnölzer, Martina

2010-01-01

Background Tardigrades are small, multicellular invertebrates which are able to survive times of unfavourable environmental conditions using their well-known capability to undergo cryptobiosis at any stage of their life cycle. Milnesium tardigradum has become a powerful model system for the analysis of cryptobiosis. While some genetic information is already available for Milnesium tardigradum the proteome is still to be discovered. Principal Findings Here we present to the best of our knowledge the first comprehensive study of Milnesium tardigradum on the protein level. To establish a proteome reference map we developed optimized protocols for protein extraction from tardigrades in the active state and for separation of proteins by high resolution two-dimensional gel electrophoresis. Since only limited sequence information of M. tardigradum on the genome and gene expression level is available to date in public databases we initiated in parallel a tardigrade EST sequencing project to allow for protein identification by electrospray ionization tandem mass spectrometry. 271 out of 606 analyzed protein spots could be identified by searching against the publicly available NCBInr database as well as our newly established tardigrade protein database corresponding to 144 unique proteins. Another 150 spots could be identified in the tardigrade clustered EST database corresponding to 36 unique contigs and ESTs. Proteins with annotated function were further categorized in more detail by their molecular function, biological process and cellular component. For the proteins of unknown function more information could be obtained by performing a protein domain annotation analysis. Our results include proteins like protein member of different heat shock protein families and LEA group 3, which might play important roles in surviving extreme conditions. Conclusions The proteome reference map of Milnesium tardigradum provides the basis for further studies in order to identify and characterize the biochemical mechanisms of tolerance to extreme desiccation. The optimized proteomics workflow will enable application of sensitive quantification techniques to detect differences in protein expression, which are characteristic of the active and anhydrobiotic states of tardigrades. PMID:20224743

Differential Proteome Analysis of a Flor Yeast Strain under Biofilm Formation

PubMed Central

Moreno-García, Jaime; Mauricio, Juan Carlos; Moreno, Juan; García-Martínez, Teresa

2017-01-01

Several Saccharomyces cerevisiae strains (flor yeasts) form a biofilm (flor velum) on the surface of Sherry wines after fermentation, when glucose is depleted. This flor velum is fundamental to biological aging of these particular wines. In this study, we identify abundant proteins in the formation of the biofilm of an industrial flor yeast strain. A database search to enrich flor yeast “biological process” and “cellular component” according to Gene Ontology Terminology (GO Terms) and, “pathways” was carried out. The most abundant proteins detected were largely involved in respiration, translation, stress damage prevention and repair, amino acid metabolism (glycine, isoleucine, leucine and arginine), glycolysis/gluconeogenesis and biosynthesis of vitamin B9 (folate). These proteins were located in cellular components as in the peroxisome, mitochondria, vacuole, cell wall and extracellular region; being these two last directly related with the flor formation. Proteins like Bgl2p, Gcv3p, Hyp2p, Mdh1p, Suc2p and Ygp1p were quantified in very high levels. This study reveals some expected processes and provides new and important information for the design of conditions and genetic constructions of flor yeasts for improving the cellular survival and, thus, to optimize biological aging of Sherry wine production. PMID:28350350

Regulatory RNA binding proteins contribute to the transcriptome-wide splicing alterations in human cellular senescence.

PubMed

Dong, Qiongye; Wei, Lei; Zhang, Michael Q; Wang, Xiaowo

2018-06-24

Dysregulation of mRNA splicing has been observed in certain cellular senescence process. However, the common splicing alterations on the whole transcriptome shared by various types of senescence are poorly understood. In order to systematically identify senescence-associated transcriptomic changes in genome-wide scale, we collected RNA sequencing datasets of different human cell types with a variety of senescence-inducing methods from public databases and performed meta-analysis. First, we discovered that a group of RNA binding proteins were consistently down-regulated in diverse senescent samples and identified 406 senescence-associated common differential splicing events. Then, eight differentially expressed RNA binding proteins were predicted to regulate these senescence-associated splicing alterations through an enrichment analysis of their RNA binding information, including motif scanning and enhanced cross-linking immunoprecipitation data. In addition, we constructed the splicing regulatory modules that might contribute to senescence-associated biological processes. Finally, it was confirmed that knockdown of the predicted senescence-associated potential splicing regulators through shRNAs in HepG2 cell line could result in senescence-like splicing changes. Taken together, our work demonstrated a broad range of common changes in mRNA splicing switches and detected their central regulatory RNA binding proteins during senescence. These findings would help to better understand the coordinating splicing alterations in cellular senescence.

Protein-protein interaction networks: unraveling the wiring of molecular machines within the cell.

PubMed

De Las Rivas, Javier; Fontanillo, Celia

2012-11-01

Mapping and understanding of the protein interaction networks with their key modules and hubs can provide deeper insights into the molecular machinery underlying complex phenotypes. In this article, we present the basic characteristics and definitions of protein networks, starting with a distinction of the different types of associations between proteins. We focus the review on protein-protein interactions (PPIs), a subset of associations defined as physical contacts between proteins that occur by selective molecular docking in a particular biological context. We present such definition as opposed to other types of protein associations derived from regulatory, genetic, structural or functional relations. To determine PPIs, a variety of binary and co-complex methods exist; however, not all the technologies provide the same information and data quality. A way of increasing confidence in a given protein interaction is to integrate orthogonal experimental evidences. The use of several complementary methods testing each single interaction assesses the accuracy of PPI data and tries to minimize the occurrence of false interactions. Following this approach there have been important efforts to unify primary databases of experimentally proven PPIs into integrated databases. These meta-databases provide a measure of the confidence of interactions based on the number of experimental proofs that report them. As a conclusion, we can state that integrated information allows the building of more reliable interaction networks. Identification of communities, cliques, modules and hubs by analysing the topological parameters and graph properties of the protein networks allows the discovery of central/critical nodes, which are candidates to regulate cellular flux and dynamics.

Discovery of small molecules binding to the normal conformation of prion by combining virtual screening and multiple biological activity evaluation methods

NASA Astrophysics Data System (ADS)

Li, Lanlan; Wei, Wei; Jia, Wen-Juan; Zhu, Yongchang; Zhang, Yan; Chen, Jiang-Huai; Tian, Jiaqi; Liu, Huanxiang; He, Yong-Xing; Yao, Xiaojun

2017-12-01

Conformational conversion of the normal cellular prion protein, PrPC, into the misfolded isoform, PrPSc, is considered to be a central event in the development of fatal neurodegenerative diseases. Stabilization of prion protein at the normal cellular form (PrPC) with small molecules is a rational and efficient strategy for treatment of prion related diseases. However, few compounds have been identified as potent prion inhibitors by binding to the normal conformation of prion. In this work, to rational screening of inhibitors capable of stabilizing cellular form of prion protein, multiple approaches combining docking-based virtual screening, steady-state fluorescence quenching, surface plasmon resonance and thioflavin T fluorescence assay were used to discover new compounds interrupting PrPC to PrPSc conversion. Compound 3253-0207 that can bind to PrPC with micromolar affinity and inhibit prion fibrillation was identified from small molecule databases. Molecular dynamics simulation indicated that compound 3253-0207 can bind to the hotspot residues in the binding pocket composed by β1, β2 and α2, which are significant structure moieties in conversion from PrPC to PrPSc.

Computational approaches for de novo design and redesign of metal-binding sites on proteins.

PubMed

Akcapinar, Gunseli Bayram; Sezerman, Osman Ugur

2017-04-28

Metal ions play pivotal roles in protein structure, function and stability. The functional and structural diversity of proteins in nature expanded with the incorporation of metal ions or clusters in proteins. Approximately one-third of these proteins in the databases contain metal ions. Many biological and chemical processes in nature involve metal ion-binding proteins, aka metalloproteins. Many cellular reactions that underpin life require metalloproteins. Most of the remarkable, complex chemical transformations are catalysed by metalloenzymes. Realization of the importance of metal-binding sites in a variety of cellular events led to the advancement of various computational methods for their prediction and characterization. Furthermore, as structural and functional knowledgebase about metalloproteins is expanding with advances in computational and experimental fields, the focus of the research is now shifting towards de novo design and redesign of metalloproteins to extend nature's own diversity beyond its limits. In this review, we will focus on the computational toolbox for prediction of metal ion-binding sites, de novo metalloprotein design and redesign. We will also give examples of tailor-made artificial metalloproteins designed with the computational toolbox. © 2017 The Author(s).

Proteomic analysis of the defense response of wheat to the powdery mildew fungus, Blumeria graminis f. sp. tritici.

PubMed

Mandal, Md Siddikun Nabi; Fu, Ying; Zhang, Sheng; Ji, Wanquan

2014-12-01

Powdery mildew of wheat is caused by Blumeria graminis f. sp. tritici (Bgt). Although many wheat cultivars resistant to this disease have been developed, little is known about their resistance mechanisms. The aim of this study was to identify proteins showing changes in abundance during the resistance response of the wheat line N0308 infected by Bgt. In two-dimensional electrophoresis analyses, 45 spots on the gels showed significant changes in abundance at 24, 48, and 72 h after inoculation, as compared to non-inoculated plants. Of these 45 proteins, 44 were identified by mass spectrometry analysis using the NCBInr database of Triticum aestivum (26 spots) and closely related species in the Triticum genus (18 spots). These proteins were associated with the defense response, photosynthesis, metabolism, and other cellular processes in wheat. Most of the up-regulated proteins were identified as stress- and defense-related proteins. In particular, the product of a specific powdery mildew resistance gene (Pm3b and its homolog) and some other defense- and pathogenesis-related proteins were overexpressed. The resistance gene product mediates the immune response and coordinates other cellular processes during the resistance response to Bgt.

Multiparametric Determination of Radiation Risk

NASA Technical Reports Server (NTRS)

Richmond, Robert C.

2003-01-01

Predicting risk of human cancer following exposure to ionizing space radiation is challenging in part because of uncertainties of low-dose distribution amongst cells, of unknown potentially synergistic effects of microgravity upon cellular protein-expression, and of processing dose-related damage within cells to produce rare and late-appearing malignant transformation, degrade the confidence of cancer risk-estimates. The NASA- specific responsibility to estimate the risks of radiogenic cancer in a limited number of astronauts is not amenable to epidemiologic study, thereby increasing this challenge. Developing adequately sensitive cellular biodosimeters that simultaneously report 1) the quantity of absorbed close after exposure to ionizing radiation, 2) the quality of radiation delivering that dose, and 3) the risk of developing malignant transformation by the cells absorbing that dose could be useful for resolving these challenges. Use of a multiparametric cellular biodosimeter is suggested using analyses of gene-expression and protein-expression whereby large datasets of cellular response to radiation-induced damage are obtained and analyzed for expression-profiles correlated with established end points and molecular markers predictive for cancer-risk. Analytical techniques of genomics and proteomics may be used to establish dose-dependency of multiple gene- and protein- expressions resulting from radiation-induced cellular damage. Furthermore, gene- and protein-expression from cells in microgravity are known to be altered relative to cells grown on the ground at 1g. Therefore, hypotheses are proposed that 1) macromolecular expression caused by radiation-induced damage in cells in microgravity may be different than on the ground, and 2) different patterns of macromolecular expression in microgravity may alter human radiogenic cancer risk relative to radiation exposure on Earth. A new paradigm is accordingly suggested as a national database wherein genomic and proteomic datasets are registered and interrogated in order to provide statistically significant dose-dependent risk estimation of radiogenic cancer in astronauts.

Functional discovery via a compendium of expression profiles.

PubMed

Hughes, T R; Marton, M J; Jones, A R; Roberts, C J; Stoughton, R; Armour, C D; Bennett, H A; Coffey, E; Dai, H; He, Y D; Kidd, M J; King, A M; Meyer, M R; Slade, D; Lum, P Y; Stepaniants, S B; Shoemaker, D D; Gachotte, D; Chakraburtty, K; Simon, J; Bard, M; Friend, S H

2000-07-07

Ascertaining the impact of uncharacterized perturbations on the cell is a fundamental problem in biology. Here, we describe how a single assay can be used to monitor hundreds of different cellular functions simultaneously. We constructed a reference database or "compendium" of expression profiles corresponding to 300 diverse mutations and chemical treatments in S. cerevisiae, and we show that the cellular pathways affected can be determined by pattern matching, even among very subtle profiles. The utility of this approach is validated by examining profiles caused by deletions of uncharacterized genes: we identify and experimentally confirm that eight uncharacterized open reading frames encode proteins required for sterol metabolism, cell wall function, mitochondrial respiration, or protein synthesis. We also show that the compendium can be used to characterize pharmacological perturbations by identifying a novel target of the commonly used drug dyclonine.

RAID: a comprehensive resource for human RNA-associated (RNA-RNA/RNA-protein) interaction.

PubMed

Zhang, Xiaomeng; Wu, Deng; Chen, Liqun; Li, Xiang; Yang, Jinxurong; Fan, Dandan; Dong, Tingting; Liu, Mingyue; Tan, Puwen; Xu, Jintian; Yi, Ying; Wang, Yuting; Zou, Hua; Hu, Yongfei; Fan, Kaili; Kang, Juanjuan; Huang, Yan; Miao, Zhengqiang; Bi, Miaoman; Jin, Nana; Li, Kongning; Li, Xia; Xu, Jianzhen; Wang, Dong

2014-07-01

Transcriptomic analyses have revealed an unexpected complexity in the eukaryote transcriptome, which includes not only protein-coding transcripts but also an expanding catalog of noncoding RNAs (ncRNAs). Diverse coding and noncoding RNAs (ncRNAs) perform functions through interaction with each other in various cellular processes. In this project, we have developed RAID (http://www.rna-society.org/raid), an RNA-associated (RNA-RNA/RNA-protein) interaction database. RAID intends to provide the scientific community with all-in-one resources for efficient browsing and extraction of the RNA-associated interactions in human. This version of RAID contains more than 6100 RNA-associated interactions obtained by manually reviewing more than 2100 published papers, including 4493 RNA-RNA interactions and 1619 RNA-protein interactions. Each entry contains detailed information on an RNA-associated interaction, including RAID ID, RNA/protein symbol, RNA/protein categories, validated method, expressing tissue, literature references (Pubmed IDs), and detailed functional description. Users can query, browse, analyze, and manipulate RNA-associated (RNA-RNA/RNA-protein) interaction. RAID provides a comprehensive resource of human RNA-associated (RNA-RNA/RNA-protein) interaction network. Furthermore, this resource will help in uncovering the generic organizing principles of cellular function network. © 2014 Zhang et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society.

Carbohydrate Structure Database: tools for statistical analysis of bacterial, plant and fungal glycomes

PubMed Central

Egorova, K.S.; Kondakova, A.N.; Toukach, Ph.V.

2015-01-01

Carbohydrates are biological blocks participating in diverse and crucial processes both at cellular and organism levels. They protect individual cells, establish intracellular interactions, take part in the immune reaction and participate in many other processes. Glycosylation is considered as one of the most important modifications of proteins and other biologically active molecules. Still, the data on the enzymatic machinery involved in the carbohydrate synthesis and processing are scattered, and the advance on its study is hindered by the vast bulk of accumulated genetic information not supported by any experimental evidences for functions of proteins that are encoded by these genes. In this article, we present novel instruments for statistical analysis of glycomes in taxa. These tools may be helpful for investigating carbohydrate-related enzymatic activities in various groups of organisms and for comparison of their carbohydrate content. The instruments are developed on the Carbohydrate Structure Database (CSDB) platform and are available freely on the CSDB web-site at http://csdb.glycoscience.ru. Database URL: http://csdb.glycoscience.ru PMID:26337239

Wheat proteomics: proteome modulation and abiotic stress acclimation

PubMed Central

Komatsu, Setsuko; Kamal, Abu H. M.; Hossain, Zahed

2014-01-01

Cellular mechanisms of stress sensing and signaling represent the initial plant responses to adverse conditions. The development of high-throughput “Omics” techniques has initiated a new era of the study of plant molecular strategies for adapting to environmental changes. However, the elucidation of stress adaptation mechanisms in plants requires the accurate isolation and characterization of stress-responsive proteins. Because the functional part of the genome, namely the proteins and their post-translational modifications, are critical for plant stress responses, proteomic studies provide comprehensive information about the fine-tuning of cellular pathways that primarily involved in stress mitigation. This review summarizes the major proteomic findings related to alterations in the wheat proteomic profile in response to abiotic stresses. Moreover, the strengths and weaknesses of different sample preparation techniques, including subcellular protein extraction protocols, are discussed in detail. The continued development of proteomic approaches in combination with rapidly evolving bioinformatics tools and interactive databases will facilitate understanding of the plant mechanisms underlying stress tolerance. PMID:25538718

Predicting Protein Relationships to Human Pathways through a Relational Learning Approach Based on Simple Sequence Features.

PubMed

García-Jiménez, Beatriz; Pons, Tirso; Sanchis, Araceli; Valencia, Alfonso

2014-01-01

Biological pathways are important elements of systems biology and in the past decade, an increasing number of pathway databases have been set up to document the growing understanding of complex cellular processes. Although more genome-sequence data are becoming available, a large fraction of it remains functionally uncharacterized. Thus, it is important to be able to predict the mapping of poorly annotated proteins to original pathway models. We have developed a Relational Learning-based Extension (RLE) system to investigate pathway membership through a function prediction approach that mainly relies on combinations of simple properties attributed to each protein. RLE searches for proteins with molecular similarities to specific pathway components. Using RLE, we associated 383 uncharacterized proteins to 28 pre-defined human Reactome pathways, demonstrating relative confidence after proper evaluation. Indeed, in specific cases manual inspection of the database annotations and the related literature supported the proposed classifications. Examples of possible additional components of the Electron transport system, Telomere maintenance and Integrin cell surface interactions pathways are discussed in detail. All the human predicted proteins in the 2009 and 2012 releases 30 and 40 of Reactome are available at http://rle.bioinfo.cnio.es.

Bioinformatics analysis on molecular mechanism of rheum officinale in treatment of jaundice

NASA Astrophysics Data System (ADS)

Shan, Si; Tu, Jun; Nie, Peng; Yan, Xiaojun

2017-01-01

Objective: To study the molecular mechanism of Rheum officinale in the treatment of Jaundice by building molecular networks and comparing canonical pathways. Methods: Target proteins of Rheum officinale and related genes of Jaundice were searched from Pubchem and Gene databases online respectively. Molecular networks and canonical pathways comparison analyses were performed by Ingenuity Pathway Analysis (IPA). Results: The molecular networks of Rheum officinale and Jaundice were complex and multifunctional. The 40 target proteins of Rheum officinale and 33 Homo sapiens genes of Jaundice were found in databases. There were 19 common pathways both related networks. Rheum officinale could regulate endothelial differentiation, Interleukin-1B (IL-1B) and Tumor Necrosis Factor (TNF) in these pathways. Conclusions: Rheum officinale treat Jaundice by regulating many effective nodes of Apoptotic pathway and cellular immunity related pathways.

USP1 deubiquitinase: cellular functions, regulatory mechanisms and emerging potential as target in cancer therapy

PubMed Central

2013-01-01

Reversible protein ubiquitination is emerging as a key process for maintaining cell homeostasis, and the enzymes that participate in this process, in particular E3 ubiquitin ligases and deubiquitinases (DUBs), are increasingly being regarded as candidates for drug discovery. Human DUBs are a group of approximately 100 proteins, whose cellular functions and regulatory mechanisms remain, with some exceptions, poorly characterized. One of the best-characterized human DUBs is ubiquitin-specific protease 1 (USP1), which plays an important role in the cellular response to DNA damage. USP1 levels, localization and activity are modulated through several mechanisms, including protein-protein interactions, autocleavage/degradation and phosphorylation, ensuring that USP1 function is carried out in a properly regulated spatio-temporal manner. Importantly, USP1 expression is deregulated in certain types of human cancer, suggesting that USP1 could represent a valid target in cancer therapy. This view has gained recent support with the finding that USP1 inhibition may contribute to revert cisplatin resistance in an in vitro model of non-small cell lung cancer (NSCLC). Here, we describe the current knowledge on the cellular functions and regulatory mechanisms of USP1. We also summarize USP1 alterations found in cancer, combining data from the literature and public databases with our own data. Finally, we discuss the emerging potential of USP1 as a target, integrating published data with our novel findings on the effects of the USP1 inhibitor pimozide in combination with cisplatin in NSCLC cells. PMID:23937906

Proteomic profile of dormant Trichophyton Rubrum conidia

PubMed Central

Leng, Wenchuan; Liu, Tao; Li, Rui; Yang, Jian; Wei, Candong; Zhang, Wenliang; Jin, Qi

2008-01-01

Background Trichophyton rubrum is the most common dermatophyte causing fungal skin infections in humans. Asexual sporulation is an important means of propagation for T. rubrum, and conidia produced by this way are thought to be the primary cause of human infections. Despite their importance in pathogenesis, the conidia of T. rubrum remain understudied. We intend to intensively investigate the proteome of dormant T. rubrum conidia to characterize its molecular and cellular features and to enhance the development of novel therapeutic strategies. Results The proteome of T. rubrum conidia was analyzed by combining shotgun proteomics with sample prefractionation and multiple enzyme digestion. In total, 1026 proteins were identified. All identified proteins were compared to those in the NCBI non-redundant protein database, the eukaryotic orthologous groups database, and the gene ontology database to obtain functional annotation information. Functional classification revealed that the identified proteins covered nearly all major biological processes. Some proteins were spore specific and related to the survival and dispersal of T. rubrum conidia, and many proteins were important to conidial germination and response to environmental conditions. Conclusion Our results suggest that the proteome of T. rubrum conidia is considerably complex, and that the maintenance of conidial dormancy is an intricate and elaborate process. This data set provides the first global framework for the dormant T. rubrum conidia proteome and is a stepping stone on the way to further study of the molecular mechanisms of T. rubrum conidial germination and the maintenance of conidial dormancy. PMID:18578874

Cellular responses during morphological transformation in Azospirillum brasilense and Its flcA knockout mutant.

PubMed

Hou, Xingsheng; McMillan, Mary; Coumans, Joëlle V F; Poljak, Anne; Raftery, Mark J; Pereg, Lily

2014-01-01

FlcA is a response regulator controlling flocculation and the morphological transformation of Azospirillum cells from vegetative to cyst-like forms. To understand the cellular responses of Azospirillum to conditions that cause morphological transformation, proteins differentially expressed under flocculation conditions in A. brasilense Sp7 and its flcA knockout mutant were investigated. Comparison of 2-DE protein profiles of wild-type (Sp7) and a flcA deletion mutant (Sp7-flcAΔ) revealed a total of 33 differentially expressed 2-DE gel spots, with 22 of these spots confidently separated to allow protein identification. Analysis of these spots by liquid chromatography-tandem mass spectrometry (LC-MS/MS) and MASCOT database searching identified 48 proteins (≥10% emPAI in each spot). The functional characteristics of these proteins included carbon metabolism (beta-ketothiolase and citrate synthase), nitrogen metabolism (Glutamine synthetase and nitric oxide synthase), stress tolerance (superoxide dismutase, Alkyl hydroperoxidase and ATP-dependent Clp protease proteolytic subunit) and morphological transformation (transducer coupling protein). The observed differences between Sp7 wild-type and flcA- strains enhance our understanding of the morphological transformation process and help to explain previous phenotypical observations. This work is a step forward in connecting the Azospirillum phenome and genome.

Cellular Responses during Morphological Transformation in Azospirillum brasilense and Its flcA Knockout Mutant

PubMed Central

Coumans, Joëlle V. F.; Poljak, Anne; Raftery, Mark J.; Pereg, Lily

2014-01-01

FlcA is a response regulator controlling flocculation and the morphological transformation of Azospirillum cells from vegetative to cyst-like forms. To understand the cellular responses of Azospirillum to conditions that cause morphological transformation, proteins differentially expressed under flocculation conditions in A. brasilense Sp7 and its flcA knockout mutant were investigated. Comparison of 2-DE protein profiles of wild-type (Sp7) and a flcA deletion mutant (Sp7-flcAΔ) revealed a total of 33 differentially expressed 2-DE gel spots, with 22 of these spots confidently separated to allow protein identification. Analysis of these spots by liquid chromatography-tandem mass spectrometry (LC-MS/MS) and MASCOT database searching identified 48 proteins (≥10% emPAI in each spot). The functional characteristics of these proteins included carbon metabolism (beta-ketothiolase and citrate synthase), nitrogen metabolism (Glutamine synthetase and nitric oxide synthase), stress tolerance (superoxide dismutase, Alkyl hydroperoxidase and ATP-dependent Clp protease proteolytic subunit) and morphological transformation (transducer coupling protein). The observed differences between Sp7 wild-type and flcA − strains enhance our understanding of the morphological transformation process and help to explain previous phenotypical observations. This work is a step forward in connecting the Azospirillum phenome and genome. PMID:25502569

Unbiased compound screening with a reporter gene assay highlights the role of p13 in the cardiac cellular stress response.

PubMed

Inoue, Naoki; Hirouchi, Taisei; Kasai, Atsushi; Higashi, Shintaro; Hiraki, Natsumi; Tanaka, Shota; Nakazawa, Takanobu; Nunomura, Kazuto; Lin, Bangzhong; Omori, Akiko; Hayata-Takano, Atsuko; Kim, Yoon-Jeong; Doi, Takefumi; Baba, Akemichi; Hashimoto, Hitoshi; Shintani, Norihito

2018-01-08

We recently showed that a 13-kDa protein (p13), the homolog protein of formation of mitochondrial complex V assembly factor 1 in yeast, acts as a potential protective factor in pancreatic islets under diabetes. Here, we aimed to identify known compounds regulating p13 mRNA expression to obtain therapeutic insight into the cellular stress response. A luciferase reporter system was developed using the putative promoter region of the human p13 gene. Overexpression of peroxisome proliferator-activated receptor gamma coactivator 1α, a master player regulating mitochondrial metabolism, increased both reporter activity and p13 expression. Following unbiased screening with 2320 known compounds in HeLa cells, 12 pharmacological agents (including 8 cardiotonics and 2 anthracyclines) that elicited >2-fold changes in p13 mRNA expression were identified. Among them, four cardiac glycosides decreased p13 expression and concomitantly elevated cellular oxidative stress. Additional database analyses showed highest p13 expression in heart, with typically decreased expression in cardiac disease. Accordingly, our results illustrate the usefulness of unbiased compound screening as a method for identifying novel functional roles of unfamiliar genes. Our findings also highlight the importance of p13 in the cellular stress response in heart. Copyright © 2017. Published by Elsevier Inc.

KEGG orthology-based annotation of the predicted proteome of Acropora digitifera: ZoophyteBase - an open access and searchable database of a coral genome

PubMed Central

2013-01-01

Background Contemporary coral reef research has firmly established that a genomic approach is urgently needed to better understand the effects of anthropogenic environmental stress and global climate change on coral holobiont interactions. Here we present KEGG orthology-based annotation of the complete genome sequence of the scleractinian coral Acropora digitifera and provide the first comprehensive view of the genome of a reef-building coral by applying advanced bioinformatics. Description Sequences from the KEGG database of protein function were used to construct hidden Markov models. These models were used to search the predicted proteome of A. digitifera to establish complete genomic annotation. The annotated dataset is published in ZoophyteBase, an open access format with different options for searching the data. A particularly useful feature is the ability to use a Google-like search engine that links query words to protein attributes. We present features of the annotation that underpin the molecular structure of key processes of coral physiology that include (1) regulatory proteins of symbiosis, (2) planula and early developmental proteins, (3) neural messengers, receptors and sensory proteins, (4) calcification and Ca2+-signalling proteins, (5) plant-derived proteins, (6) proteins of nitrogen metabolism, (7) DNA repair proteins, (8) stress response proteins, (9) antioxidant and redox-protective proteins, (10) proteins of cellular apoptosis, (11) microbial symbioses and pathogenicity proteins, (12) proteins of viral pathogenicity, (13) toxins and venom, (14) proteins of the chemical defensome and (15) coral epigenetics. Conclusions We advocate that providing annotation in an open-access searchable database available to the public domain will give an unprecedented foundation to interrogate the fundamental molecular structure and interactions of coral symbiosis and allow critical questions to be addressed at the genomic level based on combined aspects of evolutionary, developmental, metabolic, and environmental perspectives. PMID:23889801

Functional annotation from the genome sequence of the giant panda.

PubMed

Huo, Tong; Zhang, Yinjie; Lin, Jianping

2012-08-01

The giant panda is one of the most critically endangered species due to the fragmentation and loss of its habitat. Studying the functions of proteins in this animal, especially specific trait-related proteins, is therefore necessary to protect the species. In this work, the functions of these proteins were investigated using the genome sequence of the giant panda. Data on 21,001 proteins and their functions were stored in the Giant Panda Protein Database, in which the proteins were divided into two groups: 20,179 proteins whose functions can be predicted by GeneScan formed the known-function group, whereas 822 proteins whose functions cannot be predicted by GeneScan comprised the unknown-function group. For the known-function group, we further classified the proteins by molecular function, biological process, cellular component, and tissue specificity. For the unknown-function group, we developed a strategy in which the proteins were filtered by cross-Blast to identify panda-specific proteins under the assumption that proteins related to the panda-specific traits in the unknown-function group exist. After this filtering procedure, we identified 32 proteins (2 of which are membrane proteins) specific to the giant panda genome as compared against the dog and horse genomes. Based on their amino acid sequences, these 32 proteins were further analyzed by functional classification using SVM-Prot, motif prediction using MyHits, and interacting protein prediction using the Database of Interacting Proteins. Nineteen proteins were predicted to be zinc-binding proteins, thus affecting the activities of nucleic acids. The 32 panda-specific proteins will be further investigated by structural and functional analysis.

A novel cell penetrating peptide carrier for the delivery of nematocidal proteins drug

NASA Astrophysics Data System (ADS)

Kim, Jea Hyun

Nematodes have recently become a primary source of harmful diseases to the environment that inflict harsh damages to pine trees and marine species. However, nematodes cannot be killed by normal pesticides or chemicals due to their thick outer protective layer mainly composed of collagen and cuticles. Thus, a novel approach to trigger intracellular delivery of chemicals through the layers of nematodes is required. In this study, the selection of the novel CPP was carefully progressed through protein database and serial digested fragmentation, internalization of each amino sequence was analyzed through flow cytometry and confocal microscope. As one of the most effective CPP material, JH 1.6 was compared with other major CPPs and its cellular toxicity was investigated. Furthermore, JH 1.6 was attached to various RNA, DNA, and proteins and internalization efficiency was evaluated for mammalian cells. To examine its effects on nematodes in vivo, JH 1.6 was conjugated with nematocidal protein - botulinum neurotoxin (BnT) and treated in C.elegans as a model animal. The results showed that JH 1.6 had high relative internalization rate and low cellular toxicity compared to other major CPP such as TAT and GV1001 peptides.

E3Net: a system for exploring E3-mediated regulatory networks of cellular functions.

PubMed

Han, Youngwoong; Lee, Hodong; Park, Jong C; Yi, Gwan-Su

2012-04-01

Ubiquitin-protein ligase (E3) is a key enzyme targeting specific substrates in diverse cellular processes for ubiquitination and degradation. The existing findings of substrate specificity of E3 are, however, scattered over a number of resources, making it difficult to study them together with an integrative view. Here we present E3Net, a web-based system that provides a comprehensive collection of available E3-substrate specificities and a systematic framework for the analysis of E3-mediated regulatory networks of diverse cellular functions. Currently, E3Net contains 2201 E3s and 4896 substrates in 427 organisms and 1671 E3-substrate specific relations between 493 E3s and 1277 substrates in 42 organisms, extracted mainly from MEDLINE abstracts and UniProt comments with an automatic text mining method and additional manual inspection and partly from high throughput experiment data and public ubiquitination databases. The significant functions and pathways of the extracted E3-specific substrate groups were identified from a functional enrichment analysis with 12 functional category resources for molecular functions, protein families, protein complexes, pathways, cellular processes, cellular localization, and diseases. E3Net includes interactive analysis and navigation tools that make it possible to build an integrative view of E3-substrate networks and their correlated functions with graphical illustrations and summarized descriptions. As a result, E3Net provides a comprehensive resource of E3s, substrates, and their functional implications summarized from the regulatory network structures of E3-specific substrate groups and their correlated functions. This resource will facilitate further in-depth investigation of ubiquitination-dependent regulatory mechanisms. E3Net is freely available online at http://pnet.kaist.ac.kr/e3net.

DenHunt - A Comprehensive Database of the Intricate Network of Dengue-Human Interactions

PubMed Central

Arjunan, Selvam; Sastri, Narayan P.; Chandra, Nagasuma

2016-01-01

Dengue virus (DENV) is a human pathogen and its etiology has been widely established. There are many interactions between DENV and human proteins that have been reported in literature. However, no publicly accessible resource for efficiently retrieving the information is yet available. In this study, we mined all publicly available dengue–human interactions that have been reported in the literature into a database called DenHunt. We retrieved 682 direct interactions of human proteins with dengue viral components, 382 indirect interactions and 4120 differentially expressed human genes in dengue infected cell lines and patients. We have illustrated the importance of DenHunt by mapping the dengue–human interactions on to the host interactome and observed that the virus targets multiple host functional complexes of important cellular processes such as metabolism, immune system and signaling pathways suggesting a potential role of these interactions in viral pathogenesis. We also observed that 7 percent of the dengue virus interacting human proteins are also associated with other infectious and non-infectious diseases. Finally, the understanding that comes from such analyses could be used to design better strategies to counteract the diseases caused by dengue virus. The whole dataset has been catalogued in a searchable database, called DenHunt (http://proline.biochem.iisc.ernet.in/DenHunt/). PMID:27618709

DenHunt - A Comprehensive Database of the Intricate Network of Dengue-Human Interactions.

PubMed

Karyala, Prashanthi; Metri, Rahul; Bathula, Christopher; Yelamanchi, Syam K; Sahoo, Lipika; Arjunan, Selvam; Sastri, Narayan P; Chandra, Nagasuma

2016-09-01

Dengue virus (DENV) is a human pathogen and its etiology has been widely established. There are many interactions between DENV and human proteins that have been reported in literature. However, no publicly accessible resource for efficiently retrieving the information is yet available. In this study, we mined all publicly available dengue-human interactions that have been reported in the literature into a database called DenHunt. We retrieved 682 direct interactions of human proteins with dengue viral components, 382 indirect interactions and 4120 differentially expressed human genes in dengue infected cell lines and patients. We have illustrated the importance of DenHunt by mapping the dengue-human interactions on to the host interactome and observed that the virus targets multiple host functional complexes of important cellular processes such as metabolism, immune system and signaling pathways suggesting a potential role of these interactions in viral pathogenesis. We also observed that 7 percent of the dengue virus interacting human proteins are also associated with other infectious and non-infectious diseases. Finally, the understanding that comes from such analyses could be used to design better strategies to counteract the diseases caused by dengue virus. The whole dataset has been catalogued in a searchable database, called DenHunt (http://proline.biochem.iisc.ernet.in/DenHunt/).

Data on the association of the nuclear envelope protein Sun1 with nucleoli.

PubMed

Moujaber, Ossama; Omran, Nawal; Kodiha, Mohamed; Pié, Brigitte; Cooper, Ellis; Presley, John F; Stochaj, Ursula

2017-08-01

SUN proteins participate in diverse cellular activities, many of which are connected to the nuclear envelope. Recently, the family member SUN1 has been linked to novel biological activities. These include the regulation of nucleoli, intranuclear compartments that assemble ribosomal subunits. We show that SUN1 associates with nucleoli in several mammalian epithelial cell lines. This nucleolar localization is not shared by all cell types, as SUN1 concentrates at the nuclear envelope in ganglionic neurons and non-neuronal satellite cells. Database analyses and Western blotting emphasize the complexity of SUN1 protein profiles in different mammalian cells. We constructed a STRING network which identifies SUN1-related proteins as part of a larger network that includes several nucleolar proteins. Taken together, the current data highlight the diversity of SUN1 proteins and emphasize the possible links between SUN1 and nucleoli.

Towards an understanding of wheat chloroplasts: a methodical investigation of thylakoid proteome.

PubMed

Kamal, Abu Hena Mostafa; Cho, Kun; Komatsu, Setsuko; Uozumi, Nobuyuki; Choi, Jong-Soon; Woo, Sun Hee

2012-05-01

We utilized Percoll density gradient centrifugation to isolate and fractionate chloroplasts of Korean winter wheat cultivar cv. Kumgang (Triticum aestivum L.). The resulting protein fractions were separated by one dimensional polyacrylamide gel electrophoresis (1D-PAGE) coupled with LTQ-FTICR mass spectrometry. This enabled us to detect and identify 767 unique proteins. Our findings represent the most comprehensive exploration of a proteome to date. Based on annotation information from the UniProtKB/Swiss-Prot database and our analyses via WoLF PSORT and PSORT, these proteins are localized in the chloroplast (607 proteins), chloroplast stroma (145), thylakoid membrane (342), lumens (163), and integral membranes (166). In all, 67% were confirmed as chloroplast thylakoid proteins. Although nearly complete protein coverage (89% proteins) has been accomplished for the key chloroplast pathways in wheat, such as for photosynthesis, many other proteins are involved in regulating carbon metabolism. The identified proteins were assigned to 103 functional categories according to a classification system developed by the iProClass database and provided through Protein Information Resources. Those functions include electron transport, energy, cellular organization and biogenesis, transport, stress responses, and other metabolic processes. Whereas most of these proteins are associated with known complexes and metabolic pathways, about 13% of the proteins have unknown functions. The chloroplast proteome contains many proteins that are localized to the thylakoids but as yet have no known function. We propose that some of these familiar proteins participate in the photosynthetic pathway. Thus, our new and comprehensive protein profile may provide clues for better understanding that photosynthetic process in wheat.

New insights into protein-protein interaction data lead to increased estimates of the S. cerevisiae interactome size.

PubMed

Sambourg, Laure; Thierry-Mieg, Nicolas

2010-12-21

As protein interactions mediate most cellular mechanisms, protein-protein interaction networks are essential in the study of cellular processes. Consequently, several large-scale interactome mapping projects have been undertaken, and protein-protein interactions are being distilled into databases through literature curation; yet protein-protein interaction data are still far from comprehensive, even in the model organism Saccharomyces cerevisiae. Estimating the interactome size is important for evaluating the completeness of current datasets, in order to measure the remaining efforts that are required. We examined the yeast interactome from a new perspective, by taking into account how thoroughly proteins have been studied. We discovered that the set of literature-curated protein-protein interactions is qualitatively different when restricted to proteins that have received extensive attention from the scientific community. In particular, these interactions are less often supported by yeast two-hybrid, and more often by more complex experiments such as biochemical activity assays. Our analysis showed that high-throughput and literature-curated interactome datasets are more correlated than commonly assumed, but that this bias can be corrected for by focusing on well-studied proteins. We thus propose a simple and reliable method to estimate the size of an interactome, combining literature-curated data involving well-studied proteins with high-throughput data. It yields an estimate of at least 37, 600 direct physical protein-protein interactions in S. cerevisiae. Our method leads to higher and more accurate estimates of the interactome size, as it accounts for interactions that are genuine yet difficult to detect with commonly-used experimental assays. This shows that we are even further from completing the yeast interactome map than previously expected.

Update of KDBI: Kinetic Data of Bio-molecular Interaction database

PubMed Central

Kumar, Pankaj; Han, B. C.; Shi, Z.; Jia, J.; Wang, Y. P.; Zhang, Y. T.; Liang, L.; Liu, Q. F.; Ji, Z. L.; Chen, Y. Z.

2009-01-01

Knowledge of the kinetics of biomolecular interactions is important for facilitating the study of cellular processes and underlying molecular events, and is essential for quantitative study and simulation of biological systems. Kinetic Data of Bio-molecular Interaction database (KDBI) has been developed to provide information about experimentally determined kinetic data of protein–protein, protein–nucleic acid, protein–ligand, nucleic acid–ligand binding or reaction events described in the literature. To accommodate increasing demand for studying and simulating biological systems, numerous improvements and updates have been made to KDBI, including new ways to access data by pathway and molecule names, data file in System Biology Markup Language format, more efficient search engine, access to published parameter sets of simulation models of 63 pathways, and 2.3-fold increase of data (19 263 entries of 10 532 distinctive biomolecular binding and 11 954 interaction events, involving 2635 proteins/protein complexes, 847 nucleic acids, 1603 small molecules and 45 multi-step processes). KDBI is publically available at http://bidd.nus.edu.sg/group/kdbi/kdbi.asp. PMID:18971255

Data in support of the identification of neuronal and astrocyte proteins interacting with extracellularly applied oligomeric and fibrillar α-synuclein assemblies by mass spectrometry

PubMed Central

Shrivastava, Amulya Nidhi; Redeker, Virginie; Fritz, Nicolas; Pieri, Laura; Almeida, Leandro G.; Spolidoro, Maria; Liebmann, Thomas; Bousset, Luc; Renner, Marianne; Léna, Clément; Aperia, Anita; Melki, Ronald; Triller, Antoine

2016-01-01

α-Synuclein (α-syn) is the principal component of Lewy bodies, the pathophysiological hallmark of individuals affected by Parkinson disease (PD). This neuropathologic form of α-syn contributes to PD progression and propagation of α-syn assemblies between neurons. The data we present here support the proteomic analysis used to identify neuronal proteins that specifically interact with extracellularly applied oligomeric or fibrillar α-syn assemblies (conditions 1 and 2, respectively) (doi: 10.15252/embj.201591397[1]). α-syn assemblies and their cellular partner proteins were pulled down from neuronal cell lysed shortly after exposure to exogenous α-syn assemblies and the associated proteins were identified by mass spectrometry using a shotgun proteomic-based approach. We also performed experiments on pure cultures of astrocytes to identify astrocyte-specific proteins interacting with oligomeric or fibrillar α-syn (conditions 3 and 4, respectively). For each condition, proteins interacting selectively with α-syn assemblies were identified by comparison to proteins pulled-down from untreated cells used as controls. The mass spectrometry data, the database search and the peak lists have been deposited to the ProteomeXchange Consortium database via the PRIDE partner repository with the dataset identifiers PRIDE: PXD002256 to PRIDE: PXD002263 and doi: 10.6019/PXD002256 to 10.6019/PXD002263. PMID:26958642

Data in support of the identification of neuronal and astrocyte proteins interacting with extracellularly applied oligomeric and fibrillar α-synuclein assemblies by mass spectrometry.

PubMed

Shrivastava, Amulya Nidhi; Redeker, Virginie; Fritz, Nicolas; Pieri, Laura; Almeida, Leandro G; Spolidoro, Maria; Liebmann, Thomas; Bousset, Luc; Renner, Marianne; Léna, Clément; Aperia, Anita; Melki, Ronald; Triller, Antoine

2016-06-01

α-Synuclein (α-syn) is the principal component of Lewy bodies, the pathophysiological hallmark of individuals affected by Parkinson disease (PD). This neuropathologic form of α-syn contributes to PD progression and propagation of α-syn assemblies between neurons. The data we present here support the proteomic analysis used to identify neuronal proteins that specifically interact with extracellularly applied oligomeric or fibrillar α-syn assemblies (conditions 1 and 2, respectively) (doi: 10.15252/embj.201591397[1]). α-syn assemblies and their cellular partner proteins were pulled down from neuronal cell lysed shortly after exposure to exogenous α-syn assemblies and the associated proteins were identified by mass spectrometry using a shotgun proteomic-based approach. We also performed experiments on pure cultures of astrocytes to identify astrocyte-specific proteins interacting with oligomeric or fibrillar α-syn (conditions 3 and 4, respectively). For each condition, proteins interacting selectively with α-syn assemblies were identified by comparison to proteins pulled-down from untreated cells used as controls. The mass spectrometry data, the database search and the peak lists have been deposited to the ProteomeXchange Consortium database via the PRIDE partner repository with the dataset identifiers PRIDE: PXD002256 to PRIDE: PXD002263 and doi: 10.6019/PXD002256 to 10.6019/PXD002263.

Proteomic profiling of mature leaves from oil palm (Elaeis guineensis Jacq.).

PubMed

Tan, Hooi Sin; Jacoby, Richard P; Ong-Abdullah, Meilina; Taylor, Nicolas L; Liddell, Susan; Chee, Wong Wei; Chin, Chiew Foan

2017-04-01

Oil palm is one of the most productive oil bearing crops grown in Southeast Asia. Due to the dwindling availability of agricultural land and increasing demand for high yielding oil palm seedlings, clonal propagation is vital to the oil palm industry. Most commonly, leaf explants are used for in vitro micropropagation of oil palm and to optimize this process it is important to unravel the physiological and molecular mechanisms underlying somatic embryo production from leaves. In this study, a proteomic approach was used to determine protein abundance of mature oil palm leaves. To do this, leaf proteins were extracted using TCA/acetone precipitation protocol and separated by 2DE. A total of 191 protein spots were observed on the 2D gels and 67 of the most abundant protein spots that were consistently observed were selected for further analysis with 35 successfully identified using MALDI TOF/TOF MS. The majority of proteins were classified as being involved in photosynthesis, metabolism, cellular biogenesis, stress response, and transport. This study provides the first proteomic assessment of oil palm leaves in this important oil crop and demonstrates the successful identification of selected proteins spots using the Malaysian Palm Oil Board (MPOB) Elaeis guineensis EST and NCBI-protein databases. The MS data have been deposited in the ProteomeXchange Consortium database with the data set identifier PXD001307. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

HDAPD: a web tool for searching the disease-associated protein structures

PubMed Central

2010-01-01

Background The protein structures of the disease-associated proteins are important for proceeding with the structure-based drug design to against a particular disease. Up until now, proteins structures are usually searched through a PDB id or some sequence information. However, in the HDAPD database presented here the protein structure of a disease-associated protein can be directly searched through the associated disease name keyed in. Description The search in HDAPD can be easily initiated by keying some key words of a disease, protein name, protein type, or PDB id. The protein sequence can be presented in FASTA format and directly copied for a BLAST search. HDAPD is also interfaced with Jmol so that users can observe and operate a protein structure with Jmol. The gene ontological data such as cellular components, molecular functions, and biological processes are provided once a hyperlink to Gene Ontology (GO) is clicked. Further, HDAPD provides a link to the KEGG map such that where the protein is placed and its relationship with other proteins in a metabolic pathway can be found from the map. The latest literatures namely titles, journals, authors, and abstracts searched from PubMed for the protein are also presented as a length controllable list. Conclusions Since the HDAPD data content can be routinely updated through a PHP-MySQL web page built, the new database presented is useful for searching the structures for some disease-associated proteins that may play important roles in the disease developing process for performing the structure-based drug design to against the diseases. PMID:20158919

Inborn errors of metabolism and the human interactome: a systems medicine approach.

PubMed

Woidy, Mathias; Muntau, Ania C; Gersting, Søren W

2018-02-05

The group of inborn errors of metabolism (IEM) displays a marked heterogeneity and IEM can affect virtually all functions and organs of the human organism; however, IEM share that their associated proteins function in metabolism. Most proteins carry out cellular functions by interacting with other proteins, and thus are organized in biological networks. Therefore, diseases are rarely the consequence of single gene mutations but of the perturbations caused in the related cellular network. Systematic approaches that integrate multi-omics and database information into biological networks have successfully expanded our knowledge of complex disorders but network-based strategies have been rarely applied to study IEM. We analyzed IEM on a proteome scale and found that IEM-associated proteins are organized as a network of linked modules within the human interactome of protein interactions, the IEM interactome. Certain IEM disease groups formed self-contained disease modules, which were highly interlinked. On the other hand, we observed disease modules consisting of proteins from many different disease groups in the IEM interactome. Moreover, we explored the overlap between IEM and non-IEM disease genes and applied network medicine approaches to investigate shared biological pathways, clinical signs and symptoms, and links to drug targets. The provided resources may help to elucidate the molecular mechanisms underlying new IEM, to uncover the significance of disease-associated mutations, to identify new biomarkers, and to develop novel therapeutic strategies.

Proteome-wide Subcellular Topologies of E. coli Polypeptides Database (STEPdb)*

PubMed Central

Orfanoudaki, Georgia; Economou, Anastassios

2014-01-01

Cell compartmentalization serves both the isolation and the specialization of cell functions. After synthesis in the cytoplasm, over a third of all proteins are targeted to other subcellular compartments. Knowing how proteins are distributed within the cell and how they interact is a prerequisite for understanding it as a whole. Surface and secreted proteins are important pathogenicity determinants. Here we present the STEP database (STEPdb) that contains a comprehensive characterization of subcellular localization and topology of the complete proteome of Escherichia coli. Two widely used E. coli proteomes (K-12 and BL21) are presented organized into thirteen subcellular classes. STEPdb exploits the wealth of genetic, proteomic, biochemical, and functional information on protein localization, secretion, and targeting in E. coli, one of the best understood model organisms. Subcellular annotations were derived from a combination of bioinformatics prediction, proteomic, biochemical, functional, topological data and extensive literature re-examination that were refined through manual curation. Strong experimental support for the location of 1553 out of 4303 proteins was based on 426 articles and some experimental indications for another 526. Annotations were provided for another 320 proteins based on firm bioinformatic predictions. STEPdb is the first database that contains an extensive set of peripheral IM proteins (PIM proteins) and includes their graphical visualization into complexes, cellular functions, and interactions. It also summarizes all currently known protein export machineries of E. coli K-12 and pairs them, where available, with the secretory proteins that use them. It catalogs the Sec- and TAT-utilizing secretomes and summarizes their topological features such as signal peptides and transmembrane regions, transmembrane topologies and orientations. It also catalogs physicochemical and structural features that influence topology such as abundance, solubility, disorder, heat resistance, and structural domain families. Finally, STEPdb incorporates prediction tools for topology (TMHMM, SignalP, and Phobius) and disorder (IUPred) and implements the BLAST2STEP that performs protein homology searches against the STEPdb. PMID:25210196

Transcriptomic Analysis of the Endangered Neritid Species Clithon retropictus: De Novo Assembly, Functional Annotation, and Marker Discovery

PubMed Central

Park, So Young; Patnaik, Bharat Bhusan; Kang, Se Won; Hwang, Hee-Ju; Chung, Jong Min; Song, Dae Kwon; Sang, Min Kyu; Patnaik, Hongray Howrelia; Lee, Jae Bong; Noh, Mi Young; Kim, Changmu; Kim, Soonok; Park, Hong Seog; Lee, Jun Sang; Han, Yeon Soo; Lee, Yong Seok

2016-01-01

An aquatic gastropod belonging to the family Neritidae, Clithon retropictus is listed as an endangered class II species in South Korea. The lack of information on its genomic background limits the ability to obtain functional data resources and inhibits informed conservation planning for this species. In the present study, the transcriptomic sequencing and de novo assembly of C. retropictus generated a total of 241,696,750 high-quality reads. These assembled to 282,838 unigenes with mean and N50 lengths of 736.9 and 1201 base pairs, respectively. Of these, 125,616 unigenes were subjected to annotation analysis with known proteins in Protostome DB, COG, GO, and KEGG protein databases (BLASTX; E ≤ 0.00001) and with known nucleotides in the Unigene database (BLASTN; E ≤ 0.00001). The GO analysis indicated that cellular process, cell, and catalytic activity are the predominant GO terms in the biological process, cellular component, and molecular function categories, respectively. In addition, 2093 unigenes were distributed in 107 different KEGG pathways. Furthermore, 49,280 simple sequence repeats were identified in the unigenes (>1 kilobase sequences). This is the first report on the identification of transcriptomic and microsatellite resources for C. retropictus, which opens up the possibility of exploring traits related to the adaptation and acclimatization of this species. PMID:27455329

AtlasT4SS: a curated database for type IV secretion systems.

PubMed

Souza, Rangel C; del Rosario Quispe Saji, Guadalupe; Costa, Maiana O C; Netto, Diogo S; Lima, Nicholas C B; Klein, Cecília C; Vasconcelos, Ana Tereza R; Nicolás, Marisa F

2012-08-09

The type IV secretion system (T4SS) can be classified as a large family of macromolecule transporter systems, divided into three recognized sub-families, according to the well-known functions. The major sub-family is the conjugation system, which allows transfer of genetic material, such as a nucleoprotein, via cell contact among bacteria. Also, the conjugation system can transfer genetic material from bacteria to eukaryotic cells; such is the case with the T-DNA transfer of Agrobacterium tumefaciens to host plant cells. The system of effector protein transport constitutes the second sub-family, and the third one corresponds to the DNA uptake/release system. Genome analyses have revealed numerous T4SS in Bacteria and Archaea. The purpose of this work was to organize, classify, and integrate the T4SS data into a single database, called AtlasT4SS - the first public database devoted exclusively to this prokaryotic secretion system. The AtlasT4SS is a manual curated database that describes a large number of proteins related to the type IV secretion system reported so far in Gram-negative and Gram-positive bacteria, as well as in Archaea. The database was created using the RDBMS MySQL and the Catalyst Framework based in the Perl programming language and using the Model-View-Controller (MVC) design pattern for Web. The current version holds a comprehensive collection of 1,617 T4SS proteins from 58 Bacteria (49 Gram-negative and 9 Gram-Positive), one Archaea and 11 plasmids. By applying the bi-directional best hit (BBH) relationship in pairwise genome comparison, it was possible to obtain a core set of 134 clusters of orthologous genes encoding T4SS proteins. In our database we present one way of classifying orthologous groups of T4SSs in a hierarchical classification scheme with three levels. The first level comprises four classes that are based on the organization of genetic determinants, shared homologies, and evolutionary relationships: (i) F-T4SS, (ii) P-T4SS, (iii) I-T4SS, and (iv) GI-T4SS. The second level designates a specific well-known protein families otherwise an uncharacterized protein family. Finally, in the third level, each protein of an ortholog cluster is classified according to its involvement in a specific cellular process. AtlasT4SS database is open access and is available at http://www.t4ss.lncc.br.

Global De Novo Protein-Protein Interactome Elucidates Interactions of Drought-Responsive Proteins in Horse Gram (Macrotyloma uniflorum).

PubMed

Bhardwaj, Jyoti; Gangwar, Indu; Panzade, Ganesh; Shankar, Ravi; Yadav, Sudesh Kumar

2016-06-03

Inspired by the availability of de novo transcriptome of horse gram (Macrotyloma uniflorum) and recent developments in systems biology studies, the first ever global protein-protein interactome (PPI) map was constructed for this highly drought-tolerant legume. Large-scale studies of PPIs and the constructed database would provide rationale behind the interplay at cascading translational levels for drought stress-adaptive mechanisms in horse gram. Using a bidirectional approach (interolog and domain-based), a high-confidence interactome map and database for horse gram was constructed. Available transcriptomic information for shoot and root tissues of a sensitive (M-191; genotype 1) and a drought-tolerant (M-249; genotype 2) genotype of horse gram was utilized to draw comparative PPI subnetworks under drought stress. High-confidence 6804 interactions were predicted among 1812 proteins covering about one-fourth of the horse gram proteome. The highest number of interactions (33.86%) in horse gram interactome matched with Arabidopsis PPI data. The top five hub nodes mostly included ubiquitin and heat-shock-related proteins. Higher numbers of PPIs were found to be responsive in shoot tissue (416) and root tissue (2228) of genotype 2 compared with shoot tissue (136) and root tissue (579) of genotype 1. Characterization of PPIs using gene ontology analysis revealed that kinase and transferase activities involved in signal transduction, cellular processes, nucleocytoplasmic transport, protein ubiquitination, and localization of molecules were most responsive to drought stress. Hence, these could be framed in stress adaptive mechanisms of horse gram. Being the first legume global PPI map, it would provide new insights into gene and protein regulatory networks for drought stress tolerance mechanisms in horse gram. Information compiled in the form of database (MauPIR) will provide the much needed high-confidence systems biology information for horse gram genes, proteins, and involved processes. This information would ease the effort and increase the efficacy for similar studies on other legumes. Public access is available at http://14.139.59.221/MauPIR/ .

Distribution of cellular HSV-1 receptor expression in human brain.

PubMed

Lathe, Richard; Haas, Juergen G

2017-06-01

Herpes simplex virus type 1 (HSV-1) is a neurotropic virus linked to a range of acute and chronic neurological disorders affecting distinct regions of the brain. Unusually, HSV-1 entry into cells requires the interaction of viral proteins glycoprotein D (gD) and glycoprotein B (gB) with distinct cellular receptor proteins. Several different gD and gB receptors have been identified, including TNFRSF14/HVEM and PVRL1/nectin 1 as gD receptors and PILRA, MAG, and MYH9 as gB receptors. We investigated the expression of these receptor molecules in different areas of the adult and developing human brain using online transcriptome databases. Whereas all HSV-1 receptors showed distinct expression patterns in different brain areas, the Allan Brain Atlas (ABA) reported increased expression of both gD and gB receptors in the hippocampus. Specifically, for PVRL1, TNFRFS14, and MYH9, the differential z scores for hippocampal expression, a measure of relative levels of increased expression, rose to 2.9, 2.9, and 2.5, respectively, comparable to the z score for the archetypical hippocampus-enriched mineralocorticoid receptor (NR3C2, z = 3.1). These data were confirmed at the Human Brain Transcriptome (HBT) database, but HBT data indicate that MAG expression is also enriched in hippocampus. The HBT database allowed the developmental pattern of expression to be investigated; we report that all HSV1 receptors markedly increase in expression levels between gestation and the postnatal/adult periods. These results suggest that differential receptor expression levels of several HSV-1 gD and gB receptors in the adult hippocampus are likely to underlie the susceptibility of this brain region to HSV-1 infection.

Systematic Mapping and Functional Analysis of a Family of Human Epididymal Secretory Sperm-Located Proteins*

PubMed Central

Li, JianYuan; Liu, FuJun; Wang, HaiYan; Liu, Xin; Liu, Juan; Li, Ning; Wan, FengChun; Wang, WenTing; Zhang, ChengLin; Jin, ShaoHua; Liu, Jie; Zhu, Peng; Liu, YunXiang

2010-01-01

The mammalian spermatozoon has many cellular compartments, such as head and tail, permitting it to interact with the female reproductive tract and fertilize the egg. It acquires this fertilizing potential during transit through the epididymis, which secretes proteins that coat different sperm domains. Optimal levels of these proteins provide the spermatozoon with its ability to move to, bind to, fuse with, and penetrate the egg; otherwise male infertility results. As few human epididymal proteins have been characterized, this work was performed to generate a database of human epididymal sperm-located proteins involved in maturation. Two-dimensional gel electrophoresis of epididymal tissue and luminal fluid proteins, followed by identification using MALDI-TOF/MS or MALDI-TOF/TOF, revealed over a thousand spots in gels comprising 745 abundant nonstructural proteins, 408 in luminal fluids, of which 207 were present on spermatozoa. Antibodies raised to 619 recombinant or synthetic peptides, used in Western blots, histological sections, and washed sperm preparations to confirm antibody quality and protein expression, indicated their regional location in the epididymal epithelium and highly specific locations on washed functional spermatozoa. Sperm function tests suggested the role of some proteins in motility and protection against oxidative attack. A large database of these proteins, characterized by size, pI, chromosomal location, and function, was given a unified terminology reflecting their sperm domain location. These novel, secreted human epididymal proteins are potential targets for a posttesticular contraceptive acting to provide rapid, reversible, functional sterility in men and they are also biomarkers that could be used in noninvasive assessments of male fertility. PMID:20736409

HIstome--a relational knowledgebase of human histone proteins and histone modifying enzymes.

PubMed

Khare, Satyajeet P; Habib, Farhat; Sharma, Rahul; Gadewal, Nikhil; Gupta, Sanjay; Galande, Sanjeev

2012-01-01

Histones are abundant nuclear proteins that are essential for the packaging of eukaryotic DNA into chromosomes. Different histone variants, in combination with their modification 'code', control regulation of gene expression in diverse cellular processes. Several enzymes that catalyze the addition and removal of multiple histone modifications have been discovered in the past decade, enabling investigations of their role(s) in normal cellular processes and diverse pathological conditions. This sudden influx of data, however, has resulted in need of an updated knowledgebase that compiles, organizes and presents curated scientific information to the user in an easily accessible format. Here, we present HIstome, a browsable, manually curated, relational database that provides information about human histone proteins, their sites of modifications, variants and modifying enzymes. HIstome is a knowledgebase of 55 human histone proteins, 106 distinct sites of their post-translational modifications (PTMs) and 152 histone-modifying enzymes. Entries have been grouped into 5 types of histones, 8 types of post-translational modifications and 14 types of enzymes that catalyze addition and removal of these modifications. The resource will be useful for epigeneticists, pharmacologists and clinicians. HIstome: The Histone Infobase is available online at http://www.iiserpune.ac.in/∼coee/histome/ and http://www.actrec.gov.in/histome/.

NovelFam3000 – Uncharacterized human protein domains conserved across model organisms

PubMed Central

Kemmer, Danielle; Podowski, Raf M; Arenillas, David; Lim, Jonathan; Hodges, Emily; Roth, Peggy; Sonnhammer, Erik LL; Höög, Christer; Wasserman, Wyeth W

2006-01-01

Background Despite significant efforts from the research community, an extensive portion of the proteins encoded by human genes lack an assigned cellular function. Most metazoan proteins are composed of structural and/or functional domains, of which many appear in multiple proteins. Once a domain is characterized in one protein, the presence of a similar sequence in an uncharacterized protein serves as a basis for inference of function. Thus knowledge of a domain's function, or the protein within which it arises, can facilitate the analysis of an entire set of proteins. Description From the Pfam domain database, we extracted uncharacterized protein domains represented in proteins from humans, worms, and flies. A data centre was created to facilitate the analysis of the uncharacterized domain-containing proteins. The centre both provides researchers with links to dispersed internet resources containing gene-specific experimental data and enables them to post relevant experimental results or comments. For each human gene in the system, a characterization score is posted, allowing users to track the progress of characterization over time or to identify for study uncharacterized domains in well-characterized genes. As a test of the system, a subset of 39 domains was selected for analysis and the experimental results posted to the NovelFam3000 system. For 25 human protein members of these 39 domain families, detailed sub-cellular localizations were determined. Specific observations are presented based on the analysis of the integrated information provided through the online NovelFam3000 system. Conclusion Consistent experimental results between multiple members of a domain family allow for inferences of the domain's functional role. We unite bioinformatics resources and experimental data in order to accelerate the functional characterization of scarcely annotated domain families. PMID:16533400

Innovative computer-aided methods for the discovery of new kinase ligands.

PubMed

Abuhammad, Areej; Taha, Mutasem

2016-04-01

Recent evidence points to significant roles played by protein kinases in cell signaling and cellular proliferation. Faulty protein kinases are involved in cancer, diabetes and chronic inflammation. Efforts are continuously carried out to discover new inhibitors for selected protein kinases. In this review, we discuss two new computer-aided methodologies we developed to mine virtual databases for new bioactive compounds. One method is ligand-based exploration of the pharmacophoric space of inhibitors of any particular biotarget followed by quantitative structure-activity relationship-based selection of the best pharmacophore(s). The second approach is structure-based assuming that potent ligands come into contact with binding site spots distinct from those contacted by weakly potent ligands. Both approaches yield pharmacophores useful as 3D search queries for the discovery of new bioactive (kinase) inhibitors.

Purification of intact chloroplasts from marine plant Posidonia oceanica suitable for organelle proteomics.

PubMed

Piro, Amalia; Serra, Ilia Anna; Spadafora, Antonia; Cardilio, Monica; Bianco, Linda; Perrotta, Gaetano; Santos, Rui; Mazzuca, Silvia

2015-12-01

Posidonia oceanica is a marine angiosperm, or seagrass, adapted to grow to the underwater life from shallow waters to 50 m depth. This raises questions of how their photosynthesis adapted to the attenuation of light through the water column and leads to the assumption that biochemistry and metabolism of the chloroplast are the basis of adaptive capacity. In the present study, we described a protocol that was adapted from those optimized for terrestrial plants, to extract chloroplasts from as minimal tissue as possible. We obtained the best balance between tissue amount/intact chloroplasts yield using one leaf from one plant. After isopynic separations, the chloroplasts purity and integrity were evaluated by biochemical assay and using a proteomic approach. Chloroplast proteins were extracted from highly purified organelles and resolved by 1DE SDS-PAGE. Proteins were sequenced by nLC-ESI-IT-MS/MS of 1DE gel bands and identified against NCBInr green plant databases, Dr. Zompo database for seagrasses in a local customized dataset. The curated localization of proteins in sub-plastidial compartments (i.e. envelope, stroma and thylakoids) was retrieved in the AT_CHLORO database. This purification protocol and the validation of compartment markers may serve as basis for sub-cellular proteomics in P. oceanica and other seagrasses. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Physical and in silico approaches identify DNA-PK in a Tax DNA-damage response interactome

PubMed Central

Ramadan, Emad; Ward, Michael; Guo, Xin; Durkin, Sarah S; Sawyer, Adam; Vilela, Marcelo; Osgood, Christopher; Pothen, Alex; Semmes, Oliver J

2008-01-01

Background We have initiated an effort to exhaustively map interactions between HTLV-1 Tax and host cellular proteins. The resulting Tax interactome will have significant utility toward defining new and understanding known activities of this important viral protein. In addition, the completion of a full Tax interactome will also help shed light upon the functional consequences of these myriad Tax activities. The physical mapping process involved the affinity isolation of Tax complexes followed by sequence identification using tandem mass spectrometry. To date we have mapped 250 cellular components within this interactome. Here we present our approach to prioritizing these interactions via an in silico culling process. Results We first constructed an in silico Tax interactome comprised of 46 literature-confirmed protein-protein interactions. This number was then reduced to four Tax-interactions suspected to play a role in DNA damage response (Rad51, TOP1, Chk2, 53BP1). The first-neighbor and second-neighbor interactions of these four proteins were assembled from available human protein interaction databases. Through an analysis of betweenness and closeness centrality measures, and numbers of interactions, we ranked proteins in the first neighborhood. When this rank list was compared to the list of physical Tax-binding proteins, DNA-PK was the highest ranked protein common to both lists. An overlapping clustering of the Tax-specific second-neighborhood protein network showed DNA-PK to be one of three bridge proteins that link multiple clusters in the DNA damage response network. Conclusion The interaction of Tax with DNA-PK represents an important biological paradigm as suggested via consensus findings in vivo and in silico. We present this methodology as an approach to discovery and as a means of validating components of a consensus Tax interactome. PMID:18922151

Insights into the Specificity of Lysine Acetyltransferases

DOE PAGES

Tucker, Alex C.; Taylor, Keenan C.; Rank, Katherine C.; ...

2014-11-07

Reversible lysine acetylation by protein acetyltransferases is a conserved regulatory mechanism that controls diverse cellular pathways. Gcn5-related N-acetyltransferases (GNATs), named after their founding member, are found in all domains of life. GNATs are known for their role as histone acetyltransferases, but non-histone bacterial protein acetytransferases have been identified. Only structures of GNAT complexes with short histone peptide substrates are available in databases. Given the biological importance of this modification and the abundance of lysine in polypeptides, how specificity is attained for larger protein substrates is central to understanding acetyl-lysine-regulated networks. In this paper, we report the structure of a GNATmore » in complex with a globular protein substrate solved to 1.9 Å. GNAT binds the protein substrate with extensive surface interactions distinct from those reported for GNAT-peptide complexes. Finally, our data reveal determinants needed for the recognition of a protein substrate and provide insight into the specificity of GNATs.« less

Identification of osteosarcoma-related specific proteins in serum samples using surface-enhanced laser desorption/ionization-time-of-flight mass spectrometry.

PubMed

Gu, Jianli; Li, Jitian; Huang, Manyu; Zhang, Zhiyong; Li, Dongsheng; Song, Guoying; Ding, Xingpo; Li, Wuyin

2014-01-01

Osteosarcoma (OS) is the most common malignant bone tumor. To identify OS-related specific proteins for early diagnosis of OS, a novel approach, surface-enhanced laser desorption/ionization-time-of-flight mass spectrometry (SELDI-TOF-MS) to serum samples from 25 OS patients, 16 osteochondroma, and 26 age-matched normal human volunteers as controls, was performed. Two proteins showed a significantly different expression in OS serum samples from control groups. Proteomic profiles and external leave-one-out cross-validation analysis showed that the correct rate of allocation, the sensitivity, and the specificity of diagnosis were 100%. These two proteins were further identified by searching the EPO-KB database, and one of the proteins identified as Serine rich region profile is involved in various cellular signaling cascades and tumor genesis. The presence of these two proteins in OS patients but absence from premalignant and normal human controls implied that they can be potential biomarkers for early diagnosis of OS.

Global analysis of host-pathogen interactions that regulate early stage HIV-1 replication

PubMed Central

König, Renate; Zhou, Yingyao; Elleder, Daniel; Diamond, Tracy L.; Bonamy, Ghislain M.C.; Irelan, Jeffrey T.; Chiang, Chih-yuan; Tu, Buu P.; De Jesus, Paul D.; Lilley, Caroline E.; Seidel, Shannon; Opaluch, Amanda M.; Caldwell, Jeremy S.; Weitzman, Matthew D.; Kuhen, Kelli L.; Bandyopadhyay, Sourav; Ideker, Trey; Orth, Anthony P.; Miraglia, Loren J.; Bushman, Frederic D.; Young, John A.; Chanda, Sumit K.

2008-01-01

Human Immunodeficiency Viruses (HIV-1 and HIV-2) rely upon host-encoded proteins to facilitate their replication. Here we combined genome-wide siRNA analyses with interrogation of human interactome databases to assemble a host-pathogen biochemical network containing 213 confirmed host cellular factors and 11 HIV-1-encoded proteins. Protein complexes that regulate ubiquitin conjugation, proteolysis, DNA damage response and RNA splicing were identified as important modulators of early stage HIV-1 infection. Additionally, over 40 new factors were shown to specifically influence initiation and/or kinetics of HIV-1 DNA synthesis, including cytoskeletal regulatory proteins, modulators of post-translational modification, and nucleic acid binding proteins. Finally, fifteen proteins with diverse functional roles, including nuclear transport, prostaglandin synthesis, ubiquitination, and transcription, were found to influence nuclear import or viral DNA integration. Taken together, the multi-scale approach described here has uncovered multiprotein virus-host interactions that likely act in concert to facilitate early steps of HIV-1 infection. PMID:18854154

Approaches for Defining the Hsp90-dependent Proteome

PubMed Central

Hartson, Steven D.; Matts, Robert L.

2011-01-01

Hsp90 is the target of ongoing drug discovery studies seeking new compounds to treat cancer, neurodegenerative diseases, and protein folding disorders. To better understand Hsp90’s roles in cellular pathologies and in normal cells, numerous studies have utilized proteomics assays and related high-throughput tools to characterize its physical and functional protein partnerships. This review surveys these studies, and summarizes the strengths and limitations of the individual attacks. We also include downloadable spreadsheets compiling all of the Hsp90-interacting proteins identified in more than 23 studies. These tools include cross-references among gene aliases, human homologues of yeast Hsp90-interacting proteins, hyperlinks to database entries, summaries of canonical pathways that are enriched in the Hsp90 interactome, and additional bioinformatic annotations. In addition to summarizing Hsp90 proteomics studies performed to date and the insights they have provided, we identify gaps in our current understanding of Hsp90-mediated proteostasis. PMID:21906632

Profiling of cellular proteins in porcine reproductive and respiratory syndrome virus virions by proteomics analysis

PubMed Central

2010-01-01

Background Porcine reproductive and respiratory syndrome virus (PRRSV) is an enveloped virus, bearing severe economic consequences to the swine industry worldwide. Previous studies on enveloped viruses have shown that many incorporated cellular proteins associated with the virion's membranes that might play important roles in viral infectivity. In this study, we sought to proteomically profile the cellular proteins incorporated into or associated with the virions of a highly virulent PRRSV strain GDBY1, and to provide foundation for further investigations on the roles of incorporated/associated cellular proteins on PRRSV's infectivity. Results In our experiment, sixty one cellular proteins were identified in highly purified PRRSV virions by two-dimensional gel electrophoresis coupled with mass spectrometric approaches. The identified cellular proteins could be grouped into eight functional categories including cytoskeletal proteins, chaperones, macromolecular biosynthesis proteins, metabolism-associated proteins, calcium-dependent membrane-binding proteins and other functional proteins. Among the identified proteins, four have not yet been reported in other studied envelope viruses, namely, guanine nucleotide-binding proteins, tyrosine 3-monooxygenase/tryptophan 5-monooxygenase, peroxiredoxin 1 and galectin-1 protein. The presence of five selected cellular proteins (i.e., β-actin, Tubulin, Annexin A2, heat shock protein Hsp27, and calcium binding proteins S100) in the highly purified PRRSV virions was validated by Western blot and immunogold labeling assays. Conclusions Taken together, the present study has demonstrated the incorporation of cellular proteins in PRRSV virions, which provides valuable information for the further investigations for the effects of individual cellular proteins on the viral replication, assembly, and pathogenesis. PMID:20849641

PLAN2L: a web tool for integrated text mining and literature-based bioentity relation extraction.

PubMed

Krallinger, Martin; Rodriguez-Penagos, Carlos; Tendulkar, Ashish; Valencia, Alfonso

2009-07-01

There is an increasing interest in using literature mining techniques to complement information extracted from annotation databases or generated by bioinformatics applications. Here we present PLAN2L, a web-based online search system that integrates text mining and information extraction techniques to access systematically information useful for analyzing genetic, cellular and molecular aspects of the plant model organism Arabidopsis thaliana. Our system facilitates a more efficient retrieval of information relevant to heterogeneous biological topics, from implications in biological relationships at the level of protein interactions and gene regulation, to sub-cellular locations of gene products and associations to cellular and developmental processes, i.e. cell cycle, flowering, root, leaf and seed development. Beyond single entities, also predefined pairs of entities can be provided as queries for which literature-derived relations together with textual evidences are returned. PLAN2L does not require registration and is freely accessible at http://zope.bioinfo.cnio.es/plan2l.

Systems Biology Approaches for Discovering Biomarkers for Traumatic Brain Injury

PubMed Central

Feala, Jacob D.; AbdulHameed, Mohamed Diwan M.; Yu, Chenggang; Dutta, Bhaskar; Yu, Xueping; Schmid, Kara; Dave, Jitendra; Tortella, Frank

2013-01-01

Abstract The rate of traumatic brain injury (TBI) in service members with wartime injuries has risen rapidly in recent years, and complex, variable links have emerged between TBI and long-term neurological disorders. The multifactorial nature of TBI secondary cellular response has confounded attempts to find cellular biomarkers for its diagnosis and prognosis or for guiding therapy for brain injury. One possibility is to apply emerging systems biology strategies to holistically probe and analyze the complex interweaving molecular pathways and networks that mediate the secondary cellular response through computational models that integrate these diverse data sets. Here, we review available systems biology strategies, databases, and tools. In addition, we describe opportunities for applying this methodology to existing TBI data sets to identify new biomarker candidates and gain insights about the underlying molecular mechanisms of TBI response. As an exemplar, we apply network and pathway analysis to a manually compiled list of 32 protein biomarker candidates from the literature, recover known TBI-related mechanisms, and generate hypothetical new biomarker candidates. PMID:23510232

Patterns of HIV-1 Protein Interaction Identify Perturbed Host-Cellular Subsystems

PubMed Central

MacPherson, Jamie I.; Dickerson, Jonathan E.; Pinney, John W.; Robertson, David L.

2010-01-01

Human immunodeficiency virus type 1 (HIV-1) exploits a diverse array of host cell functions in order to replicate. This is mediated through a network of virus-host interactions. A variety of recent studies have catalogued this information. In particular the HIV-1, Human Protein Interaction Database (HHPID) has provided a unique depth of protein interaction detail. However, as a map of HIV-1 infection, the HHPID is problematic, as it contains curation error and redundancy; in addition, it is based on a heterogeneous set of experimental methods. Based on identifying shared patterns of HIV-host interaction, we have developed a novel methodology to delimit the core set of host-cellular functions and their associated perturbation from the HHPID. Initially, using biclustering, we identify 279 significant sets of host proteins that undergo the same types of interaction. The functional cohesiveness of these protein sets was validated using a human protein-protein interaction network, gene ontology annotation and sequence similarity. Next, using a distance measure, we group host protein sets and identify 37 distinct higher-level subsystems. We further demonstrate the biological significance of these subsystems by cross-referencing with global siRNA screens that have been used to detect host factors necessary for HIV-1 replication, and investigate the seemingly small intersect between these data sets. Our results highlight significant host-cell subsystems that are perturbed during the course of HIV-1 infection. Moreover, we characterise the patterns of interaction that contribute to these perturbations. Thus, our work disentangles the complex set of HIV-1-host protein interactions in the HHPID, reconciles these with siRNA screens and provides an accessible and interpretable map of infection. PMID:20686668

Species identification of corynebacteria by cellular fatty acid analysis.

PubMed

Van den Velde, Sandra; Lagrou, Katrien; Desmet, Koen; Wauters, Georges; Verhaegen, Jan

2006-02-01

We evaluated the usefulness of cellular fatty acid analysis for the identification of corynebacteria. Therefore, 219 well-characterized strains belonging to 21 Corynebacterium species were analyzed with the Sherlock System of MIDI (Newark, DE). Most Corynebacterium species have a qualitative different fatty acid profile. Corynebacterium coyleae (subgroup 1), Corynebacterium riegelii, Corynebacterium simulans, and Corynebacterium imitans differ only quantitatively. Corynebacterium afermentans afermentans and C. coyleae (subgroup 2) have both a similar qualitative and quantitative profile. The commercially available database (CLIN 40, MIDI) identified only one third of the 219 strains correctly at the species level. We created a new database with these 219 strains. This new database was tested with 34 clinical isolates and could identify 29 strains correctly. Strains that remained unidentified were 2 Corynebacterium aurimucosum (not included in our database), 1 C. afermentans afermentans, and 2 Corynebacterium pseudodiphtheriticum. Cellular fatty acid analysis with a self-created database can be used for the identification and differentiation of corynebacteria.

Analysis of Protein–Protein Interactions in MCF-7 and MDA-MB-231 Cell Lines Using Phthalic Acid Chemical Probes

PubMed Central

Liang, Shih-Shin; Wang, Tsu-Nai; Tsai, Eing-Mei

2014-01-01

Phthalates are a class of plasticizers that have been characterized as endocrine disrupters, and are associated with genital diseases, cardiotoxicity, hepatotoxicity, and nephrotoxicity in the GeneOntology gene/protein database. In this study, we synthesized phthalic acid chemical probes and demonstrated differing protein–protein interactions between MCF-7 cells and MDA-MB-231 breast cancer cell lines. Phthalic acid chemical probes were synthesized using silicon dioxide particle carriers, which were modified using the silanized linker 3-aminopropyl triethoxyslane (APTES). Incubation with cell lysates from breast cancer cell lines revealed interactions between phthalic acid and cellular proteins in MCF-7 and MDA-MB-231 cells. Subsequent proteomics analyses indicated 22 phthalic acid-binding proteins in both cell types, including heat shock cognate 71-kDa protein, ATP synthase subunit beta, and heat shock protein HSP 90-beta. In addition, 21 MCF-7-specific and 32 MDA-MB-231 specific phthalic acid-binding proteins were identified, including related proteasome proteins, heat shock 70-kDa protein, and NADPH dehydrogenase and ribosomal correlated proteins, ras-related proteins, and members of the heat shock protein family, respectively. PMID:25402641

Zinc Biochemistry: From a Single Zinc Enzyme to a Key Element of Life12

PubMed Central

Maret, Wolfgang

2013-01-01

The nutritional essentiality of zinc for the growth of living organisms had been recognized long before zinc biochemistry began with the discovery of zinc in carbonic anhydrase in 1939. Painstaking analytical work then demonstrated the presence of zinc as a catalytic and structural cofactor in a few hundred enzymes. In the 1980s, the field again gained momentum with the new principle of “zinc finger” proteins, in which zinc has structural functions in domains that interact with other biomolecules. Advances in structural biology and a rapid increase in the availability of gene/protein databases now made it possible to predict zinc-binding sites from metal-binding motifs detected in sequences. This procedure resulted in the definition of zinc proteomes and the remarkable estimate that the human genome encodes ∼3000 zinc proteins. More recent developments focus on the regulatory functions of zinc(II) ions in intra- and intercellular information transfer and have tantalizing implications for yet additional functions of zinc in signal transduction and cellular control. At least three dozen proteins homeostatically control the vesicular storage and subcellular distribution of zinc and the concentrations of zinc(II) ions. Novel principles emerge from quantitative investigations on how strongly zinc interacts with proteins and how it is buffered to control the remarkably low cellular and subcellular concentrations of free zinc(II) ions. It is fair to conclude that the impact of zinc for health and disease will be at least as far-reaching as that of iron. PMID:23319127

BloodChIP: a database of comparative genome-wide transcription factor binding profiles in human blood cells.

PubMed

Chacon, Diego; Beck, Dominik; Perera, Dilmi; Wong, Jason W H; Pimanda, John E

2014-01-01

The BloodChIP database (http://www.med.unsw.edu.au/CRCWeb.nsf/page/BloodChIP) supports exploration and visualization of combinatorial transcription factor (TF) binding at a particular locus in human CD34-positive and other normal and leukaemic cells or retrieval of target gene sets for user-defined combinations of TFs across one or more cell types. Increasing numbers of genome-wide TF binding profiles are being added to public repositories, and this trend is likely to continue. For the power of these data sets to be fully harnessed by experimental scientists, there is a need for these data to be placed in context and easily accessible for downstream applications. To this end, we have built a user-friendly database that has at its core the genome-wide binding profiles of seven key haematopoietic TFs in human stem/progenitor cells. These binding profiles are compared with binding profiles in normal differentiated and leukaemic cells. We have integrated these TF binding profiles with chromatin marks and expression data in normal and leukaemic cell fractions. All queries can be exported into external sites to construct TF-gene and protein-protein networks and to evaluate the association of genes with cellular processes and tissue expression.

Low temperature conditioning of garlic (Allium sativum L.) “seed” cloves induces alterations in sprouts proteome

PubMed Central

Dufoo-Hurtado, Miguel D.; Huerta-Ocampo, José Á.; Barrera-Pacheco, Alberto; Barba de la Rosa, Ana P.; Mercado-Silva, Edmundo M.

2015-01-01

Low-temperature conditioning of garlic “seed” cloves substitutes the initial climatic requirements of the crop and accelerates the cycle. We have reported that “seed” bulbs from “Coreano” variety conditioned at 5°C for 5 weeks reduces growth and plant weight as well as the crop yields and increases the synthesis of phenolic compounds and anthocyanins. Therefore, this treatment suggests a cold stress. Plant acclimation to stress is associated with deep changes in proteome composition. Since proteins are directly involved in plant stress response, proteomics studies can significantly contribute to unravel the possible relationships between protein abundance and plant stress acclimation. The aim of this work was to study the changes in the protein profiles of garlic “seed” cloves subjected to conditioning at low-temperature using proteomics approach. Two sets of garlic bulbs were used, one set was stored at room temperature (23°C), and the other was conditioned at low temperature (5°C) for 5 weeks. Total soluble proteins were extracted from sprouts of cloves and separated by two-dimensional gel electrophoresis. Protein spots showing statistically significant changes in abundance were analyzed by LC-ESI-MS/MS and identified by database search analysis using the Mascot search engine. The results revealed that low-temperature conditioning of garlic “seed” cloves causes alterations in the accumulation of proteins involved in different physiological processes such as cellular growth, antioxidative/oxidative state, macromolecules transport, protein folding and transcription regulation process. The metabolic pathways affected include protein biosynthesis and quality control system, photosynthesis, photorespiration, energy production, and carbohydrate and nucleotide metabolism. These processes can work cooperatively to establish a new cellular homeostasis that might be related with the physiological and biochemical changes observed in previous studies. PMID:26029231

Low temperature conditioning of garlic (Allium sativum L.) "seed" cloves induces alterations in sprouts proteome.

PubMed

Dufoo-Hurtado, Miguel D; Huerta-Ocampo, José Á; Barrera-Pacheco, Alberto; Barba de la Rosa, Ana P; Mercado-Silva, Edmundo M

2015-01-01

Low-temperature conditioning of garlic "seed" cloves substitutes the initial climatic requirements of the crop and accelerates the cycle. We have reported that "seed" bulbs from "Coreano" variety conditioned at 5°C for 5 weeks reduces growth and plant weight as well as the crop yields and increases the synthesis of phenolic compounds and anthocyanins. Therefore, this treatment suggests a cold stress. Plant acclimation to stress is associated with deep changes in proteome composition. Since proteins are directly involved in plant stress response, proteomics studies can significantly contribute to unravel the possible relationships between protein abundance and plant stress acclimation. The aim of this work was to study the changes in the protein profiles of garlic "seed" cloves subjected to conditioning at low-temperature using proteomics approach. Two sets of garlic bulbs were used, one set was stored at room temperature (23°C), and the other was conditioned at low temperature (5°C) for 5 weeks. Total soluble proteins were extracted from sprouts of cloves and separated by two-dimensional gel electrophoresis. Protein spots showing statistically significant changes in abundance were analyzed by LC-ESI-MS/MS and identified by database search analysis using the Mascot search engine. The results revealed that low-temperature conditioning of garlic "seed" cloves causes alterations in the accumulation of proteins involved in different physiological processes such as cellular growth, antioxidative/oxidative state, macromolecules transport, protein folding and transcription regulation process. The metabolic pathways affected include protein biosynthesis and quality control system, photosynthesis, photorespiration, energy production, and carbohydrate and nucleotide metabolism. These processes can work cooperatively to establish a new cellular homeostasis that might be related with the physiological and biochemical changes observed in previous studies.

Insights into salicylic acid responses in cucumber (Cucumis sativus L.) cotyledons based on a comparative proteomic analysis.

PubMed

Hao, J H; Dong, C J; Zhang, Z G; Wang, X L; Shang, Q M

2012-05-01

To investigate the response of cucumber seedlings to exogenous salicylic acid (SA) and gain a better understanding of SA action mechanism, we generated a proteomic profile of cucumber (Cucumis sativus L.) cotyledons treated with exogenous SA. Analysis of 1500 protein spots from each gel revealed 63 differentially expressed proteins, 59 of which were identified successfully. Of the identified proteins, 97% matched cucumber proteins using a whole cucumber protein database based on the newly completed genome established by our laboratory. The identified proteins were involved in various cellular responses and metabolic processes, including antioxidative reactions, cell defense, photosynthesis, carbohydrate metabolism, respiration and energy homeostasis, protein folding and biosynthesis. The two largest functional categories included proteins involved in antioxidative reactions (23.7%) and photosynthesis (18.6%). Furthermore, the SA-responsive protein interaction network revealed 13 key proteins, suggesting that the expression changes of these proteins could be critical for SA-induced resistance. An analysis of these changes suggested that SA-induced resistance and seedling growth might be regulated in part through pathways involving antioxidative reactions and photosynthesis. © 2012 Elsevier Ireland Ltd. All rights reserved.

Human cell structure-driven model construction for predicting protein subcellular location from biological images.

PubMed

Shao, Wei; Liu, Mingxia; Zhang, Daoqiang

2016-01-01

The systematic study of subcellular location pattern is very important for fully characterizing the human proteome. Nowadays, with the great advances in automated microscopic imaging, accurate bioimage-based classification methods to predict protein subcellular locations are highly desired. All existing models were constructed on the independent parallel hypothesis, where the cellular component classes are positioned independently in a multi-class classification engine. The important structural information of cellular compartments is missed. To deal with this problem for developing more accurate models, we proposed a novel cell structure-driven classifier construction approach (SC-PSorter) by employing the prior biological structural information in the learning model. Specifically, the structural relationship among the cellular components is reflected by a new codeword matrix under the error correcting output coding framework. Then, we construct multiple SC-PSorter-based classifiers corresponding to the columns of the error correcting output coding codeword matrix using a multi-kernel support vector machine classification approach. Finally, we perform the classifier ensemble by combining those multiple SC-PSorter-based classifiers via majority voting. We evaluate our method on a collection of 1636 immunohistochemistry images from the Human Protein Atlas database. The experimental results show that our method achieves an overall accuracy of 89.0%, which is 6.4% higher than the state-of-the-art method. The dataset and code can be downloaded from https://github.com/shaoweinuaa/. dqzhang@nuaa.edu.cn Supplementary data are available at Bioinformatics online. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Competing endogenous RNA and interactome bioinformatic analyses on human telomerase.

PubMed

Arancio, Walter; Pizzolanti, Giuseppe; Genovese, Swonild Ilenia; Baiamonte, Concetta; Giordano, Carla

2014-04-01

We present a classic interactome bioinformatic analysis and a study on competing endogenous (ce) RNAs for hTERT. The hTERT gene codes for the catalytic subunit and limiting component of the human telomerase complex. Human telomerase reverse transcriptase (hTERT) is essential for the integrity of telomeres. Telomere dysfunctions have been widely reported to be involved in aging, cancer, and cellular senescence. The hTERT gene network has been analyzed using the BioGRID interaction database (http://thebiogrid.org/) and related analysis tools such as Osprey (http://biodata.mshri.on.ca/osprey/servlet/Index) and GeneMANIA (http://genemania.org/). The network of interaction of hTERT transcripts has been further analyzed following the competing endogenous (ce) RNA hypotheses (messenger [m] RNAs cross-talk via micro [mi] RNAs) using the miRWalk database and tools (www.ma.uni-heidelberg.de/apps/zmf/mirwalk/). These analyses suggest a role for Akt, nuclear factor-κB (NF-κB), heat shock protein 90 (HSP90), p70/p80 autoantigen, 14-3-3 proteins, and dynein in telomere functions. Roles for histone acetylation/deacetylation and proteoglycan metabolism are also proposed.

A dehydration-inducible gene in the truffle Tuber borchii identifies a novel group of dehydrins

PubMed Central

Abba', Simona; Ghignone, Stefano; Bonfante, Paola

2006-01-01

Background The expressed sequence tag M6G10 was originally isolated from a screening for differentially expressed transcripts during the reproductive stage of the white truffle Tuber borchii. mRNA levels for M6G10 increased dramatically during fruiting body maturation compared to the vegetative mycelial stage. Results Bioinformatics tools, phylogenetic analysis and expression studies were used to support the hypothesis that this sequence, named TbDHN1, is the first dehydrin (DHN)-like coding gene isolated in fungi. Homologs of this gene, all defined as "coding for hypothetical proteins" in public databases, were exclusively found in ascomycetous fungi and in plants. Although complete (or almost complete) fungal genomes and EST collections of some Basidiomycota and Glomeromycota are already available, DHN-like proteins appear to be represented only in Ascomycota. A new and previously uncharacterized conserved signature pattern was identified and proposed to Uniprot database as the main distinguishing feature of this new group of DHNs. Expression studies provide experimental evidence of a transcript induction of TbDHN1 during cellular dehydration. Conclusion Expression pattern and sequence similarities to known plant DHNs indicate that TbDHN1 is the first characterized DHN-like protein in fungi. The high similarity of TbDHN1 with homolog coding sequences implies the existence of a novel fungal/plant group of LEA Class II proteins characterized by a previously undescribed signature pattern. PMID:16512918

Protein accounting in the cellular economy.

PubMed

Vázquez-Laslop, Nora; Mankin, Alexander S

2014-04-24

Knowing the copy number of cellular proteins is critical for understanding cell physiology. By being able to measure the absolute synthesis rates of the majority of cellular proteins, Li et al. gain insights into key aspects of translation regulation and fundamental principles of cellular strategies to adjust protein synthesis according to the functional needs. Copyright © 2014 Elsevier Inc. All rights reserved.

Systematic reconstruction of TRANSPATH data into Cell System Markup Language

PubMed Central

Nagasaki, Masao; Saito, Ayumu; Li, Chen; Jeong, Euna; Miyano, Satoru

2008-01-01

Background Many biological repositories store information based on experimental study of the biological processes within a cell, such as protein-protein interactions, metabolic pathways, signal transduction pathways, or regulations of transcription factors and miRNA. Unfortunately, it is difficult to directly use such information when generating simulation-based models. Thus, modeling rules for encoding biological knowledge into system-dynamics-oriented standardized formats would be very useful for fully understanding cellular dynamics at the system level. Results We selected the TRANSPATH database, a manually curated high-quality pathway database, which provides a plentiful source of cellular events in humans, mice, and rats, collected from over 31,500 publications. In this work, we have developed 16 modeling rules based on hybrid functional Petri net with extension (HFPNe), which is suitable for graphical representing and simulating biological processes. In the modeling rules, each Petri net element is incorporated with Cell System Ontology to enable semantic interoperability of models. As a formal ontology for biological pathway modeling with dynamics, CSO also defines biological terminology and corresponding icons. By combining HFPNe with the CSO features, it is possible to make TRANSPATH data to simulation-based and semantically valid models. The results are encoded into a biological pathway format, Cell System Markup Language (CSML), which eases the exchange and integration of biological data and models. Conclusion By using the 16 modeling rules, 97% of the reactions in TRANSPATH are converted into simulation-based models represented in CSML. This reconstruction demonstrates that it is possible to use our rules to generate quantitative models from static pathway descriptions. PMID:18570683

Systematic reconstruction of TRANSPATH data into cell system markup language.

PubMed

Nagasaki, Masao; Saito, Ayumu; Li, Chen; Jeong, Euna; Miyano, Satoru

2008-06-23

Many biological repositories store information based on experimental study of the biological processes within a cell, such as protein-protein interactions, metabolic pathways, signal transduction pathways, or regulations of transcription factors and miRNA. Unfortunately, it is difficult to directly use such information when generating simulation-based models. Thus, modeling rules for encoding biological knowledge into system-dynamics-oriented standardized formats would be very useful for fully understanding cellular dynamics at the system level. We selected the TRANSPATH database, a manually curated high-quality pathway database, which provides a plentiful source of cellular events in humans, mice, and rats, collected from over 31,500 publications. In this work, we have developed 16 modeling rules based on hybrid functional Petri net with extension (HFPNe), which is suitable for graphical representing and simulating biological processes. In the modeling rules, each Petri net element is incorporated with Cell System Ontology to enable semantic interoperability of models. As a formal ontology for biological pathway modeling with dynamics, CSO also defines biological terminology and corresponding icons. By combining HFPNe with the CSO features, it is possible to make TRANSPATH data to simulation-based and semantically valid models. The results are encoded into a biological pathway format, Cell System Markup Language (CSML), which eases the exchange and integration of biological data and models. By using the 16 modeling rules, 97% of the reactions in TRANSPATH are converted into simulation-based models represented in CSML. This reconstruction demonstrates that it is possible to use our rules to generate quantitative models from static pathway descriptions.

Feature selection and classification of protein-protein complexes based on their binding affinities using machine learning approaches.

PubMed

Yugandhar, K; Gromiha, M Michael

2014-09-01

Protein-protein interactions are intrinsic to virtually every cellular process. Predicting the binding affinity of protein-protein complexes is one of the challenging problems in computational and molecular biology. In this work, we related sequence features of protein-protein complexes with their binding affinities using machine learning approaches. We set up a database of 185 protein-protein complexes for which the interacting pairs are heterodimers and their experimental binding affinities are available. On the other hand, we have developed a set of 610 features from the sequences of protein complexes and utilized Ranker search method, which is the combination of Attribute evaluator and Ranker method for selecting specific features. We have analyzed several machine learning algorithms to discriminate protein-protein complexes into high and low affinity groups based on their Kd values. Our results showed a 10-fold cross-validation accuracy of 76.1% with the combination of nine features using support vector machines. Further, we observed accuracy of 83.3% on an independent test set of 30 complexes. We suggest that our method would serve as an effective tool for identifying the interacting partners in protein-protein interaction networks and human-pathogen interactions based on the strength of interactions. © 2014 Wiley Periodicals, Inc.

Endocytosis and membrane receptor internalization: implication of F-BAR protein Carom.

PubMed

Xu, Yanjie; Xia, Jixiang; Liu, Suxuan; Stein, Sam; Ramon, Cueto; Xi, Hang; Wang, Luqiao; Xiong, Xinyu; Zhang, Lixiao; He, Dingwen; Yang, William; Zhao, Xianxian; Cheng, Xiaoshu; Yang, Xiaofeng; Wang, Hong

2017-03-01

Endocytosis is a cellular process mostly responsible for membrane receptor internalization. Cell membrane receptors bind to their ligands and form a complex which can be internalized. We previously proposed that F-BAR protein initiates membrane curvature and mediates endocytosis via its binding partners. However, F-BAR protein partners involved in membrane receptor endocytosis and the regulatory mechanism remain unknown. In this study, we established database mining strategies to explore mechanisms underlying receptor-related endocytosis. We identified 34 endocytic membrane receptors and 10 regulating proteins in clathrin-dependent endocytosis (CDE), a major process of membrane receptor internalization. We found that F-BAR protein FCHSD2 (Carom) may facilitate endocytosis via 9 endocytic partners. Carom is highly expressed, along with highly expressed endocytic membrane receptors and partners, in endothelial cells and macrophages. We established 3 models of Carom-receptor complexes and their intracellular trafficking based on protein interaction and subcellular localization. We conclude that Carom may mediate receptor endocytosis and transport endocytic receptors to the cytoplasm for receptor signaling and lysosome/proteasome degradation, or to the nucleus for RNA processing, gene transcription and DNA repair.

Identification of giant Mimivirus protein functions using RNA interference

PubMed Central

Sobhy, Haitham; Scola, Bernard La; Pagnier, Isabelle; Raoult, Didier; Colson, Philippe

2015-01-01

Genomic analysis of giant viruses, such as Mimivirus, has revealed that more than half of the putative genes have no known functions (ORFans). We knocked down Mimivirus genes using short interfering RNA as a proof of concept to determine the functions of giant virus ORFans. As fibers are easy to observe, we targeted a gene encoding a protein absent in a Mimivirus mutant devoid of fibers as well as three genes encoding products identified in a protein concentrate of fibers, including one ORFan and one gene of unknown function. We found that knocking down these four genes was associated with depletion or modification of the fibers. Our strategy of silencing ORFan genes in giant viruses opens a way to identify its complete gene repertoire and may clarify the role of these genes, differentiating between junk DNA and truly used genes. Using this strategy, we were able to annotate four proteins in Mimivirus and 30 homologous proteins in other giant viruses. In addition, we were able to annotate >500 proteins from cellular organisms and 100 from metagenomic databases. PMID:25972846

Comprehensive comparative analysis and identification of RNA-binding protein domains: multi-class classification and feature selection.

PubMed

Jahandideh, Samad; Srinivasasainagendra, Vinodh; Zhi, Degui

2012-11-07

RNA-protein interaction plays an important role in various cellular processes, such as protein synthesis, gene regulation, post-transcriptional gene regulation, alternative splicing, and infections by RNA viruses. In this study, using Gene Ontology Annotated (GOA) and Structural Classification of Proteins (SCOP) databases an automatic procedure was designed to capture structurally solved RNA-binding protein domains in different subclasses. Subsequently, we applied tuned multi-class SVM (TMCSVM), Random Forest (RF), and multi-class ℓ1/ℓq-regularized logistic regression (MCRLR) for analysis and classifying RNA-binding protein domains based on a comprehensive set of sequence and structural features. In this study, we compared prediction accuracy of three different state-of-the-art predictor methods. From our results, TMCSVM outperforms the other methods and suggests the potential of TMCSVM as a useful tool for facilitating the multi-class prediction of RNA-binding protein domains. On the other hand, MCRLR by elucidating importance of features for their contribution in predictive accuracy of RNA-binding protein domains subclasses, helps us to provide some biological insights into the roles of sequences and structures in protein-RNA interactions.

Identification of host cellular proteins that interact with the M protein of a highly pathogenic porcine reproductive and respiratory syndrome virus vaccine strain.

PubMed

Wang, Qian; Li, Yanwei; Dong, Hong; Wang, Li; Peng, Jinmei; An, Tongqing; Yang, Xufu; Tian, Zhijun; Cai, Xuehui

2017-02-22

The highly pathogenic porcine reproductive and respiratory syndrome virus (HP-PRRSV) continues to pose one of the greatest threats to the swine industry. M protein is the most conserved and important structural protein of PRRSV. However, information about the host cellular proteins that interact with M protein remains limited. Host cellular proteins that interact with the M protein of HP-PRRSV were immunoprecipitated from MARC-145 cells infected with PRRSV HuN4-F112 using the M monoclonal antibody (mAb). The differentially expressed proteins were identified by LC-MS/MS. The screened proteins were used for bioinformatics analysis including Gene Ontology, the interaction network, and the enriched KEGG pathways. Some interested cellular proteins were validated to interact with M protein by CO-IP. The PRRSV HuN4-F112 infection group had 10 bands compared with the control group. The bands included 219 non-redundant cellular proteins that interact with M protein, which were identified by LC-MS/MS with high confidence. The gene ontology and Kyoto encyclopedia of genes and genomes (KEGG) pathway bioinformatic analyses indicated that the identified proteins could be assigned to several different subcellular locations and functional classes. Functional analysis of the interactome profile highlighted cellular pathways associated with protein translation, infectious disease, and signal transduction. Two interested cellular proteins-nuclear factor of activated T cells 45 kDa (NF45) and proliferating cell nuclear antigen (PCNA)-that could interact with M protein were validated by Co-IP and confocal analyses. The interactome data between PRRSV M protein and cellular proteins were identified and contribute to the understanding of the roles of M protein in the replication and pathogenesis of PRRSV. The interactome of M protein will aid studies of virus/host interactions and provide means to decrease the threat of PRRSV to the swine industry in the future.

BIG: a large-scale data integration tool for renal physiology.

PubMed

Zhao, Yue; Yang, Chin-Rang; Raghuram, Viswanathan; Parulekar, Jaya; Knepper, Mark A

2016-10-01

Due to recent advances in high-throughput techniques, we and others have generated multiple proteomic and transcriptomic databases to describe and quantify gene expression, protein abundance, or cellular signaling on the scale of the whole genome/proteome in kidney cells. The existence of so much data from diverse sources raises the following question: "How can researchers find information efficiently for a given gene product over all of these data sets without searching each data set individually?" This is the type of problem that has motivated the "Big-Data" revolution in Data Science, which has driven progress in fields such as marketing. Here we present an online Big-Data tool called BIG (Biological Information Gatherer) that allows users to submit a single online query to obtain all relevant information from all indexed databases. BIG is accessible at http://big.nhlbi.nih.gov/.

In silico re-identification of properties of drug target proteins.

PubMed

Kim, Baeksoo; Jo, Jihoon; Han, Jonghyun; Park, Chungoo; Lee, Hyunju

2017-05-31

Computational approaches in the identification of drug targets are expected to reduce time and effort in drug development. Advances in genomics and proteomics provide the opportunity to uncover properties of druggable genomes. Although several studies have been conducted for distinguishing drug targets from non-drug targets, they mainly focus on the sequences and functional roles of proteins. Many other properties of proteins have not been fully investigated. Using the DrugBank (version 3.0) database containing nearly 6,816 drug entries including 760 FDA-approved drugs and 1822 of their targets and human UniProt/Swiss-Prot databases, we defined 1578 non-redundant drug target and 17,575 non-drug target proteins. To select these non-redundant protein datasets, we built four datasets (A, B, C, and D) by considering clustering of paralogous proteins. We first reassessed the widely used properties of drug target proteins. We confirmed and extended that drug target proteins (1) are likely to have more hydrophobic, less polar, less PEST sequences, and more signal peptide sequences higher and (2) are more involved in enzyme catalysis, oxidation and reduction in cellular respiration, and operational genes. In this study, we proposed new properties (essentiality, expression pattern, PTMs, and solvent accessibility) for effectively identifying drug target proteins. We found that (1) drug targetability and protein essentiality are decoupled, (2) druggability of proteins has high expression level and tissue specificity, and (3) functional post-translational modification residues are enriched in drug target proteins. In addition, to predict the drug targetability of proteins, we exploited two machine learning methods (Support Vector Machine and Random Forest). When we predicted drug targets by combining previously known protein properties and proposed new properties, an F-score of 0.8307 was obtained. When the newly proposed properties are integrated, the prediction performance is improved and these properties are related to drug targets. We believe that our study will provide a new aspect in inferring drug-target interactions.

A proteomic chronology of gene expression through the cell cycle in human myeloid leukemia cells.

PubMed

Ly, Tony; Ahmad, Yasmeen; Shlien, Adam; Soroka, Dominique; Mills, Allie; Emanuele, Michael J; Stratton, Michael R; Lamond, Angus I

2014-01-01

Technological advances have enabled the analysis of cellular protein and RNA levels with unprecedented depth and sensitivity, allowing for an unbiased re-evaluation of gene regulation during fundamental biological processes. Here, we have chronicled the dynamics of protein and mRNA expression levels across a minimally perturbed cell cycle in human myeloid leukemia cells using centrifugal elutriation combined with mass spectrometry-based proteomics and RNA-Seq, avoiding artificial synchronization procedures. We identify myeloid-specific gene expression and variations in protein abundance, isoform expression and phosphorylation at different cell cycle stages. We dissect the relationship between protein and mRNA levels for both bulk gene expression and for over ∼6000 genes individually across the cell cycle, revealing complex, gene-specific patterns. This data set, one of the deepest surveys to date of gene expression in human cells, is presented in an online, searchable database, the Encyclopedia of Proteome Dynamics (http://www.peptracker.com/epd/). DOI: http://dx.doi.org/10.7554/eLife.01630.001.

Global Proteomics Analysis of the Response to Starvation in C. elegans*

PubMed Central

Larance, Mark; Pourkarimi, Ehsan; Wang, Bin; Brenes Murillo, Alejandro; Kent, Robert; Lamond, Angus I.; Gartner, Anton

2015-01-01

Periodic starvation of animals induces large shifts in metabolism but may also influence many other cellular systems and can lead to adaption to prolonged starvation conditions. To date, there is limited understanding of how starvation affects gene expression, particularly at the protein level. Here, we have used mass-spectrometry-based quantitative proteomics to identify global changes in the Caenorhabditis elegans proteome due to acute starvation of young adult animals. Measuring changes in the abundance of over 5,000 proteins, we show that acute starvation rapidly alters the levels of hundreds of proteins, many involved in central metabolic pathways, highlighting key regulatory responses. Surprisingly, we also detect changes in the abundance of chromatin-associated proteins, including specific linker histones, histone variants, and histone posttranslational modifications associated with the epigenetic control of gene expression. To maximize community access to these data, they are presented in an online searchable database, the Encyclopedia of Proteome Dynamics (http://www.peptracker.com/epd/). PMID:25963834

A proteomic chronology of gene expression through the cell cycle in human myeloid leukemia cells

PubMed Central

Ly, Tony; Ahmad, Yasmeen; Shlien, Adam; Soroka, Dominique; Mills, Allie; Emanuele, Michael J; Stratton, Michael R; Lamond, Angus I

2014-01-01

Technological advances have enabled the analysis of cellular protein and RNA levels with unprecedented depth and sensitivity, allowing for an unbiased re-evaluation of gene regulation during fundamental biological processes. Here, we have chronicled the dynamics of protein and mRNA expression levels across a minimally perturbed cell cycle in human myeloid leukemia cells using centrifugal elutriation combined with mass spectrometry-based proteomics and RNA-Seq, avoiding artificial synchronization procedures. We identify myeloid-specific gene expression and variations in protein abundance, isoform expression and phosphorylation at different cell cycle stages. We dissect the relationship between protein and mRNA levels for both bulk gene expression and for over ∼6000 genes individually across the cell cycle, revealing complex, gene-specific patterns. This data set, one of the deepest surveys to date of gene expression in human cells, is presented in an online, searchable database, the Encyclopedia of Proteome Dynamics (http://www.peptracker.com/epd/). DOI: http://dx.doi.org/10.7554/eLife.01630.001 PMID:24596151

MitoRes: a resource of nuclear-encoded mitochondrial genes and their products in Metazoa.

PubMed

Catalano, Domenico; Licciulli, Flavio; Turi, Antonio; Grillo, Giorgio; Saccone, Cecilia; D'Elia, Domenica

2006-01-24

Mitochondria are sub-cellular organelles that have a central role in energy production and in other metabolic pathways of all eukaryotic respiring cells. In the last few years, with more and more genomes being sequenced, a huge amount of data has been generated providing an unprecedented opportunity to use the comparative analysis approach in studies of evolution and functional genomics with the aim of shedding light on molecular mechanisms regulating mitochondrial biogenesis and metabolism. In this context, the problem of the optimal extraction of representative datasets of genomic and proteomic data assumes a crucial importance. Specialised resources for nuclear-encoded mitochondria-related proteins already exist; however, no mitochondrial database is currently available with the same features of MitoRes, which is an update of the MitoNuc database extensively modified in its structure, data sources and graphical interface. It contains data on nuclear-encoded mitochondria-related products for any metazoan species for which this type of data is available and also provides comprehensive sequence datasets (gene, transcript and protein) as well as useful tools for their extraction and export. MitoRes http://www2.ba.itb.cnr.it/MitoRes/ consolidates information from publicly external sources and automatically annotates them into a relational database. Additionally, it also clusters proteins on the basis of their sequence similarity and interconnects them with genomic data. The search engine and sequence management tools allow the query/retrieval of the database content and the extraction and export of sequences (gene, transcript, protein) and related sub-sequences (intron, exon, UTR, CDS, signal peptide and gene flanking regions) ready to be used for in silico analysis. The tool we describe here has been developed to support lab scientists and bioinformaticians alike in the characterization of molecular features and evolution of mitochondrial targeting sequences. The way it provides for the retrieval and extraction of sequences allows the user to overcome the obstacles encountered in the integrative use of different bioinformatic resources and the completeness of the sequence collection allows intra- and interspecies comparison at different biological levels (gene, transcript and protein).

Eukaryotic DING Proteins Are Endogenous: An Immunohistological Study in Mouse Tissues

PubMed Central

Collombet, Jean-Marc; Elias, Mikael; Gotthard, Guillaume; Four, Elise; Renault, Frédérique; Joffre, Aurélie; Baubichon, Dominique; Rochu, Daniel; Chabrière, Eric

2010-01-01

Background DING proteins encompass an intriguing protein family first characterized by their conserved N-terminal sequences. Some of these proteins seem to have key roles in various human diseases, e.g., rheumatoid arthritis, atherosclerosis, HIV suppression. Although this protein family seems to be ubiquitous in eukaryotes, their genes are consistently lacking from genomic databases. Such a lack has considerably hampered functional studies and has fostered therefore the hypothesis that DING proteins isolated from eukaryotes were in fact prokaryotic contaminants. Principal Findings In the framework of our study, we have performed a comprehensive immunological detection of DING proteins in mice. We demonstrate that DING proteins are present in all tissues tested as isoforms of various molecular weights (MWs). Their intracellular localization is tissue-dependant, being exclusively nuclear in neurons, but cytoplasmic and nuclear in other tissues. We also provide evidence that germ-free mouse plasma contains as much DING protein as wild-type. Significance Hence, data herein provide a valuable basis for future investigations aimed at eukaryotic DING proteins, revealing that these proteins seem ubiquitous in mouse tissue. Our results strongly suggest that mouse DING proteins are endogenous. Moreover, the determination in this study of the precise cellular localization of DING proteins constitute a precious evidence to understand their molecular involvements in their related human diseases. PMID:20161715

Evolution and function of CAG/polyglutamine repeats in protein–protein interaction networks

PubMed Central

Schaefer, Martin H.; Wanker, Erich E.; Andrade-Navarro, Miguel A.

2012-01-01

Expanded runs of consecutive trinucleotide CAG repeats encoding polyglutamine (polyQ) stretches are observed in the genes of a large number of patients with different genetic diseases such as Huntington's and several Ataxias. Protein aggregation, which is a key feature of most of these diseases, is thought to be triggered by these expanded polyQ sequences in disease-related proteins. However, polyQ tracts are a normal feature of many human proteins, suggesting that they have an important cellular function. To clarify the potential function of polyQ repeats in biological systems, we systematically analyzed available information stored in sequence and protein interaction databases. By integrating genomic, phylogenetic, protein interaction network and functional information, we obtained evidence that polyQ tracts in proteins stabilize protein interactions. This happens most likely through structural changes whereby the polyQ sequence extends a neighboring coiled-coil region to facilitate its interaction with a coiled-coil region in another protein. Alteration of this important biological function due to polyQ expansion results in gain of abnormal interactions, leading to pathological effects like protein aggregation. Our analyses suggest that research on polyQ proteins should shift focus from expanded polyQ proteins into the characterization of the influence of the wild-type polyQ on protein interactions. PMID:22287626

Distinguishing between biochemical and cellular function: Are there peptide signatures for cellular function of proteins?

PubMed

Jain, Shruti; Bhattacharyya, Kausik; Bakshi, Rachit; Narang, Ankita; Brahmachari, Vani

2017-04-01

The genome annotation and identification of gene function depends on conserved biochemical activity. However, in the cell, proteins with the same biochemical function can participate in different cellular pathways and cannot complement one another. Similarly, two proteins of very different biochemical functions are put in the same class of cellular function; for example, the classification of a gene as an oncogene or a tumour suppressor gene is not related to its biochemical function, but is related to its cellular function. We have taken an approach to identify peptide signatures for cellular function in proteins with known biochemical function. ATPases as a test case, we classified ATPases (2360 proteins) and kinases (517 proteins) from the human genome into different cellular function categories such as transcriptional, replicative, and chromatin remodelling proteins. Using publicly available tool, MEME, we identify peptide signatures shared among the members of a given category but not between cellular functional categories; for example, no motif sharing is seen between chromatin remodelling and transporter ATPases, similarly between receptor Serine/Threonine Kinase and Receptor Tyrosine Kinase. There are motifs shared within each category with significant E value and high occurrence. This concept of signature for cellular function was applied to developmental regulators, the polycomb and trithorax proteins which led to the prediction of the role of INO80, a chromatin remodelling protein, in development. This has been experimentally validated earlier for its role in homeotic gene regulation and its interaction with regulatory complexes like the Polycomb and Trithorax complex. Proteins 2017; 85:682-693. © 2016 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

A novel method for identifying disease associated protein complexes based on functional similarity protein complex networks.

PubMed

Le, Duc-Hau

2015-01-01

Protein complexes formed by non-covalent interaction among proteins play important roles in cellular functions. Computational and purification methods have been used to identify many protein complexes and their cellular functions. However, their roles in terms of causing disease have not been well discovered yet. There exist only a few studies for the identification of disease-associated protein complexes. However, they mostly utilize complicated heterogeneous networks which are constructed based on an out-of-date database of phenotype similarity network collected from literature. In addition, they only apply for diseases for which tissue-specific data exist. In this study, we propose a method to identify novel disease-protein complex associations. First, we introduce a framework to construct functional similarity protein complex networks where two protein complexes are functionally connected by either shared protein elements, shared annotating GO terms or based on protein interactions between elements in each protein complex. Second, we propose a simple but effective neighborhood-based algorithm, which yields a local similarity measure, to rank disease candidate protein complexes. Comparing the predictive performance of our proposed algorithm with that of two state-of-the-art network propagation algorithms including one we used in our previous study, we found that it performed statistically significantly better than that of these two algorithms for all the constructed functional similarity protein complex networks. In addition, it ran about 32 times faster than these two algorithms. Moreover, our proposed method always achieved high performance in terms of AUC values irrespective of the ways to construct the functional similarity protein complex networks and the used algorithms. The performance of our method was also higher than that reported in some existing methods which were based on complicated heterogeneous networks. Finally, we also tested our method with prostate cancer and selected the top 100 highly ranked candidate protein complexes. Interestingly, 69 of them were evidenced since at least one of their protein elements are known to be associated with prostate cancer. Our proposed method, including the framework to construct functional similarity protein complex networks and the neighborhood-based algorithm on these networks, could be used for identification of novel disease-protein complex associations.

PodNet, a protein-protein interaction network of the podocyte.

PubMed

Warsow, Gregor; Endlich, Nicole; Schordan, Eric; Schordan, Sandra; Chilukoti, Ravi K; Homuth, Georg; Moeller, Marcus J; Fuellen, Georg; Endlich, Karlhans

2013-07-01

Interactions between proteins crucially determine cellular structure and function. Differential analysis of the interactome may help elucidate molecular mechanisms during disease development; however, this analysis necessitates mapping of expression data on protein-protein interaction networks. These networks do not exist for the podocyte; therefore, we built PodNet, a literature-based mouse podocyte network in Cytoscape format. Using database protein-protein interactions, we expanded PodNet to XPodNet with enhanced connectivity. In order to test the performance of XPodNet in differential interactome analysis, we examined podocyte developmental differentiation and the effect of cell culture. Transcriptomes of podocytes in 10 different states were mapped on XPodNet and analyzed with the Cytoscape plugin ExprEssence, based on the law of mass action. Interactions between slit diaphragm proteins are most significantly upregulated during podocyte development and most significantly downregulated in culture. On the other hand, our analysis revealed that interactions lost during podocyte differentiation are not regained in culture, suggesting a loss rather than a reversal of differentiation for podocytes in culture. Thus, we have developed PodNet as a valuable tool for differential interactome analysis in podocytes, and we have identified established and unexplored regulated interactions in developing and cultured podocytes.

PANDORA: keyword-based analysis of protein sets by integration of annotation sources.

PubMed

Kaplan, Noam; Vaaknin, Avishay; Linial, Michal

2003-10-01

Recent advances in high-throughput methods and the application of computational tools for automatic classification of proteins have made it possible to carry out large-scale proteomic analyses. Biological analysis and interpretation of sets of proteins is a time-consuming undertaking carried out manually by experts. We have developed PANDORA (Protein ANnotation Diagram ORiented Analysis), a web-based tool that provides an automatic representation of the biological knowledge associated with any set of proteins. PANDORA uses a unique approach of keyword-based graphical analysis that focuses on detecting subsets of proteins that share unique biological properties and the intersections of such sets. PANDORA currently supports SwissProt keywords, NCBI Taxonomy, InterPro entries and the hierarchical classification terms from ENZYME, SCOP and GO databases. The integrated study of several annotation sources simultaneously allows a representation of biological relations of structure, function, cellular location, taxonomy, domains and motifs. PANDORA is also integrated into the ProtoNet system, thus allowing testing thousands of automatically generated clusters. We illustrate how PANDORA enhances the biological understanding of large, non-uniform sets of proteins originating from experimental and computational sources, without the need for prior biological knowledge on individual proteins.

A maize database resource that captures tissue-specific and subcellular-localized gene expression, via fluorescent tags and confocal imaging (Maize Cell Genomics Database).

PubMed

Krishnakumar, Vivek; Choi, Yongwook; Beck, Erin; Wu, Qingyu; Luo, Anding; Sylvester, Anne; Jackson, David; Chan, Agnes P

2015-01-01

Maize is a global crop and a powerful system among grain crops for genetic and genomic studies. However, the development of novel biological tools and resources to aid in the functional identification of gene sequences is greatly needed. Towards this goal, we have developed a collection of maize marker lines for studying native gene expression in specific cell types and subcellular compartments using fluorescent proteins (FPs). To catalog FP expression, we have developed a public repository, the Maize Cell Genomics (MCG) Database, (http://maize.jcvi.org/cellgenomics), to organize a large data set of confocal images generated from the maize marker lines. To date, the collection represents major subcellular structures and also developmentally important progenitor cell populations. The resource is available to the research community, for example to study protein localization or interactions under various experimental conditions or mutant backgrounds. A subset of the marker lines can also be used to induce misexpression of target genes through a transactivation system. For future directions, the image repository can be expanded to accept new image submissions from the research community, and to perform customized large-scale computational image analysis. This community resource will provide a suite of new tools for gaining biological insights by following the dynamics of protein expression at the subcellular, cellular and tissue levels. © The Author 2014. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.

Cellular Strategies of Protein Quality Control

PubMed Central

Chen, Bryan; Retzlaff, Marco; Roos, Thomas; Frydman, Judith

2011-01-01

Eukaryotic cells must contend with a continuous stream of misfolded proteins that compromise the cellular protein homeostasis balance and jeopardize cell viability. An elaborate network of molecular chaperones and protein degradation factors continually monitor and maintain the integrity of the proteome. Cellular protein quality control relies on three distinct yet interconnected strategies whereby misfolded proteins can either be refolded, degraded, or delivered to distinct quality control compartments that sequester potentially harmful misfolded species. Molecular chaperones play a critical role in determining the fate of misfolded proteins in the cell. Here, we discuss the spatial and temporal organization of cellular quality control strategies and their implications for human diseases linked to protein misfolding and aggregation. PMID:21746797

Identification of learning and memory genes in canine; promoter investigation and determining the selective pressure.

PubMed

Seifi Moroudi, Reihane; Masoudi, Ali Akbar; Vaez Torshizi, Rasoul; Zandi, Mohammad

2014-12-01

One of the important behaviors of dogs is trainability which is affected by learning and memory genes. These kinds of the genes have not yet been identified in dogs. In the current research, these genes were found in animal models by mining the biological data and scientific literatures. The proteins of these genes were obtained from the UniProt database in dogs and humans. Not all homologous proteins perform similar functions, thus comparison of these proteins was studied in terms of protein families, domains, biological processes, molecular functions, and cellular location of metabolic pathways in Interpro, KEGG, Quick Go and Psort databases. The results showed that some of these proteins have the same performance in the rat or mouse, dog, and human. It is anticipated that the protein of these genes may be effective in learning and memory in dogs. Then, the expression pattern of the recognized genes was investigated in the dog hippocampus using the existing information in the GEO profile. The results showed that BDNF, TAC1 and CCK genes are expressed in the dog hippocampus, therefore, these genes could be strong candidates associated with learning and memory in dogs. Subsequently, due to the importance of the promoter regions in gene function, this region was investigated in the above genes. Analysis of the promoter indicated that the HNF-4 site of BDNF gene and the transcription start site of CCK gene is exposed to methylation. Phylogenetic analysis of protein sequences of these genes showed high similarity in each of these three genes among the studied species. The dN/dS ratio for BDNF, TAC1 and CCK genes indicates a purifying selection during the evolution of the genes.

Quantitative interactome reveals that porcine reproductive and respiratory syndrome virus nonstructural protein 2 forms a complex with viral nucleocapsid protein and cellular vimentin.

PubMed

Song, Tao; Fang, Liurong; Wang, Dang; Zhang, Ruoxi; Zeng, Songlin; An, Kang; Chen, Huanchun; Xiao, Shaobo

2016-06-16

Porcine reproductive and respiratory syndrome virus (PRRSV) is an Arterivirus that has heavily impacted the global swine industry. The PRRSV nonstructural protein 2 (nsp2) plays crucial roles in viral replication and host immune regulation, most likely by interacting with viral or cellular proteins that have not yet been identified. In this study, a quantitative interactome approach based on immunoprecipitation and stable isotope labeling with amino acids in cell culture (SILAC) was performed to identify nsp2-interacting proteins in PRRSV-infected cells with an nsp2-specific monoclonal antibody. Nine viral proteins and 62 cellular proteins were identified as potential nsp2-interacting partners. Our data demonstrate that the PRRSV nsp1α, nsp1β, and nucleocapsid proteins all interact directly with nsp2. Nsp2-interacting cellular proteins were classified into different functional groups and an interactome network of nsp2 was generated. Interestingly, cellular vimentin, a known receptor for PRRSV, forms a complex with nsp2 by using viral nucleocapsid protein as an intermediate. Taken together, the nsp2 interactome under the condition of virus infection clarifies a role of nsp2 in PRRSV replication and immune evasion. Viral proteins must interact with other virus-encoded proteins and/or host cellular proteins to function, and interactome analysis is an ideal approach for identifying such interacting proteins. In this study, we used the quantitative interactome methodology to identify the viral and cellular proteins that potentially interact with the nonstructural protein 2 (nsp2) of porcine reproductive and respiratory syndrome virus (PRRSV) under virus infection conditions, thus providing a rich source of potential viral and cellular interaction partners for PRRSV nsp2. Based on the interactome data, we further demonstrated that PRRSV nsp2 and nucleocapsid protein together with cellular vimentin, form a complex that may be essential for viral attachment and replication, which partly explains the role of nsp2 in PRRSV replication and immune evasion. Copyright © 2016 Elsevier B.V. All rights reserved.

Prediction of Ras-effector interactions using position energy matrices.

PubMed

Kiel, Christina; Serrano, Luis

2007-09-01

One of the more challenging problems in biology is to determine the cellular protein interaction network. Progress has been made to predict protein-protein interactions based on structural information, assuming that structural similar proteins interact in a similar way. In a previous publication, we have determined a genome-wide Ras-effector interaction network based on homology models, with a high accuracy of predicting binding and non-binding domains. However, for a prediction on a genome-wide scale, homology modelling is a time-consuming process. Therefore, we here successfully developed a faster method using position energy matrices, where based on different Ras-effector X-ray template structures, all amino acids in the effector binding domain are sequentially mutated to all other amino acid residues and the effect on binding energy is calculated. Those pre-calculated matrices can then be used to score for binding any Ras or effector sequences. Based on position energy matrices, the sequences of putative Ras-binding domains can be scanned quickly to calculate an energy sum value. By calibrating energy sum values using quantitative experimental binding data, thresholds can be defined and thus non-binding domains can be excluded quickly. Sequences which have energy sum values above this threshold are considered to be potential binding domains, and could be further analysed using homology modelling. This prediction method could be applied to other protein families sharing conserved interaction types, in order to determine in a fast way large scale cellular protein interaction networks. Thus, it could have an important impact on future in silico structural genomics approaches, in particular with regard to increasing structural proteomics efforts, aiming to determine all possible domain folds and interaction types. All matrices are deposited in the ADAN database (http://adan-embl.ibmc.umh.es/). Supplementary data are available at Bioinformatics online.

Isolation of genes differentially expressed during development and ripening of Fragaria chiloensis fruit by suppression subtractive hybridization.

PubMed

Pimentel, Paula; Salvatierra, Ariel; Moya-León, María Alejandra; Herrera, Raúl

2010-09-15

Fragaria chiloensis, the native Chilean strawberry, is noted for its good fruit quality characters. However, it is a highly perishable fruit due to its rapid softening. With the aim to screen for genes differentially expressed during development and ripening of strawberry fruit, the subtractive suppressive hybridization (SSH) methodology was employed. Six libraries were generated contrasting transcripts from four different developmental stages. A set of 1807 genes was isolated and characterized. In our EST collection, approximately 90% of partial cDNAs showed significant similarity to proteins with known or unknown function registered in databases. Among them, proteins related to protein fate were identified in a large green fruit library and protein related with cellular transport, cell wall-related proteins, and transcription regulators were identified in a ripe fruit library. Thirteen genes were analyzed by qRT-PCR during development and ripening of the Chilean strawberry fruit. The information generated in this study provides new clues to aid the understanding of the ripening process in F. chiloensis fruit. Copyright 2010 Elsevier GmbH. All rights reserved.

In silico study of protein to protein interaction analysis of AMP-activated protein kinase and mitochondrial activity in three different farm animal species

NASA Astrophysics Data System (ADS)

Prastowo, S.; Widyas, N.

2018-03-01

AMP-activated protein kinase (AMPK) is cellular energy censor which works based on ATP and AMP concentration. This protein interacts with mitochondria in determine its activity to generate energy for cell metabolism purposes. For that, this paper aims to compare the protein to protein interaction of AMPK and mitochondrial activity genes in the metabolism of known animal farm (domesticated) that are cattle (Bos taurus), pig (Sus scrofa) and chicken (Gallus gallus). In silico study was done using STRING V.10 as prominent protein interaction database, followed with biological function comparison in KEGG PATHWAY database. Set of genes (12 in total) were used as input analysis that are PRKAA1, PRKAA2, PRKAB1, PRKAB2, PRKAG1, PRKAG2, PRKAG3, PPARGC1, ACC, CPT1B, NRF2 and SOD. The first 7 genes belong to gene in AMPK family, while the last 5 belong to mitochondrial activity genes. The protein interaction result shows 11, 8 and 5 metabolism pathways in Bos taurus, Sus scrofa and Gallus gallus, respectively. The top pathway in Bos taurus is AMPK signaling pathway (10 genes), Sus scrofa is Adipocytokine signaling pathway (8 genes) and Gallus gallus is FoxO signaling pathway (5 genes). Moreover, the common pathways found in those 3 species are Adipocytokine signaling pathway, Insulin signaling pathway and FoxO signaling pathway. Genes clustered in Adipocytokine and Insulin signaling pathway are PRKAA2, PPARGC1A, PRKAB1 and PRKAG2. While, in FoxO signaling pathway are PRKAA2, PRKAB1, PRKAG2. According to that, we found PRKAA2, PRKAB1 and PRKAG2 are the common genes. Based on the bioinformatics analysis, we can demonstrate that protein to protein interaction shows distinct different of metabolism in different species. However, further validation is needed to give a clear explanation.

The rehydration transcriptome of the desiccation-tolerant bryophyte Tortula ruralis: transcript classification and analysis

PubMed Central

Oliver, Melvin J; Dowd, Scot E; Zaragoza, Joaquin; Mauget, Steven A; Payton, Paxton R

2004-01-01

Background The cellular response of plants to water-deficits has both economic and evolutionary importance directly affecting plant productivity in agriculture and plant survival in the natural environment. Genes induced by water-deficit stress have been successfully enumerated in plants that are relatively sensitive to cellular dehydration, however we have little knowledge as to the adaptive role of these genes in establishing tolerance to water loss at the cellular level. Our approach to address this problem has been to investigate the genetic responses of plants that are capable of tolerating extremes of dehydration, in particular the desiccation-tolerant bryophyte, Tortula ruralis. To establish a sound basis for characterizing the Tortula genome in regards to desiccation tolerance, we analyzed 10,368 expressed sequence tags (ESTs) from rehydrated rapid-dried Tortula gametophytes, a stage previously determined to exhibit the maximum stress induced change in gene expression. Results The 10, 368 ESTs formed 5,563 EST clusters (contig groups representing individual genes) of which 3,321 (59.7%) exhibited similarity to genes present in the public databases and 2,242 were categorized as unknowns based on protein homology scores. The 3,321 clusters were classified by function using the Gene Ontology (GO) hierarchy and the KEGG database. The results indicate that the transcriptome contains a diverse population of transcripts that reflects, as expected, a period of metabolic upheaval in the gametophyte cells. Much of the emphasis within the transcriptome is centered on the protein synthetic machinery, ion and metabolite transport, and membrane biosynthesis and repair. Rehydrating gametophytes also have an abundance of transcripts that code for enzymes involved in oxidative stress metabolism and phosphorylating activities. The functional classifications reflect a remarkable consistency with what we have previously established with regards to the metabolic activities that are important in the recovery of the gametophytes from desiccation. A comparison of the GO distribution of Tortula clusters with an identical analysis of 9,981 clusters from the desiccation sensitive bryophyte species Physcomitrella patens, revealed, and accentuated, the differences between stressed and unstressed transcriptomes. Cross species sequence comparisons indicated that on the whole the Tortula clusters were more closely related to those from Physcomitrella than Arabidopsis (complete genome BLASTx comparison) although because of the differences in the databases there were more high scoring matches to the Arabidopsis sequences. The most abundant transcripts contained within the Tortula ESTs encode Late Embryogenesis Abundant (LEA) proteins that are normally associated with drying plant tissues. This suggests that LEAs may also play a role in recovery from desiccation when water is reintroduced into a dried tissue. Conclusion The establishment of a rehydration EST collection for Tortula ruralis, an important plant model for plant stress responses and vegetative desiccation tolerance, is an important step in understanding the genome level response to cellular dehydration. The type of transcript analysis performed here has laid the foundation for more detailed functional and genome level analyses of the genes involved in desiccation tolerance in plants. PMID:15546486

Endocytosis and membrane receptor internalization: implication of F-BAR protein Carom

PubMed Central

Xu, Yanjie; Liu, Suxuan; Xia, Jixiang; Stein, Sam; Ramon, Cueto; Xi, Hang; Wang, Luqiao; Xiong, Xinyu; Zhang, Lixiao; He, Dingwen; Yang, William; Zhao, Xianxian; Cheng, Xiaoshu; Yang, Xiaofeng; Wang, Hong

2016-01-01

Endocytosis is a cellular process mostly responsible for membrane receptor internalization. Cell membrane receptors bind to their ligands and form a complex which can be internalized. We previously proposed that F-BAR protein initiates membrane curvature and mediates endocytosis via their binding partners. However, F-BAR protein partners involved in membrane receptor endocytosis and the regulatory mechanism remain unknown. In this study, we established a group of database mining strategies to explore mechanisms underlying receptor-related endocytosis. We identified 34 endocytic membrane receptors and 10 regulating proteins for vesicle formation in clathrin-dependent endocytosis (CDE), a major process of membrane receptor internalization. We found that F-BAR protein FCHSD2 (Carom) may facilitate endocytosis via 9 endocytic partners. Carom is highly expressed, along with highly expressed endocytic membrane receptors and partners, in endothelial cells and macrophages. We established 3 models of Carom-receptor complex and their intracellular trafficking based on protein-protein interaction and subcellular localization. We conclude that Carom may mediate receptor endocytosis and transport endocytic receptors to the cytoplasm for receptor signaling and lysosome/proteasome degradation, or to the nucleus for RNA processing, gene transcription and DNA repair. PMID:28199211

Isolation and identification of peanut leaf proteins regulated by water stress.

PubMed

Akkasaeng, Chutipong; Tantisuwichwong, Napaporn; Chairam, Issariya; Prakrongrak, Narumon; Jogloy, Sanun; Pathanothai, Aran

2007-05-15

Water deficits trigger signaling cascades leading to modulation of protein expression in plant tissues. Identification of peanut leaf proteins regulated by water stress provides some insights of cellular and molecular response of peanut plants to drought stress. Peanut variety Khon Kaen 4, a water-stress sensitive variety, was grown in a growth chamber under controlled environment. Water stress was imposed on day 30 after seedling emergence by withholding watering peanut plants for 6 days as compared to plants adequately supplied with water. Total protein were prepared from a leaflet of fully expanded leaf on the main stem. Proteins were separated in duplicated gels using two-dimensional gel electrophoresis and visualized by silver nitrate staining. Image analysis was performed using ImageMaster 2D Platinum 5.0 to determine proteins regulated by water stress. Molecular mass and isoelectric point of each regulated protein were used in database queries for protein identification. One protein was induced under water stress and the homologous protein was identified as Serine/threonine-protein phosphatase PP 1. Five proteins were down-regulated by water deficit. The homologous proteins were chaperone protein DNAJ, auxin-responsive protein IAA29, peroxidase 43, caffeoyl-CoA O-methyltransferase and SNF1-related protein kinase regulatory subunit beta-2. Down-regulated proteins may be associated with sensitivity of the peanut variety to water stress.

Methods for the Analysis of Protein Phosphorylation-Mediated Cellular Signaling Networks

NASA Astrophysics Data System (ADS)

White, Forest M.; Wolf-Yadlin, Alejandro

2016-06-01

Protein phosphorylation-mediated cellular signaling networks regulate almost all aspects of cell biology, including the responses to cellular stimulation and environmental alterations. These networks are highly complex and comprise hundreds of proteins and potentially thousands of phosphorylation sites. Multiple analytical methods have been developed over the past several decades to identify proteins and protein phosphorylation sites regulating cellular signaling, and to quantify the dynamic response of these sites to different cellular stimulation. Here we provide an overview of these methods, including the fundamental principles governing each method, their relative strengths and weaknesses, and some examples of how each method has been applied to the analysis of complex signaling networks. When applied correctly, each of these techniques can provide insight into the topology, dynamics, and regulation of protein phosphorylation signaling networks.

Radiation induced pulmonary fibrosis as a model of progressive fibrosis: Contributions of DNA damage, inflammatory response and cellular senescence genes.

PubMed

Beach, Tyler A; Johnston, Carl J; Groves, Angela M; Williams, Jacqueline P; Finkelstein, Jacob N

2017-04-01

Purpose/Aim of Study: Studies of pulmonary fibrosis (PF) have resulted in DNA damage, inflammatory response, and cellular senescence being widely hypothesized to play a role in the progression of the disease. Utilizing these aforementioned terms, genomics databases were interrogated along with the term, "pulmonary fibrosis," to identify genes common among all 4 search terms. Findings were compared to data derived from a model of radiation-induced progressive pulmonary fibrosis (RIPF) to verify that these genes are similarly expressed, supporting the use of radiation as a model for diseases involving PF, such as human idiopathic pulmonary fibrosis (IPF). In an established model of RIPF, C57BL/6J mice were exposed to 12.5 Gy thorax irradiation and sacrificed at 24 hours, 1, 4, 12, and 32 weeks following exposure, and lung tissue was compared to age-matched controls by RNA sequencing. Of 176 PF associated gene transcripts identified by database interrogation, 146 (>82%) were present in our experimental model, throughout the progression of RIPF. Analysis revealed that nearly 85% of PF gene transcripts were associated with at least 1 other search term. Furthermore, of 22 genes common to all four terms, 16 were present experimentally in RIPF. This illustrates the validity of RIPF as a model of progressive PF/IPF based on the numbers of transcripts reported in both literature and observed experimentally. Well characterized genes and proteins are implicated in this model, supporting the hypotheses that DNA damage, inflammatory response and cellular senescence are associated with the pathogenesis of PF.

BIG: a large-scale data integration tool for renal physiology

PubMed Central

Zhao, Yue; Yang, Chin-Rang; Raghuram, Viswanathan; Parulekar, Jaya

2016-01-01

Due to recent advances in high-throughput techniques, we and others have generated multiple proteomic and transcriptomic databases to describe and quantify gene expression, protein abundance, or cellular signaling on the scale of the whole genome/proteome in kidney cells. The existence of so much data from diverse sources raises the following question: “How can researchers find information efficiently for a given gene product over all of these data sets without searching each data set individually?” This is the type of problem that has motivated the “Big-Data” revolution in Data Science, which has driven progress in fields such as marketing. Here we present an online Big-Data tool called BIG (Biological Information Gatherer) that allows users to submit a single online query to obtain all relevant information from all indexed databases. BIG is accessible at http://big.nhlbi.nih.gov/. PMID:27279488

Autophagy Regulatory Network - a systems-level bioinformatics resource for studying the mechanism and regulation of autophagy.

PubMed

Türei, Dénes; Földvári-Nagy, László; Fazekas, Dávid; Módos, Dezső; Kubisch, János; Kadlecsik, Tamás; Demeter, Amanda; Lenti, Katalin; Csermely, Péter; Vellai, Tibor; Korcsmáros, Tamás

2015-01-01

Autophagy is a complex cellular process having multiple roles, depending on tissue, physiological, or pathological conditions. Major post-translational regulators of autophagy are well known, however, they have not yet been collected comprehensively. The precise and context-dependent regulation of autophagy necessitates additional regulators, including transcriptional and post-transcriptional components that are listed in various datasets. Prompted by the lack of systems-level autophagy-related information, we manually collected the literature and integrated external resources to gain a high coverage autophagy database. We developed an online resource, Autophagy Regulatory Network (ARN; http://autophagy-regulation.org), to provide an integrated and systems-level database for autophagy research. ARN contains manually curated, imported, and predicted interactions of autophagy components (1,485 proteins with 4,013 interactions) in humans. We listed 413 transcription factors and 386 miRNAs that could regulate autophagy components or their protein regulators. We also connected the above-mentioned autophagy components and regulators with signaling pathways from the SignaLink 2 resource. The user-friendly website of ARN allows researchers without computational background to search, browse, and download the database. The database can be downloaded in SQL, CSV, BioPAX, SBML, PSI-MI, and in a Cytoscape CYS file formats. ARN has the potential to facilitate the experimental validation of novel autophagy components and regulators. In addition, ARN helps the investigation of transcription factors, miRNAs and signaling pathways implicated in the control of the autophagic pathway. The list of such known and predicted regulators could be important in pharmacological attempts against cancer and neurodegenerative diseases.

Responses of the Emiliania huxleyi proteome to ocean acidification.

PubMed

Jones, Bethan M; Iglesias-Rodriguez, M Debora; Skipp, Paul J; Edwards, Richard J; Greaves, Mervyn J; Young, Jeremy R; Elderfield, Henry; O'Connor, C David

2013-01-01

Ocean acidification due to rising atmospheric CO2 is expected to affect the physiology of important calcifying marine organisms, but the nature and magnitude of change is yet to be established. In coccolithophores, different species and strains display varying calcification responses to ocean acidification, but the underlying biochemical properties remain unknown. We employed an approach combining tandem mass-spectrometry with isobaric tagging (iTRAQ) and multiple database searching to identify proteins that were differentially expressed in cells of the marine coccolithophore species Emiliania huxleyi (strain NZEH) between two CO2 conditions: 395 (∼current day) and ∼1340 p.p.m.v. CO2. Cells exposed to the higher CO2 condition contained more cellular particulate inorganic carbon (CaCO3) and particulate organic nitrogen and carbon than those maintained in present-day conditions. These results are linked with the observation that cells grew slower under elevated CO2, indicating cell cycle disruption. Under high CO2 conditions, coccospheres were larger and cells possessed bigger coccoliths that did not show any signs of malformation compared to those from cells grown under present-day CO2 levels. No differences in calcification rate, particulate organic carbon production or cellular organic carbon: nitrogen ratios were observed. Results were not related to nutrient limitation or acclimation status of cells. At least 46 homologous protein groups from a variety of functional processes were quantified in these experiments, of which four (histones H2A, H3, H4 and a chloroplastic 30S ribosomal protein S7) showed down-regulation in all replicates exposed to high CO2, perhaps reflecting the decrease in growth rate. We present evidence of cellular stress responses but proteins associated with many key metabolic processes remained unaltered. Our results therefore suggest that this E. huxleyi strain possesses some acclimation mechanisms to tolerate future CO2 scenarios, although the observed decline in growth rate may be an overriding factor affecting the success of this ecotype in future oceans.

Responses of the Emiliania huxleyi Proteome to Ocean Acidification

PubMed Central

Jones, Bethan M.; Iglesias-Rodriguez, M. Debora; Skipp, Paul J.; Edwards, Richard J.; Greaves, Mervyn J.; Young, Jeremy R.; Elderfield, Henry; O'Connor, C. David

2013-01-01

Ocean acidification due to rising atmospheric CO2 is expected to affect the physiology of important calcifying marine organisms, but the nature and magnitude of change is yet to be established. In coccolithophores, different species and strains display varying calcification responses to ocean acidification, but the underlying biochemical properties remain unknown. We employed an approach combining tandem mass-spectrometry with isobaric tagging (iTRAQ) and multiple database searching to identify proteins that were differentially expressed in cells of the marine coccolithophore species Emiliania huxleyi (strain NZEH) between two CO2 conditions: 395 (∼current day) and ∼1340 p.p.m.v. CO2. Cells exposed to the higher CO2 condition contained more cellular particulate inorganic carbon (CaCO3) and particulate organic nitrogen and carbon than those maintained in present-day conditions. These results are linked with the observation that cells grew slower under elevated CO2, indicating cell cycle disruption. Under high CO2 conditions, coccospheres were larger and cells possessed bigger coccoliths that did not show any signs of malformation compared to those from cells grown under present-day CO2 levels. No differences in calcification rate, particulate organic carbon production or cellular organic carbon: nitrogen ratios were observed. Results were not related to nutrient limitation or acclimation status of cells. At least 46 homologous protein groups from a variety of functional processes were quantified in these experiments, of which four (histones H2A, H3, H4 and a chloroplastic 30S ribosomal protein S7) showed down-regulation in all replicates exposed to high CO2, perhaps reflecting the decrease in growth rate. We present evidence of cellular stress responses but proteins associated with many key metabolic processes remained unaltered. Our results therefore suggest that this E. huxleyi strain possesses some acclimation mechanisms to tolerate future CO2 scenarios, although the observed decline in growth rate may be an overriding factor affecting the success of this ecotype in future oceans. PMID:23593500

PLMD: An updated data resource of protein lysine modifications.

PubMed

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

2017-05-20

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

Yarrowia lipolytica vesicle-mediated protein transport pathways

PubMed Central

Swennen, Dominique; Beckerich, Jean-Marie

2007-01-01

Background Protein secretion is a universal cellular process involving vesicles which bud and fuse between organelles to bring proteins to their final destination. Vesicle budding is mediated by protein coats; vesicle targeting and fusion depend on Rab GTPase, tethering factors and SNARE complexes. The Génolevures II sequencing project made available entire genome sequences of four hemiascomycetous yeasts, Yarrowia lipolytica, Debaryomyces hansenii, Kluyveromyces lactis and Candida glabrata. Y. lipolytica is a dimorphic yeast and has good capacities to secrete proteins. The translocation of nascent protein through the endoplasmic reticulum membrane was well studied in Y. lipolytica and is largely co-translational as in the mammalian protein secretion pathway. Results We identified S. cerevisiae proteins involved in vesicular secretion and these protein sequences were used for the BLAST searches against Génolevures protein database (Y. lipolytica, C. glabrata, K. lactis and D. hansenii). These proteins are well conserved between these yeasts and Saccharomyces cerevisiae. We note several specificities of Y. lipolytica which may be related to its good protein secretion capacities and to its dimorphic aspect. An expansion of the Y. lipolytica Rab protein family was observed with autoBLAST and the Rab2- and Rab4-related members were identified with BLAST against NCBI protein database. An expansion of this family is also found in filamentous fungi and may reflect the greater complexity of the Y. lipolytica secretion pathway. The Rab4p-related protein may play a role in membrane recycling as rab4 deleted strain shows a modification of colony morphology, dimorphic transition and permeability. Similarly, we find three copies of the gene (SSO) encoding the plasma membrane SNARE protein. Quantification of the percentages of proteins with the greatest homology between S. cerevisiae, Y. lipolytica and animal homologues involved in vesicular transport shows that 40% of Y. lipolytica proteins are closer to animal ones, whereas they are only 13% in the case of S. cerevisiae. Conclusion These results provide further support for the idea, previously noted about the endoplasmic reticulum translocation pathway, that Y. lipolytica is more representative of vesicular secretion of animals and other fungi than is S. cerevisiae. PMID:17997821

Human Mitochondrial Protein Database

National Institute of Standards and Technology Data Gateway

SRD 131 Human Mitochondrial Protein Database (Web, free access)   The Human Mitochondrial Protein Database (HMPDb) provides comprehensive data on mitochondrial and human nuclear encoded proteins involved in mitochondrial biogenesis and function. This database consolidates information from SwissProt, LocusLink, Protein Data Bank (PDB), GenBank, Genome Database (GDB), Online Mendelian Inheritance in Man (OMIM), Human Mitochondrial Genome Database (mtDB), MITOMAP, Neuromuscular Disease Center and Human 2-D PAGE Databases. This database is intended as a tool not only to aid in studying the mitochondrion but in studying the associated diseases.

The Protein Information Resource: an integrated public resource of functional annotation of proteins

PubMed Central

Wu, Cathy H.; Huang, Hongzhan; Arminski, Leslie; Castro-Alvear, Jorge; Chen, Yongxing; Hu, Zhang-Zhi; Ledley, Robert S.; Lewis, Kali C.; Mewes, Hans-Werner; Orcutt, Bruce C.; Suzek, Baris E.; Tsugita, Akira; Vinayaka, C. R.; Yeh, Lai-Su L.; Zhang, Jian; Barker, Winona C.

2002-01-01

The Protein Information Resource (PIR) serves as an integrated public resource of functional annotation of protein data to support genomic/proteomic research and scientific discovery. The PIR, in collaboration with the Munich Information Center for Protein Sequences (MIPS) and the Japan International Protein Information Database (JIPID), produces the PIR-International Protein Sequence Database (PSD), the major annotated protein sequence database in the public domain, containing about 250 000 proteins. To improve protein annotation and the coverage of experimentally validated data, a bibliography submission system is developed for scientists to submit, categorize and retrieve literature information. Comprehensive protein information is available from iProClass, which includes family classification at the superfamily, domain and motif levels, structural and functional features of proteins, as well as cross-references to over 40 biological databases. To provide timely and comprehensive protein data with source attribution, we have introduced a non-redundant reference protein database, PIR-NREF. The database consists of about 800 000 proteins collected from PIR-PSD, SWISS-PROT, TrEMBL, GenPept, RefSeq and PDB, with composite protein names and literature data. To promote database interoperability, we provide XML data distribution and open database schema, and adopt common ontologies. The PIR web site (http://pir.georgetown.edu/) features data mining and sequence analysis tools for information retrieval and functional identification of proteins based on both sequence and annotation information. The PIR databases and other files are also available by FTP (ftp://nbrfa.georgetown.edu/pir_databases). PMID:11752247

A Brief Review of RNA–Protein Interaction Database Resources

PubMed Central

Yi, Ying; Zhao, Yue; Huang, Yan; Wang, Dong

2017-01-01

RNA–Protein interactions play critical roles in various biological processes. By collecting and analyzing the RNA–Protein interactions and binding sites from experiments and predictions, RNA–Protein interaction databases have become an essential resource for the exploration of the transcriptional and post-transcriptional regulatory network. Here, we briefly review several widely used RNA–Protein interaction database resources developed in recent years to provide a guide of these databases. The content and major functions in databases are presented. The brief description of database helps users to quickly choose the database containing information they interested. In short, these RNA–Protein interaction database resources are continually updated, but the current state shows the efforts to identify and analyze the large amount of RNA–Protein interactions. PMID:29657278

ORF phage display to identify cellular proteins with different functions.

PubMed

Li, Wei

2012-09-01

Open reading frame (ORF) phage display is a new branch of phage display aimed at improving its efficiency to identify cellular proteins with specific binding or functional activities. Despite the success of phage display with antibody libraries and random peptide libraries, phage display with cDNA libraries of cellular proteins identifies a high percentage of non-ORF clones encoding unnatural short peptides with minimal biological implications. This is mainly because of the uncontrollable reading frames of cellular proteins in conventional cDNA libraries. ORF phage display solves this problem by eliminating non-ORF clones to generate ORF cDNA libraries. Here I summarize the procedures of ORF phage display, discuss the factors influencing its efficiency, present examples of its versatile applications, and highlight evidence of its capability of identifying biologically relevant cellular proteins. ORF phage display coupled with different selection strategies is capable of delineating diverse functions of cellular proteins with unique advantages. Copyright © 2012 Elsevier Inc. All rights reserved.

Visualizing Viral Protein Structures in Cells Using Genetic Probes for Correlated Light and Electron Microscopy

PubMed Central

Ou, Horng D.; Deerinck, Thomas J.; Bushong, Eric; Ellisman, Mark H.; O’Shea, Clodagh C.

2015-01-01

Structural studies of viral proteins most often use high-resolution techniques such as X-ray crystallography, nuclear magnetic resonance, single particle negative stain, or cryo-electron microscopy (EM) to reveal atomic interactions of soluble, homogeneous viral proteins or viral protein complexes. Once viral proteins or complexes are separated from their host’s cellular environment, their natural in-situ structure and details of how they interact with other cellular components may be lost. EM has been an invaluable tool in virology since its introduction in the late 1940’s and subsequent application to cells in the 1950’s. EM studies have expanded our knowledge of viral entry, viral replication, alteration of cellular components, and viral lysis. Most of these early studies were focused on conspicuous morphological cellular changes, because classic EM metal stains were designed to highlight classes of cellular structures rather than specific molecular structures. Much later, to identify viral proteins inducing specific structural configurations at the cellular level, immunostaining with a primary antibody followed by colloidal gold secondary antibody was employed to mark the location of specific viral proteins. This technique can suffer from artifacts in cellular ultrastructure due to compromises required to provide access to the immuno-reagents. Immunolocalization methods also require the generation of highly specific antibodies, which may not be available for every viral protein. Here we discuss new methods to visualize viral proteins and structures at high resolutions in-situ using correlated light and electron microscopy (CLEM). We discuss the use of genetically encoded protein fusions that oxidize diaminobenzidine (DAB) into an osmiophilic polymer that can be visualized by EM. Detailed protocols for applying the genetically encoded photo-oxidizing protein MiniSOG to a viral protein, photo-oxidation of the fusion protein to yield DAB polymer staining, and preparation of photo-oxidized samples for TEM and serial block-face scanning EM (SBEM) for large-scale volume EM data acquisition are also presented. As an example, we discuss the recent multi-scale analysis of Adenoviral protein E4-ORF3 that reveals a new type of multi-functional polymer that disrupts multiple cellular proteins. This new capability to visualize unambiguously specific viral protein structures at high resolutions in the native cellular environment is revealing new insights into how they usurp host proteins and functions to drive pathological viral replication. PMID:26066760

Visualizing viral protein structures in cells using genetic probes for correlated light and electron microscopy.

PubMed

Ou, Horng D; Deerinck, Thomas J; Bushong, Eric; Ellisman, Mark H; O'Shea, Clodagh C

2015-11-15

Structural studies of viral proteins most often use high-resolution techniques such as X-ray crystallography, nuclear magnetic resonance, single particle negative stain, or cryo-electron microscopy (EM) to reveal atomic interactions of soluble, homogeneous viral proteins or viral protein complexes. Once viral proteins or complexes are separated from their host's cellular environment, their natural in situ structure and details of how they interact with other cellular components may be lost. EM has been an invaluable tool in virology since its introduction in the late 1940's and subsequent application to cells in the 1950's. EM studies have expanded our knowledge of viral entry, viral replication, alteration of cellular components, and viral lysis. Most of these early studies were focused on conspicuous morphological cellular changes, because classic EM metal stains were designed to highlight classes of cellular structures rather than specific molecular structures. Much later, to identify viral proteins inducing specific structural configurations at the cellular level, immunostaining with a primary antibody followed by colloidal gold secondary antibody was employed to mark the location of specific viral proteins. This technique can suffer from artifacts in cellular ultrastructure due to compromises required to provide access to the immuno-reagents. Immunolocalization methods also require the generation of highly specific antibodies, which may not be available for every viral protein. Here we discuss new methods to visualize viral proteins and structures at high resolutions in situ using correlated light and electron microscopy (CLEM). We discuss the use of genetically encoded protein fusions that oxidize diaminobenzidine (DAB) into an osmiophilic polymer that can be visualized by EM. Detailed protocols for applying the genetically encoded photo-oxidizing protein MiniSOG to a viral protein, photo-oxidation of the fusion protein to yield DAB polymer staining, and preparation of photo-oxidized samples for TEM and serial block-face scanning EM (SBEM) for large-scale volume EM data acquisition are also presented. As an example, we discuss the recent multi-scale analysis of Adenoviral protein E4-ORF3 that reveals a new type of multi-functional polymer that disrupts multiple cellular proteins. This new capability to visualize unambiguously specific viral protein structures at high resolutions in the native cellular environment is revealing new insights into how they usurp host proteins and functions to drive pathological viral replication. Copyright © 2015 Elsevier Inc. All rights reserved.

A core viral protein binds host nucleosomes to sequester immune danger signals

PubMed Central

Avgousti, Daphne C.; Herrmann, Christin; Kulej, Katarzyna; Pancholi, Neha J.; Sekulic, Nikolina; Petrescu, Joana; Molden, Rosalynn C.; Blumenthal, Daniel; Paris, Andrew J.; Reyes, Emigdio D.; Ostapchuk, Philomena; Hearing, Patrick; Seeholzer, Steven H.; Worthen, G. Scott; Black, Ben E.; Garcia, Benjamin A.; Weitzman, Matthew D.

2016-01-01

Viral proteins mimic host protein structure and function to redirect cellular processes and subvert innate defenses1. Small basic proteins compact and regulate both viral and cellular DNA genomes. Nucleosomes are the repeating units of cellular chromatin and play an important role in innate immune responses2. Viral encoded core basic proteins compact viral genomes but their impact on host chromatin structure and function remains unexplored. Adenoviruses encode a highly basic protein called protein VII that resembles cellular histones3. Although protein VII binds viral DNA and is incorporated with viral genomes into virus particles4,5, it is unknown whether protein VII impacts cellular chromatin. Our observation that protein VII alters cellular chromatin led us to hypothesize that this impacts antiviral responses during adenovirus infection. We found that protein VII forms complexes with nucleosomes and limits DNA accessibility. We identified post-translational modifications on protein VII that are responsible for chromatin localization. Furthermore, proteomic analysis demonstrated that protein VII is sufficient to alter protein composition of host chromatin. We found that protein VII is necessary and sufficient for retention in chromatin of members of the high-mobility group protein B family (HMGB1, HMGB2, and HMGB3). HMGB1 is actively released in response to inflammatory stimuli and functions as a danger signal to activate immune responses6,7. We showed that protein VII can directly bind HMGB1 in vitro and further demonstrated that protein VII expression in mouse lungs is sufficient to decrease inflammation-induced HMGB1 content and neutrophil recruitment in the bronchoalveolar lavage fluid. Together our in vitro and in vivo results show that protein VII sequesters HMGB1 and can prevent its release. This study uncovers a viral strategy in which nucleosome binding is exploited to control extracellular immune signaling. PMID:27362237

Characterization of flagellar cysteine-rich sperm proteins involved in motility, by the combination of cellular fractionation, fluorescence detection, and mass spectrometry analysis.

PubMed

Cabrillana, María E; Monclus, María A; Sáez Lancellotti, Tania E; Boarelli, Paola V; Clementi, Marisa A; Vincenti, Amanda E; Yunes, Roberto F M; Fornés, Miguel W

2011-09-01

Mammalian sperm proteins undergo thiol group (SH) oxidation to form disulfides bonds (SS) as they travel through the epididymis during cell maturation. Disulfide bonds are involved in chromatin condensation and tail organelle stabilization. In this work, we used a fluorescent thiol-selective labeling agent, monobromobimane (mBBr), to study the protein thiol status of rat sperm during maturation. Fluorescence signal decrease along the epididymal trip, more evidently in the head, but also in the tail, indicates that both sub cellular regions participate in the thiol changes. The sources of the fluorescence signal are sulfhydryls sperm proteins labeled by mBBr (mBBr-spp). Initial attempts to identify the mBBr-spp labeled were detected in the initial-caput, but not in the distal cauda-segment of the epididymis in sodium dodecyl sulfate (SDS)-PAGE analysis. This phenomenon could be due to protein resistance to solubilization. For this reason, disulfide bond reduction was accomplished by sodium dodecyl sulfate plus dithiothreitol treatment to recover the mBBr signal in SDS-PAGE. Under this protocol, a major 27 kDa protein band displays a strong signal. Protein identification by mass spectrometry and sequence database searching correlated this protein with the outer dense fiber 1 (ODF1). The mBBr specifically bound to N-terminal domain cysteine of ODF1. The mBBr reduces rat sperm motility, quantitatively and qualitatively, and the effects are dose dependent, without significantly increasing the percentage of dead sperm. Thus, we found that ODF1 is highly responsible for mBBr fluorescence detection in the sperm tail, and the motility inhibition by the fluorescence marker indicates that ODF1 N-terminal domain are related to sperm motility. © 2011 Wiley-Liss, Inc. Copyright © 2011 Wiley-Liss, Inc.

Prevalence of mutations in hepatitis C virus core protein associated with alteration of NF-kappaB activation.

PubMed

Mann, Elizabeth A; Stanford, Sandra; Sherman, Kenneth E

2006-10-01

The hepatitis C virus (HCV) core protein is a key structural element of the virion but also affects a number of cellular pathways, including nuclear factor kappaB (NF-kappaB) signaling. NF-kappaB is a transcription factor that regulates both anti-apoptotic and pro-inflammatory genes and its activation may contribute to HCV-mediated pathogenesis. Amino acid sequence divergence in core is seen at the genotype level as well as within patient isolates. Recent work has implicated amino acids 9-11 of core in the modulation of NF-kappaB activation. We report that the sequence RKT is highly conserved (93%) at this position across all HCV genotypes, based on sequences collected in the Los Alamos HCV database. Of the 13 types of variants present in the database, the two most prevalent substitutions are RQT and RKP. We further show that core encoding RKP fails to activate NF-kappaB signaling in vitro while NF-kappaB activation by core encoding RQT does not differ from control RKT core. The effect of RKP core is specific to NF-kappaB signaling as activator protein 1 (AP-1) activity is not altered. Further studies are needed to assess potential associations between specific amino acid substitutions at positions 9-11 and liver disease progression and/or response to treatment in individual patients.

Selective inhibition of influenza virus protein synthesis by inhibitors of DNA function. [UV radiation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Minor, P.D.; Dimmock, N.J.

1977-05-15

Various known inhibitors of cellular DNA function were shown to inhibit cellular RNA synthesis and influenza (fowl plague) virus multiplication. The drugs were investigated for their effect upon the synthesis of influenza virus proteins. According to this effect they could be classified with previously studied compounds as follows: Group I (ethidium bromide, proflavine, and N-nitroquinoline-N-oxide) inhibited both viral and cellular protein synthesis; Group II (nogalomycin, daunomycin and ..cap alpha..-amanitin) inhibited viral but not cellular protein synthesis, and all viral proteins were inhibited coordinately; Group III (mithramycin, echinomycin, and actinomycin D) inhibited all viral but not cellular protein synthesis at highmore » concentrations, but at a lower critical concentration inhibited the synthesis of viral haemagglutinin, neuraminidase, and M protein preferentially; Group IV(uv irradiation and camptothecin) inhibited the synthesis of viral haemagglutinin, neuraminidase, and M protein, but not other viral proteins, even at high doses. The mode of action of these inhibitors is discussed in relation to the mechanism of the nuclear events upon which influenza virus multiplication is dependent.« less

Physiological enzymology: The next frontier in understanding protein structure and function at the cellular level.

PubMed

Lee, Irene; Berdis, Anthony J

2016-01-01

Historically, the study of proteins has relied heavily on characterizing the activity of a single purified protein isolated from other cellular components. This classic approach allowed scientists to unambiguously define the intrinsic kinetic and chemical properties of that protein. The ultimate hope was to extrapolate this information toward understanding how the enzyme or receptor behaves within its native cellular context. These types of detailed in vitro analyses were necessary to reduce the innate complexities of measuring the singular activity and biochemical properties of a specific enzyme without interference from other enzymes and potential competing substrates. However, recent developments in fields encompassing cell biology, molecular imaging, and chemical biology now provide the unique chemical tools and instrumentation to study protein structure, function, and regulation in their native cellular environment. These advancements provide the foundation for a new field, coined physiological enzymology, which quantifies the function and regulation of enzymes and proteins at the cellular level. In this Special Edition, we explore the area of Physiological Enzymology and Protein Function through a series of review articles that focus on the tools and techniques used to measure the cellular activity of proteins inside living cells. This article is part of a Special Issue entitled: Physiological Enzymology and Protein Functions. Copyright © 2015 Elsevier B.V. All rights reserved.

Cellular immune response in MDR-TB patients to different protein expression of MDR and susceptible Mycobacterium tuberculosis: Rv0147, a novel MDR-TB biomarker.

PubMed

Hadizadeh Tasbiti, Alireza; Yari, Shamsi; Siadat, Seyed Davar; Tabarsi, Payam; Saeedfar, Kayvan; Yari, Fatemeh

2018-02-01

Tuberculosis (TB) is a crucial public health problem with prevalence of multidrug resistant (MDR) rising. An accurate TB biomarker is urgently needed to monitor the response to treatment in patients with MDR tuberculosis. To analyze interaction between selected MDR-TB purified protein and immune cells, dendritic cells from MDR-TB patients and healthy subjects were stimulated by 55KDa protein fractions (Rv0147). The purified proteins identified by proteomic techniques (two-dimensional gel electrophoresis, mass spectrometry) and peptide sequences are known to bind a MHC class I alleles which are extracted from the Immune Epitope Database and Analysis Resource database ( www.iedb.org ). T cells were isolated from PBMC by negative selection and cells were cultured in RPMI-1640 at 37 °C and 5% CO 2 . Cell culture was assayed for cytokine IL-10 and INF-γ by ELISA. We found that INF-γ production was significantly (335 ± 35.5 pg/ml, P ˂ 0.05) upregulated after protein candidate (Rv0147) stimulation by dendritic cells from MDR-TB patients, whereas IL-10 production was greatly reduced compared with production in healthy subjects (212 ± 9.94 pg/ml, P ˂ 0.05). In fact, the purified protein, Rv0147, stimulated dendritic cells from MDR-TB patients, failed to produce IL-10 and directly stimulates INF-γ production by T cells. These results suggest that the purified protein, Rv0147, may stimulate Th1 type protective cytokine response in MDR-TB patients but not in normal subjects. The production of INF-γ but not IL-10 in the presence of purified protein, Rv0147, may be shifted to Th1 responses in MDR-TB patients and supports its potential as protein vaccine candidates against TB.

Protein Information Resource: a community resource for expert annotation of protein data

PubMed Central

Barker, Winona C.; Garavelli, John S.; Hou, Zhenglin; Huang, Hongzhan; Ledley, Robert S.; McGarvey, Peter B.; Mewes, Hans-Werner; Orcutt, Bruce C.; Pfeiffer, Friedhelm; Tsugita, Akira; Vinayaka, C. R.; Xiao, Chunlin; Yeh, Lai-Su L.; Wu, Cathy

2001-01-01

The Protein Information Resource, in collaboration with the Munich Information Center for Protein Sequences (MIPS) and the Japan International Protein Information Database (JIPID), produces the most comprehensive and expertly annotated protein sequence database in the public domain, the PIR-International Protein Sequence Database. To provide timely and high quality annotation and promote database interoperability, the PIR-International employs rule-based and classification-driven procedures based on controlled vocabulary and standard nomenclature and includes status tags to distinguish experimentally determined from predicted protein features. The database contains about 200 000 non-redundant protein sequences, which are classified into families and superfamilies and their domains and motifs identified. Entries are extensively cross-referenced to other sequence, classification, genome, structure and activity databases. The PIR web site features search engines that use sequence similarity and database annotation to facilitate the analysis and functional identification of proteins. The PIR-Inter­national databases and search tools are accessible on the PIR web site at http://pir.georgetown.edu/ and at the MIPS web site at http://www.mips.biochem.mpg.de. The PIR-International Protein Sequence Database and other files are also available by FTP. PMID:11125041

Dynamic interactions between 14-3-3 proteins and phosphoproteins regulate diverse cellular processes

PubMed Central

2004-01-01

14-3-3 proteins exert an extraordinarily widespread influence on cellular processes in all eukaryotes. They operate by binding to specific phosphorylated sites on diverse target proteins, thereby forcing conformational changes or influencing interactions between their targets and other molecules. In these ways, 14-3-3s ‘finish the job’ when phosphorylation alone lacks the power to drive changes in the activities of intracellular proteins. By interacting dynamically with phosphorylated proteins, 14-3-3s often trigger events that promote cell survival – in situations from preventing metabolic imbalances caused by sudden darkness in leaves to mammalian cell-survival responses to growth factors. Recent work linking specific 14-3-3 isoforms to genetic disorders and cancers, and the cellular effects of 14-3-3 agonists and antagonists, indicate that the cellular complement of 14-3-3 proteins may integrate the specificity and strength of signalling through to different cellular responses. PMID:15167810

Human HOXA5 homeodomain enhances protein transduction and its application to vascular inflammation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lee, Ji Young; Park, Kyoung sook; Cho, Eun Jung

2011-07-01

Highlights: {yields} We have developed an E. coli protein expression vector including human specific gene sequences for protein cellular delivery. {yields} The plasmid was generated by ligation the nucleotides 770-817 of the homeobox A5 mRNA sequence. {yields} HOXA5-APE1/Ref-1 inhibited TNF-alpha-induced monocyte adhesion to endothelial cells. {yields} Human HOXA5-PTD vector provides a powerful research tools for uncovering cellular functions of proteins or for the generation of human PTD-containing proteins. -- Abstract: Cellular protein delivery is an emerging technique by which exogenous recombinant proteins are delivered into mammalian cells across the membrane. We have developed an Escherichia coli expression vector including humanmore » specific gene sequences for protein cellular delivery. The plasmid was generated by ligation the nucleotides 770-817 of the homeobox A5 mRNA sequence which was matched with protein transduction domain (PTD) of homeodomain protein A5 (HOXA5) into pET expression vector. The cellular uptake of HOXA5-PTD-EGFP was detected in 1 min and its transduction reached a maximum at 1 h within cell lysates. The cellular uptake of HOXA5-EGFP at 37 {sup o}C was greater than in 4 {sup o}C. For study for the functional role of human HOXA5-PTD, we purified HOXA5-APE1/Ref-1 and applied it on monocyte adhesion. Pretreatment with HOXA5-APE1/Ref-1 (100 nM) inhibited TNF-{alpha}-induced monocyte adhesion to endothelial cells, compared with HOXA5-EGFP. Taken together, our data suggested that human HOXA5-PTD vector provides a powerful research tools for uncovering cellular functions of proteins or for the generation of human PTD-containing proteins.« less

Proteomic Analysis and Functional Studies of Baicalin on Proteins Associated with Skin Cancer.

PubMed

Li, Dan; Lin, Bingjiang; Yusuf, Nabiha; Burns, Erin M; Yu, Xiuqin; Luo, Dan; Min, Wei

2017-01-01

Abundant evidence supports the key role of ultraviolet radiation (UVR) in skin cancer development. The human skin, especially the epidermal layer, is the main defense against UV radiation. Baicalin is a major bioactive component of Scutellaria baicalensis Georgi, a plant which has been found to exhibit antitumor activity. The anticarcinogenic mechanism of baicalin is not completely understood. We have reported that baicalin inhibited UVB-induced photo-damage and apoptosis in HaCaT cells (human skin keratinocytes). The aim of the present study is to investigate the cellular gene targets responsible for baicalin's antitumor activity by performing two-dimensional electrophoresis liquid chromatography-mass spectrometry/mass spectrometry (2-DE LC-MS/MS) with HaCaT cells following UVB and baicalin exposure. Two-DE for protein separation was performed, followed by matrix-assisted laser desorption/ionization mass spectrometry and database searches. Nucleophosmin (NPM)-specific siRNA was designed and synthesized, and the small interfering RNA was transfected into skin squamous cancer A431 cells to knockdown the NPM expression. Proliferation and cell cycle status were assessed by CCK8 and flow cytometric analyses, respectively. We have identified 38 protein spots that are differentially expressed in HaCaT cells exposed to baicalin and/or UVB irradiation These proteins are involved in detoxification, proliferation, metabolism, cytoskeleton and motility. In particular, we found several proteins that have been linked to tumor progression and resistance, such as NPM. Baicalin treatment reduced the cellular proliferation rate and induced arrest during the S-phase of the cell cycle in A431 cells. NPM1 silencing significantly enhanced the effect of baicalin. Our data indicated that baicalin results in the significant inhibition of tumor growth in the A431 cell line, which may be associated with the regulation of the NPM gene expression.

Searching for cellular partners of hantaviral nonstructural protein NSs: Y2H screening of mouse cDNA library and analysis of cellular interactome.

PubMed

Rönnberg, Tuomas; Jääskeläinen, Kirsi; Blot, Guillaume; Parviainen, Ville; Vaheri, Antti; Renkonen, Risto; Bouloy, Michele; Plyusnin, Alexander

2012-01-01

Hantaviruses (Bunyaviridae) are negative-strand RNA viruses with a tripartite genome. The small (S) segment encodes the nucleocapsid protein and, in some hantaviruses, also the nonstructural protein (NSs). The aim of this study was to find potential cellular partners for the hantaviral NSs protein. Toward this aim, yeast two-hybrid (Y2H) screening of mouse cDNA library was performed followed by a search for potential NSs protein counterparts via analyzing a cellular interactome. The resulting interaction network was shown to form logical, clustered structures. Furthermore, several potential binding partners for the NSs protein, for instance ACBD3, were identified and, to prove the principle, interaction between NSs and ACBD3 proteins was demonstrated biochemically.

Cellular Binding of Anionic Nanoparticles is Inhibited by Serum Proteins Independent of Nanoparticle Composition.

PubMed

Fleischer, Candace C; Kumar, Umesh; Payne, Christine K

2013-09-01

Nanoparticles used in biological applications encounter a complex mixture of extracellular proteins. Adsorption of these proteins on the nanoparticle surface results in the formation of a "protein corona," which can dominate the interaction of the nanoparticle with the cellular environment. The goal of this research was to determine how nanoparticle composition and surface modification affect the cellular binding of protein-nanoparticle complexes. We examined the cellular binding of a collection of commonly used anionic nanoparticles: quantum dots, colloidal gold nanoparticles, and low-density lipoprotein particles, in the presence and absence of extracellular proteins. These experiments have the advantage of comparing different nanoparticles under identical conditions. Using a combination of fluorescence and dark field microscopy, flow cytometry, and spectroscopy, we find that cellular binding of these anionic nanoparticles is inhibited by serum proteins independent of nanoparticle composition or surface modification. We expect these results will aid in the design of nanoparticles for in vivo applications.

Intracellular cargo delivery by virus capsid protein-based vehicles: From nano to micro.

PubMed

Gao, Ding; Lin, Xiu-Ping; Zhang, Zhi-Ping; Li, Wei; Men, Dong; Zhang, Xian-En; Cui, Zong-Qiang

2016-02-01

Cellular delivery is an important concern for the efficiency of medicines and sensors for disease diagnoses and therapy. However, this task is quite challenging. Self-assembly virus capsid proteins might be developed as building blocks for multifunctional cellular delivery vehicles. In this work, we found that SV40 VP1 (Simian virus 40 major capsid protein) could function as a new cell-penetrating protein. The VP1 protein could carry foreign proteins into cells in a pentameric structure. A double color structure, with red QDs (Quantum dots) encapsulated by viral capsids fused with EGFP, was created for imaging cargo delivery and release from viral capsids. The viral capsids encapsulating QDs were further used for cellular delivery of micron-sized iron oxide particles (MPIOs). MPIOs were efficiently delivered into live cells and controlled by a magnetic field. Therefore, our study built virus-based cellular delivery systems for different sizes of cargos: protein molecules, nanoparticles, and micron-sized particles. Much research is being done to investigate methods for efficient and specific cellular delivery of drugs, proteins or genetic material. In this article, the authors describe their approach in using self-assembly virus capsid proteins SV40 VP1 (Simian virus 40 major capsid protein). The cell-penetrating behavior provided excellent cellular delivery and should give a new method for biomedical applications. Copyright © 2015 Elsevier Inc. All rights reserved.

Signaling gateway molecule pages—a data model perspective

PubMed Central

Dinasarapu, Ashok Reddy; Saunders, Brian; Ozerlat, Iley; Azam, Kenan; Subramaniam, Shankar

2011-01-01

Summary: The Signaling Gateway Molecule Pages (SGMP) database provides highly structured data on proteins which exist in different functional states participating in signal transduction pathways. A molecule page starts with a state of a native protein, without any modification and/or interactions. New states are formed with every post-translational modification or interaction with one or more proteins, small molecules or class molecules and with each change in cellular location. State transitions are caused by a combination of one or more modifications, interactions and translocations which then might be associated with one or more biological processes. In a characterized biological state, a molecule can function as one of several entities or their combinations, including channel, receptor, enzyme, transcription factor and transporter. We have also exported SGMP data to the Biological Pathway Exchange (BioPAX) and Systems Biology Markup Language (SBML) as well as in our custom XML. Availability: SGMP is available at www.signaling-gateway.org/molecule. Contact: shankar@ucsd.edu Supplementary information: Supplementary data are available at Bioinformatics online. PMID:21505029

ProBiS-database: precalculated binding site similarities and local pairwise alignments of PDB structures.

PubMed

Konc, Janez; Cesnik, Tomo; Konc, Joanna Trykowska; Penca, Matej; Janežič, Dušanka

2012-02-27

ProBiS-Database is a searchable repository of precalculated local structural alignments in proteins detected by the ProBiS algorithm in the Protein Data Bank. Identification of functionally important binding regions of the protein is facilitated by structural similarity scores mapped to the query protein structure. PDB structures that have been aligned with a query protein may be rapidly retrieved from the ProBiS-Database, which is thus able to generate hypotheses concerning the roles of uncharacterized proteins. Presented with uncharacterized protein structure, ProBiS-Database can discern relationships between such a query protein and other better known proteins in the PDB. Fast access and a user-friendly graphical interface promote easy exploration of this database of over 420 million local structural alignments. The ProBiS-Database is updated weekly and is freely available online at http://probis.cmm.ki.si/database.

A family of cellular proteins related to snake venom disintegrins.

PubMed

Weskamp, G; Blobel, C P

1994-03-29

Disintegrins are short soluble integrin ligands that were initially identified in snake venom. A previously recognized cellular protein with a disintegrin domain was the guinea pig sperm protein PH-30, a protein implicated in sperm-egg membrane binding and fusion. Here we present peptide sequences that are characteristic for several cellular disintegrin-domain proteins. These peptide sequences were deduced from cDNA sequence tags that were generated by polymerase chain reaction from various mouse tissue and a mouse muscle cell line. Northern blot analysis with four sequence tags revealed distinct mRNA expression patterns. Evidently, cellular proteins containing a disintegrin domain define a superfamily of potential integrin ligands that are likely to function in important cell-cell and cell-matrix interactions.

An ontology for major histocompatibility restriction.

PubMed

Vita, Randi; Overton, James A; Seymour, Emily; Sidney, John; Kaufman, Jim; Tallmadge, Rebecca L; Ellis, Shirley; Hammond, John; Butcher, Geoff W; Sette, Alessandro; Peters, Bjoern

2016-01-01

MHC molecules are a highly diverse family of proteins that play a key role in cellular immune recognition. Over time, different techniques and terminologies have been developed to identify the specific type(s) of MHC molecule involved in a specific immune recognition context. No consistent nomenclature exists across different vertebrate species. To correctly represent MHC related data in The Immune Epitope Database (IEDB), we built upon a previously established MHC ontology and created an ontology to represent MHC molecules as they relate to immunological experiments. This ontology models MHC protein chains from 16 species, deals with different approaches used to identify MHC, such as direct sequencing verses serotyping, relates engineered MHC molecules to naturally occurring ones, connects genetic loci, alleles, protein chains and multi-chain proteins, and establishes evidence codes for MHC restriction. Where available, this work is based on existing ontologies from the OBO foundry. Overall, representing MHC molecules provides a challenging and practically important test case for ontology building, and could serve as an example of how to integrate other ontology building efforts into web resources.

Cassandra retrotransposons carry independently transcribed 5S RNA

PubMed Central

Kalendar, Ruslan; Tanskanen, Jaakko; Chang, Wei; Antonius, Kristiina; Sela, Hanan; Peleg, Ofer; Schulman, Alan H.

2008-01-01

We report a group of TRIMs (terminal-repeat retrotransposons in miniature), which are small nonautonomous retrotransposons. These elements, named Cassandra, universally carry conserved 5S RNA sequences and associated RNA polymerase (pol) III promoters and terminators in their long terminal repeats (LTRs). They were found in all vascular plants investigated. Uniquely for LTR retrotransposons, Cassandra produces noncapped, polyadenylated transcripts from the 5S pol III promoter. Capped, read-through transcripts containing Cassandra sequences can also be detected in RNA and in EST databases. The predicted Cassandra RNA 5S secondary structures resemble those for cellular 5S rRNA, with high information content specifically in the pol III promoter region. Genic integration sites are common for Cassandra, an unusual feature for abundant retrotransposons. The 5S in each LTR produces a tandem 5S arrangement with an inter-5S spacing resembling that of cellular 5S. The distribution of 5S genes is very variable in flowering plants and may be partially explained by Cassandra activity. Cassandra thus appears both to have adapted a ubiquitous cellular gene for ribosomal RNA for use as a promoter and to parasitize an as-yet-unidentified group of retrotransposons for the proteins needed in its lifecycle. PMID:18408163

Purine inhibitors of protein kinases, G proteins and polymerases

DOEpatents

Gray, Nathanael S.; Schultz, Peter; Kim, Sung-Hou; Meijer, Laurent

2001-07-03

The present invention relates to purine analogs that inhibit, inter alia, protein kinases, G-proteins and polymerases. In addition, the present invention relates to methods of using such purine analogs to inhibit protein kinases, G-proteins, polymerases and other cellular processes and to treat cellular proliferative diseases.

Choosing an Optimal Database for Protein Identification from Tandem Mass Spectrometry Data.

PubMed

Kumar, Dhirendra; Yadav, Amit Kumar; Dash, Debasis

2017-01-01

Database searching is the preferred method for protein identification from digital spectra of mass to charge ratios (m/z) detected for protein samples through mass spectrometers. The search database is one of the major influencing factors in discovering proteins present in the sample and thus in deriving biological conclusions. In most cases the choice of search database is arbitrary. Here we describe common search databases used in proteomic studies and their impact on final list of identified proteins. We also elaborate upon factors like composition and size of the search database that can influence the protein identification process. In conclusion, we suggest that choice of the database depends on the type of inferences to be derived from proteomics data. However, making additional efforts to build a compact and concise database for a targeted question should generally be rewarding in achieving confident protein identifications.

From hatching to dispatching: the multiple cellular roles of the Hsp70 molecular chaperone machinery.

PubMed

Meimaridou, Eirini; Gooljar, Sakina B; Chapple, J Paul

2009-01-01

Molecular chaperones are best recognized for their roles in de novo protein folding and the cellular response to stress. However, many molecular chaperones, and in particular the Hsp70 chaperone machinery, have multiple diverse cellular functions. At the molecular level, chaperones are mediators of protein conformational change. To facilitate conformational change of client/substrate proteins, in manifold contexts, chaperone power must be closely regulated and harnessed to specific cellular locales--this is controlled by cochaperones. This review considers specialized functions of the Hsp70 chaperone machinery mediated by its cochaperones. We focus on vesicular trafficking, protein degradation and a potential role in G protein-coupled receptor processing.

ATLAS of Biochemistry: A Repository of All Possible Biochemical Reactions for Synthetic Biology and Metabolic Engineering Studies.

PubMed

Hadadi, Noushin; Hafner, Jasmin; Shajkofci, Adrian; Zisaki, Aikaterini; Hatzimanikatis, Vassily

2016-10-21

Because the complexity of metabolism cannot be intuitively understood or analyzed, computational methods are indispensable for studying biochemistry and deepening our understanding of cellular metabolism to promote new discoveries. We used the computational framework BNICE.ch along with cheminformatic tools to assemble the whole theoretical reactome from the known metabolome through expansion of the known biochemistry presented in the Kyoto Encyclopedia of Genes and Genomes (KEGG) database. We constructed the ATLAS of Biochemistry, a database of all theoretical biochemical reactions based on known biochemical principles and compounds. ATLAS includes more than 130 000 hypothetical enzymatic reactions that connect two or more KEGG metabolites through novel enzymatic reactions that have never been reported to occur in living organisms. Moreover, ATLAS reactions integrate 42% of KEGG metabolites that are not currently present in any KEGG reaction into one or more novel enzymatic reactions. The generated repository of information is organized in a Web-based database ( http://lcsb-databases.epfl.ch/atlas/ ) that allows the user to search for all possible routes from any substrate compound to any product. The resulting pathways involve known and novel enzymatic steps that may indicate unidentified enzymatic activities and provide potential targets for protein engineering. Our approach of introducing novel biochemistry into pathway design and associated databases will be important for synthetic biology and metabolic engineering.

Interaction Network of Proteins Associated with Human Cytomegalovirus IE2-p86 Protein during Infection: A Proteomic Analysis

PubMed Central

Du, Guixin; Stinski, Mark F.

2013-01-01

Human cytomegalovirus protein IE2-p86 exerts its functions through interaction with other viral and cellular proteins. To further delineate its protein interaction network, we generated a recombinant virus expressing SG-tagged IE2-p86 and used tandem affinity purification coupled with mass spectrometry. A total of 9 viral proteins and 75 cellular proteins were found to associate with IE2-p86 protein during the first 48 hours of infection. The protein profile at 8, 24, and 48 h post infection revealed that UL84 tightly associated with IE2-p86, and more viral and cellular proteins came into association with IE2-p86 with the progression of virus infection. A computational analysis of the protein-protein interaction network indicated that all of the 9 viral proteins and most of the cellular proteins identified in the study are interconnected to varying degrees. Of the cellular proteins that were confirmed to associate with IE2-p86 by immunoprecipitation, C1QBP was further shown to be upregulated by HCMV infection and colocalized with IE2-p86, UL84 and UL44 in the virus replication compartment of the nucleus. The IE2-p86 interactome network demonstrated the temporal development of stable and abundant protein complexes that associate with IE2-p86 and provided a framework to benefit future studies of various protein complexes during HCMV infection. PMID:24358118

Software Tool for Researching Annotations of Proteins (STRAP): Open-Source Protein Annotation Software with Data Visualization

PubMed Central

Bhatia, Vivek N.; Perlman, David H.; Costello, Catherine E.; McComb, Mark E.

2009-01-01

In order that biological meaning may be derived and testable hypotheses may be built from proteomics experiments, assignments of proteins identified by mass spectrometry or other techniques must be supplemented with additional notation, such as information on known protein functions, protein-protein interactions, or biological pathway associations. Collecting, organizing, and interpreting this data often requires the input of experts in the biological field of study, in addition to the time-consuming search for and compilation of information from online protein databases. Furthermore, visualizing this bulk of information can be challenging due to the limited availability of easy-to-use and freely available tools for this process. In response to these constraints, we have undertaken the design of software to automate annotation and visualization of proteomics data in order to accelerate the pace of research. Here we present the Software Tool for Researching Annotations of Proteins (STRAP) – a user-friendly, open-source C# application. STRAP automatically obtains gene ontology (GO) terms associated with proteins in a proteomics results ID list using the freely accessible UniProtKB and EBI GOA databases. Summarized in an easy-to-navigate tabular format, STRAP includes meta-information on the protein in addition to complimentary GO terminology. Additionally, this information can be edited by the user so that in-house expertise on particular proteins may be integrated into the larger dataset. STRAP provides a sortable tabular view for all terms, as well as graphical representations of GO-term association data in pie (biological process, cellular component and molecular function) and bar charts (cross comparison of sample sets) to aid in the interpretation of large datasets and differential analyses experiments. Furthermore, proteins of interest may be exported as a unique FASTA-formatted file to allow for customizable re-searching of mass spectrometry data, and gene names corresponding to the proteins in the lists may be encoded in the Gaggle microformat for further characterization, including pathway analysis. STRAP, a tutorial, and the C# source code are freely available from http://cpctools.sourceforge.net. PMID:19839595

Metabotropic glutamate receptor 5 couples cellular prion protein to intracellular signalling in Alzheimer’s disease

PubMed Central

Haas, Laura T.; Salazar, Santiago V.; Kostylev, Mikhail A.; Um, Ji Won; Kaufman, Adam C.

2016-01-01

Alzheimer’s disease-related phenotypes in mice can be rescued by blockade of either cellular prion protein or metabotropic glutamate receptor 5. We sought genetic and biochemical evidence that these proteins function cooperatively as an obligate complex in the brain. We show that cellular prion protein associates via transmembrane metabotropic glutamate receptor 5 with the intracellular protein mediators Homer1b/c, calcium/calmodulin-dependent protein kinase II, and the Alzheimer’s disease risk gene product protein tyrosine kinase 2 beta. Coupling of cellular prion protein to these intracellular proteins is modified by soluble amyloid-β oligomers, by mouse brain Alzheimer’s disease transgenes or by human Alzheimer’s disease pathology. Amyloid-β oligomer-triggered phosphorylation of intracellular protein mediators and impairment of synaptic plasticity in vitro requires Prnp–Grm5 genetic interaction, being absent in transheterozygous loss-of-function, but present in either single heterozygote. Importantly, genetic coupling between Prnp and Grm5 is also responsible for signalling, for survival and for synapse loss in Alzheimer’s disease transgenic model mice. Thus, the interaction between metabotropic glutamate receptor 5 and cellular prion protein has a central role in Alzheimer’s disease pathogenesis, and the complex is a potential target for disease-modifying intervention. PMID:26667279

MIPS: analysis and annotation of proteins from whole genomes.

PubMed

Mewes, H W; Amid, C; Arnold, R; Frishman, D; Güldener, U; Mannhaupt, G; Münsterkötter, M; Pagel, P; Strack, N; Stümpflen, V; Warfsmann, J; Ruepp, A

2004-01-01

The Munich Information Center for Protein Sequences (MIPS-GSF), Neuherberg, Germany, provides protein sequence-related information based on whole-genome analysis. The main focus of the work is directed toward the systematic organization of sequence-related attributes as gathered by a variety of algorithms, primary information from experimental data together with information compiled from the scientific literature. MIPS maintains automatically generated and manually annotated genome-specific databases, develops systematic classification schemes for the functional annotation of protein sequences and provides tools for the comprehensive analysis of protein sequences. This report updates the information on the yeast genome (CYGD), the Neurospora crassa genome (MNCDB), the database of complete cDNAs (German Human Genome Project, NGFN), the database of mammalian protein-protein interactions (MPPI), the database of FASTA homologies (SIMAP), and the interface for the fast retrieval of protein-associated information (QUIPOS). The Arabidopsis thaliana database, the rice database, the plant EST databases (MATDB, MOsDB, SPUTNIK), as well as the databases for the comprehensive set of genomes (PEDANT genomes) are described elsewhere in the 2003 and 2004 NAR database issues, respectively. All databases described, and the detailed descriptions of our projects can be accessed through the MIPS web server (http://mips.gsf.de).

Adenovirus Core Protein VII Downregulates the DNA Damage Response on the Host Genome

PubMed Central

Avgousti, Daphne C.; Della Fera, Ashley N.; Otter, Clayton J.; Herrmann, Christin; Pancholi, Neha J.

2017-01-01

ABSTRACT Viral manipulation of cellular proteins allows viruses to suppress host defenses and generate infectious progeny. Due to the linear double-stranded DNA nature of the adenovirus genome, the cellular DNA damage response (DDR) is considered a barrier to successful infection. The adenovirus genome is packaged with protein VII, a virally encoded histone-like core protein that is suggested to protect incoming viral genomes from detection by the cellular DNA damage machinery. We showed that protein VII localizes to host chromatin during infection, leading us to hypothesize that protein VII may affect DNA damage responses on the cellular genome. Here we show that protein VII at cellular chromatin results in a significant decrease in accumulation of phosphorylated H2AX (γH2AX) following irradiation, indicating that protein VII inhibits DDR signaling. The oncoprotein SET was recently suggested to modulate the DDR by affecting access of repair proteins to chromatin. Since protein VII binds SET, we investigated a role for SET in DDR inhibition by protein VII. We show that knockdown of SET partially rescues the protein VII-induced decrease in γH2AX accumulation on the host genome, suggesting that SET is required for inhibition. Finally, we show that knockdown of SET also allows ATM to localize to incoming viral genomes bound by protein VII during infection with a mutant lacking early region E4. Together, our data suggest that the protein VII-SET interaction contributes to DDR evasion by adenovirus. Our results provide an additional example of a strategy used by adenovirus to abrogate the host DDR and show how viruses can modify cellular processes through manipulation of host chromatin. IMPORTANCE The DNA damage response (DDR) is a cellular network that is crucial for maintaining genome integrity. DNA viruses replicating in the nucleus challenge the resident genome and must overcome cellular responses, including the DDR. Adenoviruses are prevalent human pathogens that can cause a multitude of diseases, such as respiratory infections and conjunctivitis. Here we describe how a small adenovirus core protein that localizes to host chromatin during infection can globally downregulate the DDR. Our study focuses on key players in the damage signaling pathway and highlights how viral manipulation of chromatin may influence access of DDR proteins to the host genome. PMID:28794020

Characterizing Protein Interactions Employing a Genome-Wide siRNA Cellular Phenotyping Screen

PubMed Central

Suratanee, Apichat; Schaefer, Martin H.; Betts, Matthew J.; Soons, Zita; Mannsperger, Heiko; Harder, Nathalie; Oswald, Marcus; Gipp, Markus; Ramminger, Ellen; Marcus, Guillermo; Männer, Reinhard; Rohr, Karl; Wanker, Erich; Russell, Robert B.; Andrade-Navarro, Miguel A.; Eils, Roland; König, Rainer

2014-01-01

Characterizing the activating and inhibiting effect of protein-protein interactions (PPI) is fundamental to gain insight into the complex signaling system of a human cell. A plethora of methods has been suggested to infer PPI from data on a large scale, but none of them is able to characterize the effect of this interaction. Here, we present a novel computational development that employs mitotic phenotypes of a genome-wide RNAi knockdown screen and enables identifying the activating and inhibiting effects of PPIs. Exemplarily, we applied our technique to a knockdown screen of HeLa cells cultivated at standard conditions. Using a machine learning approach, we obtained high accuracy (82% AUC of the receiver operating characteristics) by cross-validation using 6,870 known activating and inhibiting PPIs as gold standard. We predicted de novo unknown activating and inhibiting effects for 1,954 PPIs in HeLa cells covering the ten major signaling pathways of the Kyoto Encyclopedia of Genes and Genomes, and made these predictions publicly available in a database. We finally demonstrate that the predicted effects can be used to cluster knockdown genes of similar biological processes in coherent subgroups. The characterization of the activating or inhibiting effect of individual PPIs opens up new perspectives for the interpretation of large datasets of PPIs and thus considerably increases the value of PPIs as an integrated resource for studying the detailed function of signaling pathways of the cellular system of interest. PMID:25255318

Interaction of the tick immune system with transmitted pathogens

PubMed Central

Hajdušek, Ondřej; Šíma, Radek; Ayllón, Nieves; Jalovecká, Marie; Perner, Jan; de la Fuente, José; Kopáček, Petr

2013-01-01

Ticks are hematophagous arachnids transmitting a wide variety of pathogens including viruses, bacteria, and protozoans to their vertebrate hosts. The tick vector competence has to be intimately linked to the ability of transmitted pathogens to evade tick defense mechanisms encountered on their route through the tick body comprising midgut, hemolymph, salivary glands or ovaries. Tick innate immunity is, like in other invertebrates, based on an orchestrated action of humoral and cellular immune responses. The direct antimicrobial defense in ticks is accomplished by a variety of small molecules such as defensins, lysozymes or by tick-specific antimicrobial compounds such as microplusin/hebraein or 5.3-kDa family proteins. Phagocytosis of the invading microbes by tick hemocytes is likely mediated by the primordial complement-like system composed of thioester-containing proteins, fibrinogen-related lectins and convertase-like factors. Moreover, an important role in survival of the ingested microbes seems to be played by host proteins and redox balance maintenance in the tick midgut. Here, we summarize recent knowledge about the major components of tick immune system and focus on their interaction with the relevant tick-transmitted pathogens, represented by spirochetes (Borrelia), rickettsiae (Anaplasma), and protozoans (Babesia). Availability of the tick genomic database and feasibility of functional genomics based on RNA interference greatly contribute to the understanding of molecular and cellular interplay at the tick-pathogen interface and may provide new targets for blocking the transmission of tick pathogens. PMID:23875177

New glycoproteomics software, GlycoPep Evaluator, generates decoy glycopeptides de novo and enables accurate false discovery rate analysis for small data sets.

PubMed

Zhu, Zhikai; Su, Xiaomeng; Go, Eden P; Desaire, Heather

2014-09-16

Glycoproteins are biologically significant large molecules that participate in numerous cellular activities. In order to obtain site-specific protein glycosylation information, intact glycopeptides, with the glycan attached to the peptide sequence, are characterized by tandem mass spectrometry (MS/MS) methods such as collision-induced dissociation (CID) and electron transfer dissociation (ETD). While several emerging automated tools are developed, no consensus is present in the field about the best way to determine the reliability of the tools and/or provide the false discovery rate (FDR). A common approach to calculate FDRs for glycopeptide analysis, adopted from the target-decoy strategy in proteomics, employs a decoy database that is created based on the target protein sequence database. Nonetheless, this approach is not optimal in measuring the confidence of N-linked glycopeptide matches, because the glycopeptide data set is considerably smaller compared to that of peptides, and the requirement of a consensus sequence for N-glycosylation further limits the number of possible decoy glycopeptides tested in a database search. To address the need to accurately determine FDRs for automated glycopeptide assignments, we developed GlycoPep Evaluator (GPE), a tool that helps to measure FDRs in identifying glycopeptides without using a decoy database. GPE generates decoy glycopeptides de novo for every target glycopeptide, in a 1:20 target-to-decoy ratio. The decoys, along with target glycopeptides, are scored against the ETD data, from which FDRs can be calculated accurately based on the number of decoy matches and the ratio of the number of targets to decoys, for small data sets. GPE is freely accessible for download and can work with any search engine that interprets ETD data of N-linked glycopeptides. The software is provided at https://desairegroup.ku.edu/research.

Proteomic analysis of secreted proteins by human bronchial epithelial cells in response to cadmium toxicity.

PubMed

Chen, De-Ju; Xu, Yan-Ming; Zheng, Wei; Huang, Dong-Yang; Wong, Wing-Yan; Tai, William Chi-Shing; Cho, Yong-Yeon; Lau, Andy T Y

2015-09-01

For years, many studies have been conducted to investigate the intracellular response of cells challenged with toxic metal(s), yet, the corresponding secretome responses, especially in human lung cells, are largely unexplored. Here, we provide a secretome analysis of human bronchial epithelial cells (BEAS-2B) treated with cadmium chloride (CdCl2 ), with the aim of identifying secreted proteins in response to Cd toxicity. Proteins from control and spent media were separated by two-dimensional electrophoresis and visualized by silver staining. Differentially-secreted proteins were identified by MALDI-TOF-MS analysis and database searching. We characterized, for the first time, the extracellular proteome changes of BEAS-2B dosed with Cd. Our results unveiled that Cd treatment led to the marked upregulation of molecular chaperones, antioxidant enzymes, enzymes associated with glutathione metabolic process, proteins involved in cellular energy metabolism, as well as tumor-suppressors. Pretreatment of cells with the thiol antioxidant glutathione before Cd treatment effectively abrogated the secretion of these proteins and prevented cell death. Taken together, our results demonstrate that Cd causes oxidative stress-induced cytotoxicity; and the differentially-secreted protein signatures could be considered as targets for potential use as extracellular biomarkers upon Cd exposure. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Integrating mRNA and Protein Sequencing Enables the Detection and Quantitative Profiling of Natural Protein Sequence Variants of Populus trichocarpa.

PubMed

Abraham, Paul E; Wang, Xiaojing; Ranjan, Priya; Nookaew, Intawat; Zhang, Bing; Tuskan, Gerald A; Hettich, Robert L

2015-12-04

Next-generation sequencing has transformed the ability to link genotypes to phenotypes and facilitates the dissection of genetic contribution to complex traits. However, it is challenging to link genetic variants with the perturbed functional effects on proteins encoded by such genes. Here we show how RNA sequencing can be exploited to construct genotype-specific protein sequence databases to assess natural variation in proteins, providing information about the molecular toolbox driving cellular processes. For this study, we used two natural genotypes selected from a recent genome-wide association study of Populus trichocarpa, an obligate outcrosser with tremendous phenotypic variation across the natural population. This strategy allowed us to comprehensively catalogue proteins containing single amino acid polymorphisms (SAAPs), as well as insertions and deletions. We profiled the frequency of 128 types of naturally occurring amino acid substitutions, including both expected (neutral) and unexpected (non-neutral) SAAPs, with a subset occurring in regions of the genome having strong polymorphism patterns consistent with recent positive and/or divergent selection. By zeroing in on the molecular signatures of these important regions that might have previously been uncharacterized, we now provide a high-resolution molecular inventory that should improve accessibility and subsequent identification of natural protein variants in future genotype-to-phenotype studies.

Proteomic Analysis of Pathogenic and Attenuated Alcelaphine Herpesvirus 1▿

PubMed Central

Dry, Inga; Haig, David M.; Inglis, Neil F.; Imrie, Lisa; Stewart, James P.; Russell, George C.

2008-01-01

The gammaherpesvirus alcelaphine herpesvirus 1 (AlHV-1) causes malignant catarrhal fever in susceptible ungulates but infects its natural host, wildebeest, without obvious clinical signs. In tissue culture, AlHV-1 is initially predominantly cell associated and virulent but on extended culture becomes cell-free and attenuated. We wanted to determine what changes in protein composition had taken place during the transition from virulent to attenuated virus in culture. Purified virus preparations were fractionated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and proteins were analyzed by liquid chromatography-electrospray ionization-tandem mass spectrometry. Peptides were identified in serial gel slices by using MASCOT software to interrogate virus-specific and nonredundant sequence databases. Twenty-three AlHV-1-encoded proteins and six cellular proteins were identified in the attenuated and virulent viruses. Two polypeptides were detected in only the virulent virus preparations, while one other protein was found in only the attenuated virus. Two of these virus-specific proteins were identified by a single peptide, suggesting that these may be low-abundance virion proteins rather than markers of attenuation or pathogenesis. The results suggest that attenuation of AlHV-1 is not the result of gross changes in the composition of the virus particle but probably due to altered viral gene expression in the infected cell. PMID:18353942

Semantic integration to identify overlapping functional modules in protein interaction networks

PubMed Central

Cho, Young-Rae; Hwang, Woochang; Ramanathan, Murali; Zhang, Aidong

2007-01-01

Background The systematic analysis of protein-protein interactions can enable a better understanding of cellular organization, processes and functions. Functional modules can be identified from the protein interaction networks derived from experimental data sets. However, these analyses are challenging because of the presence of unreliable interactions and the complex connectivity of the network. The integration of protein-protein interactions with the data from other sources can be leveraged for improving the effectiveness of functional module detection algorithms. Results We have developed novel metrics, called semantic similarity and semantic interactivity, which use Gene Ontology (GO) annotations to measure the reliability of protein-protein interactions. The protein interaction networks can be converted into a weighted graph representation by assigning the reliability values to each interaction as a weight. We presented a flow-based modularization algorithm to efficiently identify overlapping modules in the weighted interaction networks. The experimental results show that the semantic similarity and semantic interactivity of interacting pairs were positively correlated with functional co-occurrence. The effectiveness of the algorithm for identifying modules was evaluated using functional categories from the MIPS database. We demonstrated that our algorithm had higher accuracy compared to other competing approaches. Conclusion The integration of protein interaction networks with GO annotation data and the capability of detecting overlapping modules substantially improve the accuracy of module identification. PMID:17650343

Exploring Site-Specific N-Glycosylation Microheterogeneity of Haptoglobin using Glycopeptide CID Tandem Mass Spectra and Glycan Database Search

PubMed Central

Chandler, Kevin Brown; Pompach, Petr; Goldman, Radoslav

2013-01-01

Glycosylation is a common protein modification with a significant role in many vital cellular processes and human diseases, making the characterization of protein-attached glycan structures important for understanding cell biology and disease processes. Direct analysis of protein N-glycosylation by tandem mass spectrometry of glycopeptides promises site-specific elucidation of N-glycan microheterogeneity, something which detached N-glycan and de-glycosylated peptide analyses cannot provide. However, successful implementation of direct N-glycopeptide analysis by tandem mass spectrometry remains a challenge. In this work, we consider algorithmic techniques for the analysis of LC-MS/MS data acquired from glycopeptide-enriched fractions of enzymatic digests of purified proteins. We implement a computational strategy which takes advantage of the properties of CID fragmentation spectra of N-glycopeptides, matching the MS/MS spectra to peptide-glycan pairs from protein sequences and glycan structure databases. Significantly, we also propose a novel false-discovery-rate estimation technique to estimate and manage the number of false identifications. We use a human glycoprotein standard, haptoglobin, digested with trypsin and GluC, enriched for glycopeptides using HILIC chromatography, and analyzed by LC-MS/MS to demonstrate our algorithmic strategy and evaluate its performance. Our software, GlycoPeptideSearch (GPS), assigned glycopeptide identifications to 246 of the spectra at false-discovery-rate 5.58%, identifying 42 distinct haptoglobin peptide-glycan pairs at each of the four haptoglobin N-linked glycosylation sites. We further demonstrate the effectiveness of this approach by analyzing plasma-derived haptoglobin, identifying 136 N-linked glycopeptide spectra at false-discovery-rate 0.4%, representing 15 distinct glycopeptides on at least three of the four N-linked glycosylation sites. The software, GlycoPeptideSearch, is available for download from http://edwardslab.bmcb.georgetown.edu/GPS. PMID:23829323

Comprehensive inventory of protein complexes in the Protein Data Bank from consistent classification of interfaces.

PubMed

Bordner, Andrew J; Gorin, Andrey A

2008-05-12

Protein-protein interactions are ubiquitous and essential for all cellular processes. High-resolution X-ray crystallographic structures of protein complexes can reveal the details of their function and provide a basis for many computational and experimental approaches. Differentiation between biological and non-biological contacts and reconstruction of the intact complex is a challenging computational problem. A successful solution can provide additional insights into the fundamental principles of biological recognition and reduce errors in many algorithms and databases utilizing interaction information extracted from the Protein Data Bank (PDB). We have developed a method for identifying protein complexes in the PDB X-ray structures by a four step procedure: (1) comprehensively collecting all protein-protein interfaces; (2) clustering similar protein-protein interfaces together; (3) estimating the probability that each cluster is relevant based on a diverse set of properties; and (4) combining these scores for each PDB entry in order to predict the complex structure. The resulting clusters of biologically relevant interfaces provide a reliable catalog of evolutionary conserved protein-protein interactions. These interfaces, as well as the predicted protein complexes, are available from the Protein Interface Server (PInS) website (see Availability and requirements section). Our method demonstrates an almost two-fold reduction of the annotation error rate as evaluated on a large benchmark set of complexes validated from the literature. We also estimate relative contributions of each interface property to the accurate discrimination of biologically relevant interfaces and discuss possible directions for further improving the prediction method.

Shotgun proteomic analysis of Emiliania huxleyi, a marine phytoplankton species of major biogeochemical importance.

PubMed

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.

IFITM proteins-cellular inhibitors of viral entry.

PubMed

Smith, Se; Weston, S; Kellam, P; Marsh, M

2014-02-01

Interferon inducible transmembrane (IFITM) proteins are a recently discovered family of cellular anti-viral proteins that restrict the replication of a number of enveloped and non-enveloped viruses. IFITM proteins are located in the plasma membrane and endosomal membranes, the main portals of entry for many viruses. Biochemical and membrane fusion studies suggest IFITM proteins have the ability to inhibit viral entry, possibly by modulating the fluidity of cellular membranes. Here we discuss the IFITM proteins, recent work on their mode of action, and future directions for research. Copyright © 2014 Elsevier B.V. All rights reserved.

Transient Expression and Cellular Localization of Recombinant Proteins in Cultured Insect Cells.

PubMed

Fabrick, Jeffrey A; Hull, J Joe

2017-04-20

Heterologous protein expression systems are used for the production of recombinant proteins, the interpretation of cellular trafficking/localization, and the determination of the biochemical function of proteins at the sub-organismal level. Although baculovirus expression systems are increasingly used for protein production in numerous biotechnological, pharmaceutical, and industrial applications, nonlytic systems that do not involve viral infection have clear benefits but are often overlooked and underutilized. Here, we describe a method for generating nonlytic expression vectors and transient recombinant protein expression. This protocol allows for the efficient cellular localization of recombinant proteins and can be used to rapidly discern protein trafficking within the cell. We show the expression of four recombinant proteins in a commercially available insect cell line, including two aquaporin proteins from the insect Bemisia tabaci, as well as subcellular marker proteins specific for the cell plasma membrane and for intracellular lysosomes. All recombinant proteins were produced as chimeras with fluorescent protein markers at their carboxyl termini, which allows for the direct detection of the recombinant proteins. The double transfection of cells with plasmids harboring constructs for the genes of interest and a known subcellular marker allows for live cell imaging and improved validation of cellular protein localization.

Nuclear Cytoplasmic Trafficking of Proteins is a Major Response of Human Fibroblasts to Oxidative Stress

PubMed Central

Baqader, Noor O.; Radulovic, Marko; Crawford, Mark; Stoeber, Kai; Godovac-Zimmermann, Jasminka

2014-01-01

We have used a subcellular spatial razor approach based on LC–MS/MS-based proteomics with SILAC isotope labeling to determine changes in protein abundances in the nuclear and cytoplasmic compartments of human IMR90 fibroblasts subjected to mild oxidative stress. We show that response to mild tert-butyl hydrogen peroxide treatment includes redistribution between the nucleus and cytoplasm of numerous proteins not previously associated with oxidative stress. The 121 proteins with the most significant changes encompass proteins with known functions in a wide variety of subcellular locations and of cellular functional processes (transcription, signal transduction, autophagy, iron metabolism, TCA cycle, ATP synthesis) and are consistent with functional networks that are spatially dispersed across the cell. Both nuclear respiratory factor 2 and the proline regulatory axis appear to contribute to the cellular metabolic response. Proteins involved in iron metabolism or with iron/heme as a cofactor as well as mitochondrial proteins are prominent in the response. Evidence suggesting that nuclear import/export and vesicle-mediated protein transport contribute to the cellular response was obtained. We suggest that measurements of global changes in total cellular protein abundances need to be complemented with measurements of the dynamic subcellular spatial redistribution of proteins to obtain comprehensive pictures of cellular function. PMID:25133973

Prioritizing and modelling of putative drug target proteins of Candida albicans by systems biology approach.

PubMed

Ismail, Tariq; Fatima, Nighat; Muhammad, Syed Aun; Zaidi, Syed Saoud; Rehman, Nisar; Hussain, Izhar; Tariq, Najam Us Sahr; Amirzada, Imran; Mannan, Abdul

2018-01-01

Candida albicans (Candida albicans) is one of the major sources of nosocomial infections in humans which may prove fatal in 30% of cases. The hospital acquired infection is very difficult to treat affectively due to the presence of drug resistant pathogenic strains, therefore there is a need to find alternative drug targets to cure this infection. In silico and computational level frame work was used to prioritize and establish antifungal drug targets of Candida albicans. The identification of putative drug targets was based on acquiring 5090 completely annotated genes of Candida albicans from available databases which were categorized into essential and non-essential genes. The result indicated that 9% of proteins were essential and could become potential candidates for intervention which might result in pathogen eradication. We studied cluster of orthologs and the subtractive genomic analysis of these essential proteins against human genome was made as a reference to minimize the side effects. It was seen that 14% of Candida albicans proteins were evolutionary related to the human proteins while 86% are non-human homologs. In the next step of compatible drug target selections, the non-human homologs were sequentially compared to the human microbiome data to minimize the potential effects against gut flora which accumulated to 38% of the essential genome. The sub-cellular localization of these candidate proteins in fungal cellular systems indicated that 80% of them are cytoplasmic, 10% are mitochondrial and the remaining 10% are associated with the cell wall. The role of these non-human and non-gut flora putative target proteins in Candida albicans biological pathways was studied. Due to their integrated and critical role in Candida albicans replication cycle, four proteins were selected for molecular modeling. For drug designing and development, four high quality and reliable protein models with more than 70% sequence identity were constructed. These proteins are used for the docking studies of the known and new ligands (unpublished data). Our study will be an effective framework for drug target identifications of pathogenic microbial strains and development of new therapies against the infections they cause.

Enhancing a Pathway-Genome Database (PGDB) to capture subcellular localization of metabolites and enzymes: the nucleotide-sugar biosynthetic pathways of Populus trichocarpa.

PubMed

Nag, Ambarish; Karpinets, Tatiana V; Chang, Christopher H; Bar-Peled, Maor

2012-01-01

Understanding how cellular metabolism works and is regulated requires that the underlying biochemical pathways be adequately represented and integrated with large metabolomic data sets to establish a robust network model. Genetically engineering energy crops to be less recalcitrant to saccharification requires detailed knowledge of plant polysaccharide structures and a thorough understanding of the metabolic pathways involved in forming and regulating cell-wall synthesis. Nucleotide-sugars are building blocks for synthesis of cell wall polysaccharides. The biosynthesis of nucleotide-sugars is catalyzed by a multitude of enzymes that reside in different subcellular organelles, and precise representation of these pathways requires accurate capture of this biological compartmentalization. The lack of simple localization cues in genomic sequence data and annotations however leads to missing compartmentalization information for eukaryotes in automatically generated databases, such as the Pathway-Genome Databases (PGDBs) of the SRI Pathway Tools software that drives much biochemical knowledge representation on the internet. In this report, we provide an informal mechanism using the existing Pathway Tools framework to integrate protein and metabolite sub-cellular localization data with the existing representation of the nucleotide-sugar metabolic pathways in a prototype PGDB for Populus trichocarpa. The enhanced pathway representations have been successfully used to map SNP abundance data to individual nucleotide-sugar biosynthetic genes in the PGDB. The manually curated pathway representations are more conducive to the construction of a computational platform that will allow the simulation of natural and engineered nucleotide-sugar precursor fluxes into specific recalcitrant polysaccharide(s). Database URL: The curated Populus PGDB is available in the BESC public portal at http://cricket.ornl.gov/cgi-bin/beocyc_home.cgi and the nucleotide-sugar biosynthetic pathways can be directly accessed at http://cricket.ornl.gov:1555/PTR/new-image?object=SUGAR-NUCLEOTIDES.

Enhancing a Pathway-Genome Database (PGDB) to capture subcellular localization of metabolites and enzymes: the nucleotide-sugar biosynthetic pathways of Populus trichocarpa

PubMed Central

Nag, Ambarish; Karpinets, Tatiana V.; Chang, Christopher H.; Bar-Peled, Maor

2012-01-01

Understanding how cellular metabolism works and is regulated requires that the underlying biochemical pathways be adequately represented and integrated with large metabolomic data sets to establish a robust network model. Genetically engineering energy crops to be less recalcitrant to saccharification requires detailed knowledge of plant polysaccharide structures and a thorough understanding of the metabolic pathways involved in forming and regulating cell-wall synthesis. Nucleotide-sugars are building blocks for synthesis of cell wall polysaccharides. The biosynthesis of nucleotide-sugars is catalyzed by a multitude of enzymes that reside in different subcellular organelles, and precise representation of these pathways requires accurate capture of this biological compartmentalization. The lack of simple localization cues in genomic sequence data and annotations however leads to missing compartmentalization information for eukaryotes in automatically generated databases, such as the Pathway-Genome Databases (PGDBs) of the SRI Pathway Tools software that drives much biochemical knowledge representation on the internet. In this report, we provide an informal mechanism using the existing Pathway Tools framework to integrate protein and metabolite sub-cellular localization data with the existing representation of the nucleotide-sugar metabolic pathways in a prototype PGDB for Populus trichocarpa. The enhanced pathway representations have been successfully used to map SNP abundance data to individual nucleotide-sugar biosynthetic genes in the PGDB. The manually curated pathway representations are more conducive to the construction of a computational platform that will allow the simulation of natural and engineered nucleotide-sugar precursor fluxes into specific recalcitrant polysaccharide(s). Database URL: The curated Populus PGDB is available in the BESC public portal at http://cricket.ornl.gov/cgi-bin/beocyc_home.cgi and the nucleotide-sugar biosynthetic pathways can be directly accessed at http://cricket.ornl.gov:1555/PTR/new-image?object=SUGAR-NUCLEOTIDES. PMID:22465851

MIPS: a database for genomes and protein sequences.

PubMed Central

Mewes, H W; Heumann, K; Kaps, A; Mayer, K; Pfeiffer, F; Stocker, S; Frishman, D

1999-01-01

The Munich Information Center for Protein Sequences (MIPS-GSF), Martinsried near Munich, Germany, develops and maintains genome oriented databases. It is commonplace that the amount of sequence data available increases rapidly, but not the capacity of qualified manual annotation at the sequence databases. Therefore, our strategy aims to cope with the data stream by the comprehensive application of analysis tools to sequences of complete genomes, the systematic classification of protein sequences and the active support of sequence analysis and functional genomics projects. This report describes the systematic and up-to-date analysis of genomes (PEDANT), a comprehensive database of the yeast genome (MYGD), a database reflecting the progress in sequencing the Arabidopsis thaliana genome (MATD), the database of assembled, annotated human EST clusters (MEST), and the collection of protein sequence data within the framework of the PIR-International Protein Sequence Database (described elsewhere in this volume). MIPS provides access through its WWW server (http://www.mips.biochem.mpg.de) to a spectrum of generic databases, including the above mentioned as well as a database of protein families (PROTFAM), the MITOP database, and the all-against-all FASTA database. PMID:9847138

The Halophile protein database.

PubMed

Sharma, Naveen; Farooqi, Mohammad Samir; Chaturvedi, Krishna Kumar; Lal, Shashi Bhushan; Grover, Monendra; Rai, Anil; Pandey, Pankaj

2014-01-01

Halophilic archaea/bacteria adapt to different salt concentration, namely extreme, moderate and low. These type of adaptations may occur as a result of modification of protein structure and other changes in different cell organelles. Thus proteins may play an important role in the adaptation of halophilic archaea/bacteria to saline conditions. The Halophile protein database (HProtDB) is a systematic attempt to document the biochemical and biophysical properties of proteins from halophilic archaea/bacteria which may be involved in adaptation of these organisms to saline conditions. In this database, various physicochemical properties such as molecular weight, theoretical pI, amino acid composition, atomic composition, estimated half-life, instability index, aliphatic index and grand average of hydropathicity (Gravy) have been listed. These physicochemical properties play an important role in identifying the protein structure, bonding pattern and function of the specific proteins. This database is comprehensive, manually curated, non-redundant catalogue of proteins. The database currently contains 59 897 proteins properties extracted from 21 different strains of halophilic archaea/bacteria. The database can be accessed through link. Database URL: http://webapp.cabgrid.res.in/protein/ © The Author(s) 2014. Published by Oxford University Press.

The Safety Dance: Biophysics of Membrane Protein Folding and Misfolding in a Cellular Context

PubMed Central

Schlebach, Jonathan P.; Sanders, Charles R.

2015-01-01

Most biological processes require the production and degradation of proteins, a task that weighs heavily on the cell. Mutations that compromise the conformational stability of proteins place both specific and general burdens on cellular protein homeostasis (proteostasis) in ways that contribute to numerous diseases. Efforts to elucidate the chain of molecular events responsible for diseases of protein folding address one of the foremost challenges in biomedical science. However, relatively little is known about the processes by which mutations prompt the misfolding of α-helical membrane proteins, which rely on an intricate network of cellular machinery to acquire and maintain their functional structures within cellular membranes. In this review, we summarize the current understanding of the physical principles that guide membrane protein biogenesis and folding in the context of mammalian cells. Additionally, we explore how pathogenic mutations that influence biogenesis may differ from those that disrupt folding and assembly, as well as how this may relate to disease mechanisms and therapeutic intervention. These perspectives indicate an imperative for the use of information from structural, cellular, and biochemical studies of membrane proteins in the design of novel therapeutics and in personalized medicine. PMID:25420508

Transient expression and cellular localization of recombinant proteins in cultured insect cells

USDA-ARS?s Scientific Manuscript database

Heterologous protein expression systems are used for production of recombinant proteins, interpretation of cellular trafficking/localization, and for the determination of biochemical function of proteins at the sub-organismal level. Although baculovirus expression systems are increasingly used for ...

A Proteomic Approach to Investigate the Drought Response in the Orphan Crop Eragrostis tef.

PubMed

Kamies, Rizqah; Farrant, Jill M; Tadele, Zerihun; Cannarozzi, Gina; Rafudeen, Mohammed Suhail

2017-11-15

The orphan crop, Eragrostis tef , was subjected to controlled drought conditions to observe the physiological parameters and proteins changing in response to dehydration stress. Physiological measurements involving electrolyte leakage, chlorophyll fluorescence and ultra-structural analysis showed tef plants tolerated water loss to 50% relative water content (RWC) before adverse effects in leaf tissues were observed. Proteomic analysis using isobaric tag for relative and absolute quantification (iTRAQ) mass spectrometry and appropriate database searching enabled the detection of 5727 proteins, of which 211 proteins, including a number of spliced variants, were found to be differentially regulated with the imposed stress conditions. Validation of the iTRAQ dataset was done with selected stress-related proteins, fructose-bisphosphate aldolase (FBA) and the protective antioxidant proteins, monodehydroascorbate reductase (MDHAR) and peroxidase (POX). Western blot analyses confirmed protein presence and showed increased protein abundance levels during water deficit while enzymatic activity for FBA, MDHAR and POX increased at selected RWC points. Gene ontology (GO)-term enrichment and analysis revealed terms involved in biotic and abiotic stress response, signaling, transport, cellular homeostasis and pentose metabolic processes, to be enriched in tef upregulated proteins, while terms linked to reactive oxygen species (ROS)-producing processes under water-deficit, such as photosynthesis and associated light harvesting reactions, manganese transport and homeostasis, the synthesis of sugars and cell wall catabolism and modification, to be enriched in tef downregulated proteins.

MIPS: analysis and annotation of proteins from whole genomes

PubMed Central

Mewes, H. W.; Amid, C.; Arnold, R.; Frishman, D.; Güldener, U.; Mannhaupt, G.; Münsterkötter, M.; Pagel, P.; Strack, N.; Stümpflen, V.; Warfsmann, J.; Ruepp, A.

2004-01-01

The Munich Information Center for Protein Sequences (MIPS-GSF), Neuherberg, Germany, provides protein sequence-related information based on whole-genome analysis. The main focus of the work is directed toward the systematic organization of sequence-related attributes as gathered by a variety of algorithms, primary information from experimental data together with information compiled from the scientific literature. MIPS maintains automatically generated and manually annotated genome-specific databases, develops systematic classification schemes for the functional annotation of protein sequences and provides tools for the comprehensive analysis of protein sequences. This report updates the information on the yeast genome (CYGD), the Neurospora crassa genome (MNCDB), the database of complete cDNAs (German Human Genome Project, NGFN), the database of mammalian protein–protein interactions (MPPI), the database of FASTA homologies (SIMAP), and the interface for the fast retrieval of protein-associated information (QUIPOS). The Arabidopsis thaliana database, the rice database, the plant EST databases (MATDB, MOsDB, SPUTNIK), as well as the databases for the comprehensive set of genomes (PEDANT genomes) are described elsewhere in the 2003 and 2004 NAR database issues, respectively. All databases described, and the detailed descriptions of our projects can be accessed through the MIPS web server (http://mips.gsf.de). PMID:14681354

Temporal proteomic analysis of HIV infection reveals remodelling of the host phosphoproteome by lentiviral Vif variants

PubMed Central

Greenwood, Edward JD; Matheson, Nicholas J; Wals, Kim; van den Boomen, Dick JH; Antrobus, Robin; Williamson, James C; Lehner, Paul J

2016-01-01

Viruses manipulate host factors to enhance their replication and evade cellular restriction. We used multiplex tandem mass tag (TMT)-based whole cell proteomics to perform a comprehensive time course analysis of >6500 viral and cellular proteins during HIV infection. To enable specific functional predictions, we categorized cellular proteins regulated by HIV according to their patterns of temporal expression. We focussed on proteins depleted with similar kinetics to APOBEC3C, and found the viral accessory protein Vif to be necessary and sufficient for CUL5-dependent proteasomal degradation of all members of the B56 family of regulatory subunits of the key cellular phosphatase PP2A (PPP2R5A-E). Quantitative phosphoproteomic analysis of HIV-infected cells confirmed Vif-dependent hyperphosphorylation of >200 cellular proteins, particularly substrates of the aurora kinases. The ability of Vif to target PPP2R5 subunits is found in primate and non-primate lentiviral lineages, and remodeling of the cellular phosphoproteome is therefore a second ancient and conserved Vif function. DOI: http://dx.doi.org/10.7554/eLife.18296.001 PMID:27690223

Identification and characterization of a novel ISG15-ubiquitin mixed chain and its role in regulating protein homeostasis

PubMed Central

Fan, Jun-Bao; Arimoto, Kei-lchiro; Motamedchaboki, Khatereh; Yan, Ming; Wolf, Dieter A.; Zhang, Dong-Er

2015-01-01

As a ubiquitin-like modifier, ISG15 is conjugated to many cellular proteins in a process termed protein ISGylation. However, the crosstalk between protein ISGylation and the ubiquitin proteasome system is not fully understood. Here, we report that cellular ubiquitin is a substrate of ISG15 and Lys 29 on ubiquitin is the major ISG15 acceptor site. Using a model substrate, we demonstrate that ISG15 can modify ubiquitin, which is immobilized on its substrate, to form ISG15-ubiquitin mixed chains. Furthermore, our results indicate that ISG15-ubiquitin mixed chains do not serve as degradation signals for a ubiquitin fusion degradation substrate. Accordingly, an ISG15-ubiquitin fusion protein, which mimics an ISG15-ubiquitin mixed chain, negatively regulates cellular turnover of ubiquitylated proteins. In addition, ISG15-ubiquitin mixed chains, which are detectable on endogenously ubiquitylated proteins, dampen cellular turnover of these proteins. Thus, our studies unveil an unanticipated interplay between two protein modification systems and highlight its role in coordinating protein homeostasis. PMID:26226047

The Nuclear Protein Database (NPD): sub-nuclear localisation and functional annotation of the nuclear proteome

PubMed Central

Dellaire, G.; Farrall, R.; Bickmore, W.A.

2003-01-01

The Nuclear Protein Database (NPD) is a curated database that contains information on more than 1300 vertebrate proteins that are thought, or are known, to localise to the cell nucleus. Each entry is annotated with information on predicted protein size and isoelectric point, as well as any repeats, motifs or domains within the protein sequence. In addition, information on the sub-nuclear localisation of each protein is provided and the biological and molecular functions are described using Gene Ontology (GO) terms. The database is searchable by keyword, protein name, sub-nuclear compartment and protein domain/motif. Links to other databases are provided (e.g. Entrez, SWISS-PROT, OMIM, PubMed, PubMed Central). Thus, NPD provides a gateway through which the nuclear proteome may be explored. The database can be accessed at http://npd.hgu.mrc.ac.uk and is updated monthly. PMID:12520015

Phylogenetic Characterization of Transport Protein Superfamilies: Superiority of SuperfamilyTree Programs over Those Based on Multiple Alignments

PubMed Central

Chen, Jonathan S.; Reddy, Vamsee; Chen, Joshua H.; Shlykov, Maksim A.; Zheng, Wei Hao; Cho, Jaehoon; Yen, Ming Ren; Saier, Milton H.

2012-01-01

Transport proteins function in the translocation of ions, solutes and macromolecules across cellular and organellar membranes. These integral membrane proteins fall into >600 families as tabulated in the Transporter Classification Database (www.tcdb.org). Recent studies, some of which are reported here, define distant phylogenetic relationships between families with the creation of superfamilies. Several of these are analyzed using a novel set of programs designed to allow reliable prediction of phylogenetic trees when sequence divergence is too great to allow the use of multiple alignments. These new programs, called SuperfamilyTree1 and 2 (SFT1 and 2), allow display of protein and family relationships, respectively, based on thousands of comparative BLAST scores rather than multiple alignments. Superfamilies analyzed include: (1) Aerolysins, (2) RTX Toxins, (3) Defensins, (4) Ion Transporters, (5) Bile/Arsenite/Riboflavin Transporters, (6) Cation: Proton Antiporters, and (7) the Glucose/Fructose/Lactose superfamily within the prokaryotic phosphoenol pyruvate-dependent Phosphotransferase System. In addition to defining the phylogenetic relationships of the proteins and families within these seven superfamilies, evidence is provided showing that the SFT programs outperform programs that are based on multiple alignments whenever sequence divergence of superfamily members is extensive. The SFT programs should be applicable to virtually any superfamily of proteins or nucleic acids. PMID:22286036

Identification of Modules in Protein-Protein Interaction Networks

NASA Astrophysics Data System (ADS)

Erten, Sinan; Koyutürk, Mehmet

In biological systems, most processes are carried out through orchestration of multiple interacting molecules. These interactions are often abstracted using network models. A key feature of cellular networks is their modularity, which contributes significantly to the robustness, as well as adaptability of biological systems. Therefore, modularization of cellular networks is likely to be useful in obtaining insights into the working principles of cellular systems, as well as building tractable models of cellular organization and dynamics. A common, high-throughput source of data on molecular interactions is in the form of physical interactions between proteins, which are organized into protein-protein interaction (PPI) networks. This chapter provides an overview on identification and analysis of functional modules in PPI networks, which has been an active area of research in the last decade.

Scaffolding protein RanBPM and its interactions in diverse signaling pathways in health and disease.

PubMed

Das, Soumyadip; Haq, Saba; Ramakrishna, Suresh

2018-04-01

Ran-binding protein in the microtubule-organizing center (RanBPM) is an evolutionarily conserved, nucleocytoplasmic scaffolding protein involved in various cellular processes and several signal transduction pathways. RanBPM has a crucial role in mediating disease pathology by interacting with diverse proteins to regulate their functions. Previously, we compiled diverse cellular functions of RanBPM. Since then the functions of RanBPM have increased exponentially. In this article, we have updated the functions of RanBPM through its manifold interactions that have been investigated to date, according to their roles in protein stability, transcriptional activity, cellular development, neurobiology, and the cell cycle. Our review provides a complete guide on RanBPM interactors, the physiological role of RanBPM in cellular functions, and potential applications in disease therapeutics.

Proteogenomic database construction driven from large scale RNA-seq data.

PubMed

Woo, Sunghee; Cha, Seong Won; Merrihew, Gennifer; He, Yupeng; Castellana, Natalie; Guest, Clark; MacCoss, Michael; Bafna, Vineet

2014-01-03

The advent of inexpensive RNA-seq technologies and other deep sequencing technologies for RNA has the promise to radically improve genomic annotation, providing information on transcribed regions and splicing events in a variety of cellular conditions. Using MS-based proteogenomics, many of these events can be confirmed directly at the protein level. However, the integration of large amounts of redundant RNA-seq data and mass spectrometry data poses a challenging problem. Our paper addresses this by construction of a compact database that contains all useful information expressed in RNA-seq reads. Applying our method to cumulative C. elegans data reduced 496.2 GB of aligned RNA-seq SAM files to 410 MB of splice graph database written in FASTA format. This corresponds to 1000× compression of data size, without loss of sensitivity. We performed a proteogenomics study using the custom data set, using a completely automated pipeline, and identified a total of 4044 novel events, including 215 novel genes, 808 novel exons, 12 alternative splicings, 618 gene-boundary corrections, 245 exon-boundary changes, 938 frame shifts, 1166 reverse strands, and 42 translated UTRs. Our results highlight the usefulness of transcript + proteomic integration for improved genome annotations.

Purine inhibitors of protein kinases, G proteins and polymerases

DOEpatents

Gray, Nathanael S.; Schultz, Peter; Kim, Sung-Hou; Meijer, Laurent

2004-10-12

The present invention relates to 2-N-substituted 6-(4-methoxybenzylamino)-9-isopropylpurines that inhibit, inter alia, protein kinases, G-proteins and polymerases. In addition, the present invention relates to methods of using such 2-N-substituted 6-(4-methoxybenzylamino)-9-isopropylpurines to inhibit protein kinases, G-proteins, polymerases and other cellular processes and to treat cellular proliferative diseases.

Two potato proteins, including a novel RING finger protein (HIP1), interact with the potyviral multifunctional protein HCpro.

PubMed

Guo, Deyin; Spetz, Carl; Saarma, Mart; Valkonen, Jari P T

2003-05-01

Potyviral helper-component proteinase (HCpro) is a multifunctional protein exerting its cellular functions in interaction with putative host proteins. In this study, cellular protein partners of the HCpro encoded by Potato virus A (PVA) (genus Potyvirus) were screened in a potato leaf cDNA library using a yeast two-hybrid system. Two cellular proteins were obtained that interact specifically with PVA HCpro in yeast and in the two in vitro binding assays used. Both proteins are encoded by single-copy genes in the potato genome. Analysis of the deduced amino acid sequences revealed that one (HIP1) of the two HCpro interactors is a novel RING finger protein. The sequence of the other protein (HIP2) showed no resemblance to the protein sequences available from databanks and has known biological functions.

Generic framework for mining cellular automata models on protein-folding simulations.

PubMed

Diaz, N; Tischer, I

2016-05-13

Cellular automata model identification is an important way of building simplified simulation models. In this study, we describe a generic architectural framework to ease the development process of new metaheuristic-based algorithms for cellular automata model identification in protein-folding trajectories. Our framework was developed by a methodology based on design patterns that allow an improved experience for new algorithms development. The usefulness of the proposed framework is demonstrated by the implementation of four algorithms, able to obtain extremely precise cellular automata models of the protein-folding process with a protein contact map representation. Dynamic rules obtained by the proposed approach are discussed, and future use for the new tool is outlined.

BioImaging Database

DOE Office of Scientific and Technical Information (OSTI.GOV)

David Nix, Lisa Simirenko

2006-10-25

The Biolmaging Database (BID) is a relational database developed to store the data and meta-data for the 3D gene expression in early Drosophila embryo development on a cellular level. The schema was written to be used with the MySQL DBMS but with minor modifications can be used on any SQL compliant relational DBMS.

MIPS: a database for genomes and protein sequences

PubMed Central

Mewes, H. W.; Frishman, D.; Güldener, U.; Mannhaupt, G.; Mayer, K.; Mokrejs, M.; Morgenstern, B.; Münsterkötter, M.; Rudd, S.; Weil, B.

2002-01-01

The Munich Information Center for Protein Sequences (MIPS-GSF, Neuherberg, Germany) continues to provide genome-related information in a systematic way. MIPS supports both national and European sequencing and functional analysis projects, develops and maintains automatically generated and manually annotated genome-specific databases, develops systematic classification schemes for the functional annotation of protein sequences, and provides tools for the comprehensive analysis of protein sequences. This report updates the information on the yeast genome (CYGD), the Neurospora crassa genome (MNCDB), the databases for the comprehensive set of genomes (PEDANT genomes), the database of annotated human EST clusters (HIB), the database of complete cDNAs from the DHGP (German Human Genome Project), as well as the project specific databases for the GABI (Genome Analysis in Plants) and HNB (Helmholtz–Netzwerk Bioinformatik) networks. The Arabidospsis thaliana database (MATDB), the database of mitochondrial proteins (MITOP) and our contribution to the PIR International Protein Sequence Database have been described elsewhere [Schoof et al. (2002) Nucleic Acids Res., 30, 91–93; Scharfe et al. (2000) Nucleic Acids Res., 28, 155–158; Barker et al. (2001) Nucleic Acids Res., 29, 29–32]. All databases described, the protein analysis tools provided and the detailed descriptions of our projects can be accessed through the MIPS World Wide Web server (http://mips.gsf.de). PMID:11752246

MIPS: a database for genomes and protein sequences.

PubMed

Mewes, H W; Frishman, D; Güldener, U; Mannhaupt, G; Mayer, K; Mokrejs, M; Morgenstern, B; Münsterkötter, M; Rudd, S; Weil, B

2002-01-01

The Munich Information Center for Protein Sequences (MIPS-GSF, Neuherberg, Germany) continues to provide genome-related information in a systematic way. MIPS supports both national and European sequencing and functional analysis projects, develops and maintains automatically generated and manually annotated genome-specific databases, develops systematic classification schemes for the functional annotation of protein sequences, and provides tools for the comprehensive analysis of protein sequences. This report updates the information on the yeast genome (CYGD), the Neurospora crassa genome (MNCDB), the databases for the comprehensive set of genomes (PEDANT genomes), the database of annotated human EST clusters (HIB), the database of complete cDNAs from the DHGP (German Human Genome Project), as well as the project specific databases for the GABI (Genome Analysis in Plants) and HNB (Helmholtz-Netzwerk Bioinformatik) networks. The Arabidospsis thaliana database (MATDB), the database of mitochondrial proteins (MITOP) and our contribution to the PIR International Protein Sequence Database have been described elsewhere [Schoof et al. (2002) Nucleic Acids Res., 30, 91-93; Scharfe et al. (2000) Nucleic Acids Res., 28, 155-158; Barker et al. (2001) Nucleic Acids Res., 29, 29-32]. All databases described, the protein analysis tools provided and the detailed descriptions of our projects can be accessed through the MIPS World Wide Web server (http://mips.gsf.de).

Control of human adenovirus type 5 gene expression by cellular Daxx/ATRX chromatin-associated complexes

PubMed Central

Schreiner, Sabrina; Bürck, Carolin; Glass, Mandy; Groitl, Peter; Wimmer, Peter; Kinkley, Sarah; Mund, Andreas; Everett, Roger D.; Dobner, Thomas

2013-01-01

Death domain–associated protein (Daxx) cooperates with X-linked α-thalassaemia retardation syndrome protein (ATRX), a putative member of the sucrose non-fermentable 2 family of ATP-dependent chromatin-remodelling proteins, acting as the core ATPase subunit in this complex, whereas Daxx is the targeting factor, leading to histone deacetylase recruitment, H3.3 deposition and transcriptional repression of cellular promoters. Despite recent findings on the fundamental importance of chromatin modification in host-cell gene regulation, it remains unclear whether adenovirus type 5 (Ad5) transcription is regulated by cellular chromatin remodelling to allow efficient virus gene expression. Here, we focus on the repressive role of the Daxx/ATRX complex during Ad5 replication, which depends on intact protein–protein interaction, as negative regulation could be relieved with a Daxx mutant that is unable to interact with ATRX. To ensure efficient viral replication, Ad5 E1B-55K protein inhibits Daxx and targets ATRX for proteasomal degradation in cooperation with early region 4 open reading frame protein 6 and cellular components of a cullin-dependent E3-ubiquitin ligase. Our studies illustrate the importance and diversity of viral factors antagonizing Daxx/ATRX-mediated repression of viral gene expression and shed new light on the modulation of cellular chromatin remodelling factors by Ad5. We show for the first time that cellular Daxx/ATRX chromatin remodelling complexes play essential roles in Ad gene expression and illustrate the importance of early viral proteins to counteract cellular chromatin remodelling. PMID:23396441

An Internal Signal Sequence Directs Intramembrane Proteolysis of a Cellular Immunoglobulin Domain Protein*S⃞

PubMed Central

Robakis, Thalia; Bak, Beata; Lin, Shu-huei; Bernard, Daniel J.; Scheiffele, Peter

2008-01-01

Precursor proteolysis is a crucial mechanism for regulating protein structure and function. Signal peptidase (SP) is an enzyme with a well defined role in cleaving N-terminal signal sequences but no demonstrated function in the proteolysis of cellular precursor proteins. We provide evidence that SP mediates intraprotein cleavage of IgSF1, a large cellular Ig domain protein that is processed into two separate Ig domain proteins. In addition, our results suggest the involvement of signal peptide peptidase (SPP), an intramembrane protease, which acts on substrates that have been previously cleaved by SP. We show that IgSF1 is processed through sequential proteolysis by SP and SPP. Cleavage is directed by an internal signal sequence and generates two separate Ig domain proteins from a polytopic precursor. Our findings suggest that SP and SPP function are not restricted to N-terminal signal sequence cleavage but also contribute to the processing of cellular transmembrane proteins. PMID:18981173

Designer amphiphilic proteins as building blocks for the intracellular formation of organelle-like compartments

NASA Astrophysics Data System (ADS)

Huber, Matthias C.; Schreiber, Andreas; von Olshausen, Philipp; Varga, Balázs R.; Kretz, Oliver; Joch, Barbara; Barnert, Sabine; Schubert, Rolf; Eimer, Stefan; Kele, Péter; Schiller, Stefan M.

2015-01-01

Nanoscale biological materials formed by the assembly of defined block-domain proteins control the formation of cellular compartments such as organelles. Here, we introduce an approach to intentionally ‘program’ the de novo synthesis and self-assembly of genetically encoded amphiphilic proteins to form cellular compartments, or organelles, in Escherichia coli. These proteins serve as building blocks for the formation of artificial compartments in vivo in a similar way to lipid-based organelles. We investigated the formation of these organelles using epifluorescence microscopy, total internal reflection fluorescence microscopy and transmission electron microscopy. The in vivo modification of these protein-based de novo organelles, by means of site-specific incorporation of unnatural amino acids, allows the introduction of artificial chemical functionalities. Co-localization of membrane proteins results in the formation of functionalized artificial organelles combining artificial and natural cellular function. Adding these protein structures to the cellular machinery may have consequences in nanobiotechnology, synthetic biology and materials science, including the constitution of artificial cells and bio-based metamaterials.

Intelligent Interfaces for Mining Large-Scale RNAi-HCS Image Databases

PubMed Central

Lin, Chen; Mak, Wayne; Hong, Pengyu; Sepp, Katharine; Perrimon, Norbert

2010-01-01

Recently, High-content screening (HCS) has been combined with RNA interference (RNAi) to become an essential image-based high-throughput method for studying genes and biological networks through RNAi-induced cellular phenotype analyses. However, a genome-wide RNAi-HCS screen typically generates tens of thousands of images, most of which remain uncategorized due to the inadequacies of existing HCS image analysis tools. Until now, it still requires highly trained scientists to browse a prohibitively large RNAi-HCS image database and produce only a handful of qualitative results regarding cellular morphological phenotypes. For this reason we have developed intelligent interfaces to facilitate the application of the HCS technology in biomedical research. Our new interfaces empower biologists with computational power not only to effectively and efficiently explore large-scale RNAi-HCS image databases, but also to apply their knowledge and experience to interactive mining of cellular phenotypes using Content-Based Image Retrieval (CBIR) with Relevance Feedback (RF) techniques. PMID:21278820

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

PubMed

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 genome sequences, though there are over lapped proteins. Based on the demonstrated application of data stored in the database for functional analyses, it is suggested that these data will be useful for analyses of biological mechanisms in soybean. Furthermore, coupled with recent advances in information and communication technology, the usefulness of this database would increase in the analyses of biological mechanisms. Copyright © 2017 Elsevier B.V. All rights reserved.

Navigating through the Jungle of Allergens: Features and Applications of Allergen Databases.

PubMed

Radauer, Christian

2017-01-01

The increasing number of available data on allergenic proteins demanded the establishment of structured, freely accessible allergen databases. In this review article, features and applications of 6 of the most widely used allergen databases are discussed. The WHO/IUIS Allergen Nomenclature Database is the official resource of allergen designations. Allergome is the most comprehensive collection of data on allergens and allergen sources. AllergenOnline is aimed at providing a peer-reviewed database of allergen sequences for prediction of allergenicity of proteins, such as those planned to be inserted into genetically modified crops. The Structural Database of Allergenic Proteins (SDAP) provides a database of allergen sequences, structures, and epitopes linked to bioinformatics tools for sequence analysis and comparison. The Immune Epitope Database (IEDB) is the largest repository of T-cell, B-cell, and major histocompatibility complex protein epitopes including epitopes of allergens. AllFam classifies allergens into families of evolutionarily related proteins using definitions from the Pfam protein family database. These databases contain mostly overlapping data, but also show differences in terms of their targeted users, the criteria for including allergens, data shown for each allergen, and the availability of bioinformatics tools. © 2017 S. Karger AG, Basel.

Detection of alternative splice variants at the proteome level in Aspergillus flavus.

PubMed

Chang, Kung-Yen; Georgianna, D Ryan; Heber, Steffen; Payne, Gary A; Muddiman, David C

2010-03-05

Identification of proteins from proteolytic peptides or intact proteins plays an essential role in proteomics. Researchers use search engines to match the acquired peptide sequences to the target proteins. However, search engines depend on protein databases to provide candidates for consideration. Alternative splicing (AS), the mechanism where the exon of pre-mRNAs can be spliced and rearranged to generate distinct mRNA and therefore protein variants, enable higher eukaryotic organisms, with only a limited number of genes, to have the requisite complexity and diversity at the proteome level. Multiple alternative isoforms from one gene often share common segments of sequences. However, many protein databases only include a limited number of isoforms to keep minimal redundancy. As a result, the database search might not identify a target protein even with high quality tandem MS data and accurate intact precursor ion mass. We computationally predicted an exhaustive list of putative isoforms of Aspergillus flavus proteins from 20 371 expressed sequence tags to investigate whether an alternative splicing protein database can assign a greater proportion of mass spectrometry data. The newly constructed AS database provided 9807 new alternatively spliced variants in addition to 12 832 previously annotated proteins. The searches of the existing tandem MS spectra data set using the AS database identified 29 new proteins encoded by 26 genes. Nine fungal genes appeared to have multiple protein isoforms. In addition to the discovery of splice variants, AS database also showed potential to improve genome annotation. In summary, the introduction of an alternative splicing database helps identify more proteins and unveils more information about a proteome.

Endoplasmic reticulum mediated signaling in cellular microdomains

PubMed Central

Biwer, Lauren; Isakson, Brant E

2016-01-01

The endoplasmic reticulum (ER) is a prime mediator of cellular signaling due to its functions as an internal cellular store for calcium, as well as a site for synthesis of proteins and lipids. Its peripheral network of sheets and tubules facilitate calcium and lipid signaling, especially in areas of the cell that are more distant to the main cytoplasmic network. Specific membrane proteins shape the peripheral ER architecture and influence the network stability in order to project into restricted spaces. The signaling microdomains are anatomically separate from the cytoplasm as a whole and exhibit localized protein, ion channel and cytoskeletal element expression. Signaling can also occur between the ER and other organelles, such as the Golgi or mitochondria. Lipids made in the ER membrane can be sent to the Golgi via specialized transfer proteins and specific phospholipid synthases are enriched at ER-mitochondria junctions to more efficiently expedite phospholipid transfer. As a hub for protein and lipid synthesis, a store for intracellular calcium [Ca2+]i, and a mediator of cellular stress, the ER is an important cellular organelle. Its ability to organize into tubules and project into restricted spaces allows for discrete and temporal signaling, which is important for cellular physiology and organism homeostasis. PMID:26973141

Protein-protein interaction networks identify targets which rescue the MPP+ cellular model of Parkinson’s disease

NASA Astrophysics Data System (ADS)

Keane, Harriet; Ryan, Brent J.; Jackson, Brendan; Whitmore, Alan; Wade-Martins, Richard

2015-11-01

Neurodegenerative diseases are complex multifactorial disorders characterised by the interplay of many dysregulated physiological processes. As an exemplar, Parkinson’s disease (PD) involves multiple perturbed cellular functions, including mitochondrial dysfunction and autophagic dysregulation in preferentially-sensitive dopamine neurons, a selective pathophysiology recapitulated in vitro using the neurotoxin MPP+. Here we explore a network science approach for the selection of therapeutic protein targets in the cellular MPP+ model. We hypothesised that analysis of protein-protein interaction networks modelling MPP+ toxicity could identify proteins critical for mediating MPP+ toxicity. Analysis of protein-protein interaction networks constructed to model the interplay of mitochondrial dysfunction and autophagic dysregulation (key aspects of MPP+ toxicity) enabled us to identify four proteins predicted to be key for MPP+ toxicity (P62, GABARAP, GBRL1 and GBRL2). Combined, but not individual, knockdown of these proteins increased cellular susceptibility to MPP+ toxicity. Conversely, combined, but not individual, over-expression of the network targets provided rescue of MPP+ toxicity associated with the formation of autophagosome-like structures. We also found that modulation of two distinct proteins in the protein-protein interaction network was necessary and sufficient to mitigate neurotoxicity. Together, these findings validate our network science approach to multi-target identification in complex neurological diseases.

Deregulation of F-box proteins and its consequence on cancer development, progression and metastasis

PubMed Central

Heo, Jinho; Eki, Rebeka; Abbas, Tarek

2015-01-01

F-box proteins are substrate receptors of the SCF (SKP1-Cullin 1-F-box protein) E3 ubiquitin ligase that play important roles in a number of physiological processes and activities. Through their ability to assemble distinct E3 ubiquitin ligases and target key regulators of cellular activities for ubiquitylation and degradation, this versatile group of proteins is able to regulate the abundance of cellular proteins whose deregulated expression or activity contributes to disease. In this review, we describe the important roles of select F-box proteins in regulating cellular activities, the perturbation of which contributes to the initiation and progression of a number of human malignancies. PMID:26432751

The Protein Identifier Cross-Referencing (PICR) service: reconciling protein identifiers across multiple source databases.

PubMed

Côté, Richard G; Jones, Philip; Martens, Lennart; Kerrien, Samuel; Reisinger, Florian; Lin, Quan; Leinonen, Rasko; Apweiler, Rolf; Hermjakob, Henning

2007-10-18

Each major protein database uses its own conventions when assigning protein identifiers. Resolving the various, potentially unstable, identifiers that refer to identical proteins is a major challenge. This is a common problem when attempting to unify datasets that have been annotated with proteins from multiple data sources or querying data providers with one flavour of protein identifiers when the source database uses another. Partial solutions for protein identifier mapping exist but they are limited to specific species or techniques and to a very small number of databases. As a result, we have not found a solution that is generic enough and broad enough in mapping scope to suit our needs. We have created the Protein Identifier Cross-Reference (PICR) service, a web application that provides interactive and programmatic (SOAP and REST) access to a mapping algorithm that uses the UniProt Archive (UniParc) as a data warehouse to offer protein cross-references based on 100% sequence identity to proteins from over 70 distinct source databases loaded into UniParc. Mappings can be limited by source database, taxonomic ID and activity status in the source database. Users can copy/paste or upload files containing protein identifiers or sequences in FASTA format to obtain mappings using the interactive interface. Search results can be viewed in simple or detailed HTML tables or downloaded as comma-separated values (CSV) or Microsoft Excel (XLS) files suitable for use in a local database or a spreadsheet. Alternatively, a SOAP interface is available to integrate PICR functionality in other applications, as is a lightweight REST interface. We offer a publicly available service that can interactively map protein identifiers and protein sequences to the majority of commonly used protein databases. Programmatic access is available through a standards-compliant SOAP interface or a lightweight REST interface. The PICR interface, documentation and code examples are available at http://www.ebi.ac.uk/Tools/picr.

The Protein Identifier Cross-Referencing (PICR) service: reconciling protein identifiers across multiple source databases

PubMed Central

Côté, Richard G; Jones, Philip; Martens, Lennart; Kerrien, Samuel; Reisinger, Florian; Lin, Quan; Leinonen, Rasko; Apweiler, Rolf; Hermjakob, Henning

2007-01-01

Background Each major protein database uses its own conventions when assigning protein identifiers. Resolving the various, potentially unstable, identifiers that refer to identical proteins is a major challenge. This is a common problem when attempting to unify datasets that have been annotated with proteins from multiple data sources or querying data providers with one flavour of protein identifiers when the source database uses another. Partial solutions for protein identifier mapping exist but they are limited to specific species or techniques and to a very small number of databases. As a result, we have not found a solution that is generic enough and broad enough in mapping scope to suit our needs. Results We have created the Protein Identifier Cross-Reference (PICR) service, a web application that provides interactive and programmatic (SOAP and REST) access to a mapping algorithm that uses the UniProt Archive (UniParc) as a data warehouse to offer protein cross-references based on 100% sequence identity to proteins from over 70 distinct source databases loaded into UniParc. Mappings can be limited by source database, taxonomic ID and activity status in the source database. Users can copy/paste or upload files containing protein identifiers or sequences in FASTA format to obtain mappings using the interactive interface. Search results can be viewed in simple or detailed HTML tables or downloaded as comma-separated values (CSV) or Microsoft Excel (XLS) files suitable for use in a local database or a spreadsheet. Alternatively, a SOAP interface is available to integrate PICR functionality in other applications, as is a lightweight REST interface. Conclusion We offer a publicly available service that can interactively map protein identifiers and protein sequences to the majority of commonly used protein databases. Programmatic access is available through a standards-compliant SOAP interface or a lightweight REST interface. The PICR interface, documentation and code examples are available at . PMID:17945017

Identification of Human N-Myristoylated Proteins from Human Complementary DNA Resources by Cell-Free and Cellular Metabolic Labeling Analyses.

PubMed

Takamitsu, Emi; Otsuka, Motoaki; Haebara, Tatsuki; Yano, Manami; Matsuzaki, Kanako; Kobuchi, Hirotsugu; Moriya, Koko; Utsumi, Toshihiko

2015-01-01

To identify physiologically important human N-myristoylated proteins, 90 cDNA clones predicted to encode human N-myristoylated proteins were selected from a human cDNA resource (4,369 Kazusa ORFeome project human cDNA clones) by two bioinformatic N-myristoylation prediction systems, NMT-The MYR Predictor and Myristoylator. After database searches to exclude known human N-myristoylated proteins, 37 cDNA clones were selected as potential human N-myristoylated proteins. The susceptibility of these cDNA clones to protein N-myristoylation was first evaluated using fusion proteins in which the N-terminal ten amino acid residues were fused to an epitope-tagged model protein. Then, protein N-myristoylation of the gene products of full-length cDNAs was evaluated by metabolic labeling experiments both in an insect cell-free protein synthesis system and in transfected human cells. As a result, the products of 13 cDNA clones (FBXL7, PPM1B, SAMM50, PLEKHN, AIFM3, C22orf42, STK32A, FAM131C, DRICH1, MCC1, HID1, P2RX5, STK32B) were found to be human N-myristoylated proteins. Analysis of the role of protein N-myristoylation on the intracellular localization of SAMM50, a mitochondrial outer membrane protein, revealed that protein N-myristoylation was required for proper targeting of SAMM50 to mitochondria. Thus, the strategy used in this study is useful for the identification of physiologically important human N-myristoylated proteins from human cDNA resources.

Identification of Human N-Myristoylated Proteins from Human Complementary DNA Resources by Cell-Free and Cellular Metabolic Labeling Analyses

PubMed Central

Takamitsu, Emi; Otsuka, Motoaki; Haebara, Tatsuki; Yano, Manami; Matsuzaki, Kanako; Kobuchi, Hirotsugu; Moriya, Koko; Utsumi, Toshihiko

2015-01-01

To identify physiologically important human N-myristoylated proteins, 90 cDNA clones predicted to encode human N-myristoylated proteins were selected from a human cDNA resource (4,369 Kazusa ORFeome project human cDNA clones) by two bioinformatic N-myristoylation prediction systems, NMT-The MYR Predictor and Myristoylator. After database searches to exclude known human N-myristoylated proteins, 37 cDNA clones were selected as potential human N-myristoylated proteins. The susceptibility of these cDNA clones to protein N-myristoylation was first evaluated using fusion proteins in which the N-terminal ten amino acid residues were fused to an epitope-tagged model protein. Then, protein N-myristoylation of the gene products of full-length cDNAs was evaluated by metabolic labeling experiments both in an insect cell-free protein synthesis system and in transfected human cells. As a result, the products of 13 cDNA clones (FBXL7, PPM1B, SAMM50, PLEKHN, AIFM3, C22orf42, STK32A, FAM131C, DRICH1, MCC1, HID1, P2RX5, STK32B) were found to be human N-myristoylated proteins. Analysis of the role of protein N-myristoylation on the intracellular localization of SAMM50, a mitochondrial outer membrane protein, revealed that protein N-myristoylation was required for proper targeting of SAMM50 to mitochondria. Thus, the strategy used in this study is useful for the identification of physiologically important human N-myristoylated proteins from human cDNA resources. PMID:26308446

Quantitative imaging with fluorescent biosensors.

PubMed

Okumoto, Sakiko; Jones, Alexander; Frommer, Wolf B

2012-01-01

Molecular activities are highly dynamic and can occur locally in subcellular domains or compartments. Neighboring cells in the same tissue can exist in different states. Therefore, quantitative information on the cellular and subcellular dynamics of ions, signaling molecules, and metabolites is critical for functional understanding of organisms. Mass spectrometry is generally used for monitoring ions and metabolites; however, its temporal and spatial resolution are limited. Fluorescent proteins have revolutionized many areas of biology-e.g., fluorescent proteins can report on gene expression or protein localization in real time-yet promoter-based reporters are often slow to report physiologically relevant changes such as calcium oscillations. Therefore, novel tools are required that can be deployed in specific cells and targeted to subcellular compartments in order to quantify target molecule dynamics directly. We require tools that can measure enzyme activities, protein dynamics, and biophysical processes (e.g., membrane potential or molecular tension) with subcellular resolution. Today, we have an extensive suite of tools at our disposal to address these challenges, including translocation sensors, fluorescence-intensity sensors, and Förster resonance energy transfer sensors. This review summarizes sensor design principles, provides a database of sensors for more than 70 different analytes/processes, and gives examples of applications in quantitative live cell imaging.

UniPROBE, update 2011: expanded content and search tools in the online database of protein-binding microarray data on protein-DNA interactions.

PubMed

Robasky, Kimberly; Bulyk, Martha L

2011-01-01

The Universal PBM Resource for Oligonucleotide-Binding Evaluation (UniPROBE) database is a centralized repository of information on the DNA-binding preferences of proteins as determined by universal protein-binding microarray (PBM) technology. Each entry for a protein (or protein complex) in UniPROBE provides the quantitative preferences for all possible nucleotide sequence variants ('words') of length k ('k-mers'), as well as position weight matrix (PWM) and graphical sequence logo representations of the k-mer data. In this update, we describe >130% expansion of the database content, incorporation of a protein BLAST (blastp) tool for finding protein sequence matches in UniPROBE, the introduction of UniPROBE accession numbers and additional database enhancements. The UniPROBE database is available at http://uniprobe.org.

Modification of an acetone-sodium dodecyl sulfate disruption method for cellular protein extraction from neuropathogenic Clostridium botulinum

USDA-ARS?s Scientific Manuscript database

An acetone-sodium dodecyl sulfate (SDS) disruption method was used for the extraction of cellular proteins from neurotoxigenic Clostridium botulinum. The amount of protein extracted per gram of dry weight and the protein profile as revealed by polyacrylamide gel electrophoresis (PAGE) was comparabl...

Targeting Virus-host Interactions of HIV Replication.

PubMed

Weydert, Caroline; De Rijck, Jan; Christ, Frauke; Debyser, Zeger

2016-01-01

Cellular proteins that are hijacked by HIV in order to complete its replication cycle, form attractive new targets for antiretroviral therapy. In particular, the protein-protein interactions between these cellular proteins (cofactors) and viral proteins are of great interest to develop new therapies. Research efforts have led to the validation of different cofactors and some successes in therapeutic applications. Maraviroc, the first cofactor inhibitor approved for human medicinal use, provided a proof of concept. Furthermore, compounds developed as Integrase-LEDGF/p75 interaction inhibitors (LEDGINs) have advanced to early clinical trials. Other compounds targeting cofactors and cofactor-viral protein interactions are currently under development. Likewise, interactions between cellular restriction factors and their counteracting HIV protein might serve as interesting targets in order to impair HIV replication. In this respect, compounds targeting the Vif-APOBEC3G interaction have been described. In this review, we focus on compounds targeting the Integrase- LEDGF/p75 interaction, the Tat-P-TEFb interaction and the Vif-APOBEC3G interaction. Additionally we give an overview of currently discovered compounds presumably targeting cellular cofactor-HIV protein interactions.

Identification of a cytoplasmic interaction partner of the large regulatory proteins Rep78/Rep68 of adeno-associated virus type 2 (AAV-2)

DOE Office of Scientific and Technical Information (OSTI.GOV)

Weger, Stefan; Hammer, Eva; Goetz, Anne

2007-05-25

Through yeast two-hybrid analysis and coimmunoprecipitation studies, we have identified a novel cellular AAV-2 Rep78/Rep68 interaction partner located predominantly in the cytoplasm. In public databases, it has been assigned as KCTD5, because of a region of high similarity to the cytoplasmic tetramerization domain of voltage-gated potassium channels. Whereas Rep/KCTD5 interaction relied on the region surrounding the Rep nuclear localization signal, nuclear accumulation of Rep was not required. Wildtype Rep78/Rep68 proteins induced the translocation of large portions of KCTD5 into the nucleus pointing to functional interactions both in the cytoplasm and the nucleus. In line with an anticipated functional interference inmore » the cytoplasm, KCTD5 overexpression completely abrogated Rep68-mediated posttranscriptional activation of a HIV-LTR driven luciferase reporter gene. Our study expands the panel of already identified nuclear Rep interaction partners to a cytoplasmic protein, which raises the awareness that important steps in the AAV life cycle may be regulated in this compartment.« less

Identification of immunity-related genes in the larvae of Protaetia brevitarsis seulensis (Coleoptera: Cetoniidae) by a next-generation sequencing-based transcriptome analysis.

PubMed

Bang, Kyeongrin; Hwang, Sejung; Lee, Jiae; Cho, Saeyoull

2015-01-01

To identify immune-related genes in the larvae of white-spotted flower chafers, next-generation sequencing was conducted with an Illumina HiSeq2000, resulting in 100 million cDNA reads with sequence information from over 10 billion base pairs (bp) and >50× transcriptome coverage. A subset of 77,336 contigs was created, and ∼35,532 sequences matched entries against the NCBI nonredundant database (cutoff, e < 10(-5)). Statistical analysis was performed on the 35,532 contigs. For profiling of the immune response, samples were analyzed by aligning 42 base sequence tags to the de novo reference assembly, comparing levels in immunized larvae to control levels of expression. Of the differentially expressed genes, 3,440 transcripts were upregulated and 3,590 transcripts were downregulated. Many of these genes were confirmed as immune-related genes such as pattern recognition proteins, immune-related signal transduction proteins, antimicrobial peptides, and cellular response proteins, by comparison to published data. © The Author 2015. Published by Oxford University Press on behalf of the Entomological Society of America.

Aberrant localization of lamin B receptor (LBR) in cellular senescence in human cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Arai, Rumi; En, Atsuki; Ukekawa, Ryo

2016-05-13

5-Bromodeoxyuridine (BrdU), a thymidine analogue, induces cellular senescence in mammalian cells. BrdU induces cellular senescence probably through the regulation of chromatin because BrdU destabilizes or disrupts nucleosome positioning and decondenses heterochromatin. Since heterochromatin is tethered to the nuclear periphery through the interaction with the nuclear envelope proteins, we examined the localization of the several nuclear envelope proteins such as lamins, lamin-interacting proteins, nuclear pore complex proteins, and nuclear transport proteins in senescent cells. We have shown here that lamin B receptor (LBR) showed a change in localization in both BrdU-induced and replicative senescent cells.

Determining the sub-cellular localization of proteins within Caenorhabditis elegans body wall muscle.

PubMed

Meissner, Barbara; Rogalski, Teresa; Viveiros, Ryan; Warner, Adam; Plastino, Lorena; Lorch, Adam; Granger, Laure; Segalat, Laurent; Moerman, Donald G

2011-01-01

Determining the sub-cellular localization of a protein within a cell is often an essential step towards understanding its function. In Caenorhabditis elegans, the relatively large size of the body wall muscle cells and the exquisite organization of their sarcomeres offer an opportunity to identify the precise position of proteins within cell substructures. Our goal in this study is to generate a comprehensive "localizome" for C. elegans body wall muscle by GFP-tagging proteins expressed in muscle and determining their location within the cell. For this project, we focused on proteins that we know are expressed in muscle and are orthologs or at least homologs of human proteins. To date we have analyzed the expression of about 227 GFP-tagged proteins that show localized expression in the body wall muscle of this nematode (e.g. dense bodies, M-lines, myofilaments, mitochondria, cell membrane, nucleus or nucleolus). For most proteins analyzed in this study no prior data on sub-cellular localization was available. In addition to discrete sub-cellular localization we observe overlapping patterns of localization including the presence of a protein in the dense body and the nucleus, or the dense body and the M-lines. In total we discern more than 14 sub-cellular localization patterns within nematode body wall muscle. The localization of this large set of proteins within a muscle cell will serve as an invaluable resource in our investigation of muscle sarcomere assembly and function.

A Decade of Boon or Burden: What Has the CHIP Ever Done for Cellular Protein Quality Control Mechanism Implicated in Neurodegeneration and Aging?

PubMed Central

Joshi, Vibhuti; Amanullah, Ayeman; Upadhyay, Arun; Mishra, Ribhav; Kumar, Amit; Mishra, Amit

2016-01-01

Cells regularly synthesize new proteins to replace old and abnormal proteins for normal cellular functions. Two significant protein quality control pathways inside the cellular milieu are ubiquitin proteasome system (UPS) and autophagy. Autophagy is known for bulk clearance of cytoplasmic aggregated proteins, whereas the specificity of protein degradation by UPS comes from E3 ubiquitin ligases. Few E3 ubiquitin ligases, like C-terminus of Hsc70-interacting protein (CHIP) not only take part in protein quality control pathways, but also plays a key regulatory role in other cellular processes like signaling, development, DNA damage repair, immunity and aging. CHIP targets misfolded proteins for their degradation through proteasome, as well as autophagy; simultaneously, with the help of chaperones, it also regulates folding attempts for misfolded proteins. The broad range of CHIP substrates and their associations with multiple pathologies make it a key molecule to work upon and focus for future therapeutic interventions. E3 ubiquitin ligase CHIP interacts and degrades many protein inclusions formed in neurodegenerative diseases. The presence of CHIP at various nodes of cellular protein-protein interaction network presents this molecule as a potential candidate for further research. In this review, we have explored a wide range of functionality of CHIP inside cells by a detailed presentation of its co-chaperone, E3 and E4 enzyme like functions, with central focus on its protein quality control roles in neurodegenerative diseases. We have also raised many unexplored but expected fundamental questions regarding CHIP functions, which generate hopes for its future applications in research, as well as drug discovery. PMID:27757073

Combined Mass Spectrometry Imaging and Top-down Microproteomics Reveals Evidence of a Hidden Proteome in Ovarian Cancer.

PubMed

Delcourt, Vivian; Franck, Julien; Leblanc, Eric; Narducci, Fabrice; Robin, Yves-Marie; Gimeno, Jean-Pascal; Quanico, Jusal; Wisztorski, Maxence; Kobeissy, Firas; Jacques, Jean-François; Roucou, Xavier; Salzet, Michel; Fournier, Isabelle

2017-07-01

Recently, it was demonstrated that proteins can be translated from alternative open reading frames (altORFs), increasing the size of the actual proteome. Top-down mass spectrometry-based proteomics allows the identification of intact proteins containing post-translational modifications (PTMs) as well as truncated forms translated from reference ORFs or altORFs. Top-down tissue microproteomics was applied on benign, tumor and necrotic-fibrotic regions of serous ovarian cancer biopsies, identifying proteins exhibiting region-specific cellular localization and PTMs. The regions of interest (ROIs) were determined by MALDI mass spectrometry imaging and spatial segmentation. Analysis with a customized protein sequence database containing reference and alternative proteins (altprots) identified 15 altprots, including alternative G protein nucleolar 1 (AltGNL1) found in the tumor, and translated from an altORF nested within the GNL1 canonical coding sequence. Co-expression of GNL1 and altGNL1 was validated by transfection in HEK293 and HeLa cells with an expression plasmid containing a GNL1-FLAG (V5) construct. Western blot and immunofluorescence experiments confirmed constitutive co-expression of altGNL1-V5 with GNL1-FLAG. Taken together, our approach provides means to evaluate protein changes in the case of serous ovarian cancer, allowing the detection of potential markers that have never been considered. Copyright © 2017 The Author(s). Published by Elsevier B.V. All rights reserved.

NRfamPred: a proteome-scale two level method for prediction of nuclear receptor proteins and their sub-families.

PubMed

Kumar, Ravindra; Kumari, Bandana; Srivastava, Abhishikha; Kumar, Manish

2014-10-29

Nuclear receptor proteins (NRP) are transcription factor that regulate many vital cellular processes in animal cells. NRPs form a super-family of phylogenetically related proteins and divided into different sub-families on the basis of ligand characteristics and their functions. In the post-genomic era, when new proteins are being added to the database in a high-throughput mode, it becomes imperative to identify new NRPs using information from amino acid sequence alone. In this study we report a SVM based two level prediction systems, NRfamPred, using dipeptide composition of proteins as input. At the 1st level, NRfamPred screens whether the query protein is NRP or non-NRP; if the query protein belongs to NRP class, prediction moves to 2nd level and predicts the sub-family. Using leave-one-out cross-validation, we were able to achieve an overall accuracy of 97.88% at the 1st level and an overall accuracy of 98.11% at the 2nd level with dipeptide composition. Benchmarking on independent datasets showed that NRfamPred had comparable accuracy to other existing methods, developed on the same dataset. Our method predicted the existence of 76 NRPs in the human proteome, out of which 14 are novel NRPs. NRfamPred also predicted the sub-families of these 14 NRPs.

A gene network bioinformatics analysis for pemphigoid autoimmune blistering diseases.

PubMed

Barone, Antonio; Toti, Paolo; Giuca, Maria Rita; Derchi, Giacomo; Covani, Ugo

2015-07-01

In this theoretical study, a text mining search and clustering analysis of data related to genes potentially involved in human pemphigoid autoimmune blistering diseases (PAIBD) was performed using web tools to create a gene/protein interaction network. The Search Tool for the Retrieval of Interacting Genes/Proteins (STRING) database was employed to identify a final set of PAIBD-involved genes and to calculate the overall significant interactions among genes: for each gene, the weighted number of links, or WNL, was registered and a clustering procedure was performed using the WNL analysis. Genes were ranked in class (leader, B, C, D and so on, up to orphans). An ontological analysis was performed for the set of 'leader' genes. Using the above-mentioned data network, 115 genes represented the final set; leader genes numbered 7 (intercellular adhesion molecule 1 (ICAM-1), interferon gamma (IFNG), interleukin (IL)-2, IL-4, IL-6, IL-8 and tumour necrosis factor (TNF)), class B genes were 13, whereas the orphans were 24. The ontological analysis attested that the molecular action was focused on extracellular space and cell surface, whereas the activation and regulation of the immunity system was widely involved. Despite the limited knowledge of the present pathologic phenomenon, attested by the presence of 24 genes revealing no protein-protein direct or indirect interactions, the network showed significant pathways gathered in several subgroups: cellular components, molecular functions, biological processes and the pathologic phenomenon obtained from the Kyoto Encyclopaedia of Genes and Genomes (KEGG) database. The molecular basis for PAIBD was summarised and expanded, which will perhaps give researchers promising directions for the identification of new therapeutic targets.

FunGene: the functional gene pipeline and repository.

PubMed

Fish, Jordan A; Chai, Benli; Wang, Qiong; Sun, Yanni; Brown, C Titus; Tiedje, James M; Cole, James R

2013-01-01

Ribosomal RNA genes have become the standard molecular markers for microbial community analysis for good reasons, including universal occurrence in cellular organisms, availability of large databases, and ease of rRNA gene region amplification and analysis. As markers, however, rRNA genes have some significant limitations. The rRNA genes are often present in multiple copies, unlike most protein-coding genes. The slow rate of change in rRNA genes means that multiple species sometimes share identical 16S rRNA gene sequences, while many more species share identical sequences in the short 16S rRNA regions commonly analyzed. In addition, the genes involved in many important processes are not distributed in a phylogenetically coherent manner, potentially due to gene loss or horizontal gene transfer. While rRNA genes remain the most commonly used markers, key genes in ecologically important pathways, e.g., those involved in carbon and nitrogen cycling, can provide important insights into community composition and function not obtainable through rRNA analysis. However, working with ecofunctional gene data requires some tools beyond those required for rRNA analysis. To address this, our Functional Gene Pipeline and Repository (FunGene; http://fungene.cme.msu.edu/) offers databases of many common ecofunctional genes and proteins, as well as integrated tools that allow researchers to browse these collections and choose subsets for further analysis, build phylogenetic trees, test primers and probes for coverage, and download aligned sequences. Additional FunGene tools are specialized to process coding gene amplicon data. For example, FrameBot produces frameshift-corrected protein and DNA sequences from raw reads while finding the most closely related protein reference sequence. These tools can help provide better insight into microbial communities by directly studying key genes involved in important ecological processes.

MIPS: a database for protein sequences, homology data and yeast genome information.

PubMed Central

Mewes, H W; Albermann, K; Heumann, K; Liebl, S; Pfeiffer, F

1997-01-01

The MIPS group (Martinsried Institute for Protein Sequences) at the Max-Planck-Institute for Biochemistry, Martinsried near Munich, Germany, collects, processes and distributes protein sequence data within the framework of the tripartite association of the PIR-International Protein Sequence Database (,). MIPS contributes nearly 50% of the data input to the PIR-International Protein Sequence Database. The database is distributed on CD-ROM together with PATCHX, an exhaustive supplement of unique, unverified protein sequences from external sources compiled by MIPS. Through its WWW server (http://www.mips.biochem.mpg.de/ ) MIPS permits internet access to sequence databases, homology data and to yeast genome information. (i) Sequence similarity results from the FASTA program () are stored in the FASTA database for all proteins from PIR-International and PATCHX. The database is dynamically maintained and permits instant access to FASTA results. (ii) Starting with FASTA database queries, proteins have been classified into families and superfamilies (PROT-FAM). (iii) The HPT (hashed position tree) data structure () developed at MIPS is a new approach for rapid sequence and pattern searching. (iv) MIPS provides access to the sequence and annotation of the complete yeast genome (), the functional classification of yeast genes (FunCat) and its graphical display, the 'Genome Browser' (). A CD-ROM based on the JAVA programming language providing dynamic interactive access to the yeast genome and the related protein sequences has been compiled and is available on request. PMID:9016498

CellNetVis: a web tool for visualization of biological networks using force-directed layout constrained by cellular components.

PubMed

Heberle, Henry; Carazzolle, Marcelo Falsarella; Telles, Guilherme P; Meirelles, Gabriela Vaz; Minghim, Rosane

2017-09-13

The advent of "omics" science has brought new perspectives in contemporary biology through the high-throughput analyses of molecular interactions, providing new clues in protein/gene function and in the organization of biological pathways. Biomolecular interaction networks, or graphs, are simple abstract representations where the components of a cell (e.g. proteins, metabolites etc.) are represented by nodes and their interactions are represented by edges. An appropriate visualization of data is crucial for understanding such networks, since pathways are related to functions that occur in specific regions of the cell. The force-directed layout is an important and widely used technique to draw networks according to their topologies. Placing the networks into cellular compartments helps to quickly identify where network elements are located and, more specifically, concentrated. Currently, only a few tools provide the capability of visually organizing networks by cellular compartments. Most of them cannot handle large and dense networks. Even for small networks with hundreds of nodes the available tools are not able to reposition the network while the user is interacting, limiting the visual exploration capability. Here we propose CellNetVis, a web tool to easily display biological networks in a cell diagram employing a constrained force-directed layout algorithm. The tool is freely available and open-source. It was originally designed for networks generated by the Integrated Interactome System and can be used with networks from others databases, like InnateDB. CellNetVis has demonstrated to be applicable for dynamic investigation of complex networks over a consistent representation of a cell on the Web, with capabilities not matched elsewhere.

NPIDB: Nucleic acid-Protein Interaction DataBase.

PubMed

Kirsanov, Dmitry D; Zanegina, Olga N; Aksianov, Evgeniy A; Spirin, Sergei A; Karyagina, Anna S; Alexeevski, Andrei V

2013-01-01

The Nucleic acid-Protein Interaction DataBase (http://npidb.belozersky.msu.ru/) contains information derived from structures of DNA-protein and RNA-protein complexes extracted from the Protein Data Bank (3846 complexes in October 2012). It provides a web interface and a set of tools for extracting biologically meaningful characteristics of nucleoprotein complexes. The content of the database is updated weekly. The current version of the Nucleic acid-Protein Interaction DataBase is an upgrade of the version published in 2007. The improvements include a new web interface, new tools for calculation of intermolecular interactions, a classification of SCOP families that contains DNA-binding protein domains and data on conserved water molecules on the DNA-protein interface.

De Novo Transcriptome Analysis of Allium cepa L. (Onion) Bulb to Identify Allergens and Epitopes.

PubMed

Rajkumar, Hemalatha; Ramagoni, Ramesh Kumar; Anchoju, Vijayendra Chary; Vankudavath, Raju Naik; Syed, Arshi Uz Zaman

2015-01-01

Allium cepa (onion) is a diploid plant with one of the largest nuclear genomes among all diploids. Onion is an example of an under-researched crop which has a complex heterozygous genome. There are no allergenic proteins and genomic data available for onions. This study was conducted to establish a transcriptome catalogue of onion bulb that will enable us to study onion related genes involved in medicinal use and allergies. Transcriptome dataset generated from onion bulb using the Illumina HiSeq 2000 technology showed a total of 99,074,309 high quality raw reads (~20 Gb). Based on sequence homology onion genes were categorized into 49 different functional groups. Most of the genes however, were classified under 'unknown' in all three gene ontology categories. Of the categorized genes, 61.2% showed metabolic functions followed by cellular components such as binding, cellular processes; catalytic activity and cell part. With BLASTx top hit analysis, a total of 2,511 homologous allergenic sequences were found, which had 37-100% similarity with 46 different types of allergens existing in the database. From the 46 contigs or allergens, 521 B-cell linear epitopes were identified using BepiPred linear epitope prediction tool. This is the first comprehensive insight into the transcriptome of onion bulb tissue using the NGS technology, which can be used to map IgE epitopes and prediction of structures and functions of various proteins.

Unique spatial and cellular expression patterns of Hoxa5, Hoxb4 and Hoxb6 proteins in normal developing murine lung are modified in pulmonary hypoplasia

PubMed Central

Volpe, MaryAnn Vitoria; Wang, Karen Ting Wai; Nielsen, Heber Carl; Chinoy, Mala Romeshchandra

2009-01-01

Background Hox transcription factors modulate signaling pathways controlling organ morphogenesis and maintain cell fate and differentiation in adults. Retinoid signaling, key in regulating Hox expression, is altered in pulmonary hypoplasia. Information on pattern-specific expression of Hox proteins in normal lung development and in pulmonary hypoplasia is minimal. Our objective was to determine how pulmonary hypoplasia alters temporal, spatial and cellular expression of Hoxa5, Hoxb4 and Hoxb6 proteins compared to normal lung development. Methods Temporal, spatial and cellular Hoxa5, Hoxb4 and Hoxb6 expression was studied in normal (untreated) and nitrofen-induced hypoplastic (NT-PH) lungs from gestational day 13.5, 16, 19 fetuses and neonates using western blot and immunohistochemistry. Results Modification of protein levels and spatial and cellular Hox expression patterns in NT-PH lungs was consistent with delayed lung development. Distinct protein isoforms were detected for each Hox protein. Expression levels of the Hoxa5 and Hoxb6 isoforms changed with development and further in NT-PH lungs. Compared to normal lungs, Gd19 and neonatal NT-PH lungs had decreased Hoxb6 and increased Hoxa5 and Hoxb4. Hoxa5 cellular localization changed from mesenchyme to epithelia earlier in normal lungs. Hoxb4 was expressed in mesenchyme and epithelial cells throughout development. Hoxb6 remained mainly in mesenchymal cells around distal airways. Conclusions Unique spatial and cellular expression of Hoxa5, Hoxb4 and Hoxb6 participates in branching morphogenesis and terminal sac formation. Altered Hox protein temporal and cellular balance of expression either contributes to pulmonary hypoplasia or functions as a compensatory mechanism attempting to correct abnormal lung development and maturation in this condition. PMID:18553509

The PMDB Protein Model Database

PubMed Central

Castrignanò, Tiziana; De Meo, Paolo D'Onorio; Cozzetto, Domenico; Talamo, Ivano Giuseppe; Tramontano, Anna

2006-01-01

The Protein Model Database (PMDB) is a public resource aimed at storing manually built 3D models of proteins. The database is designed to provide access to models published in the scientific literature, together with validating experimental data. It is a relational database and it currently contains >74 000 models for ∼240 proteins. The system is accessible at and allows predictors to submit models along with related supporting evidence and users to download them through a simple and intuitive interface. Users can navigate in the database and retrieve models referring to the same target protein or to different regions of the same protein. Each model is assigned a unique identifier that allows interested users to directly access the data. PMID:16381873

PDB_TM: selection and membrane localization of transmembrane proteins in the protein data bank.

PubMed

Tusnády, Gábor E; Dosztányi, Zsuzsanna; Simon, István

2005-01-01

PDB_TM is a database for transmembrane proteins with known structures. It aims to collect all transmembrane proteins that are deposited in the protein structure database (PDB) and to determine their membrane-spanning regions. These assignments are based on the TMDET algorithm, which uses only structural information to locate the most likely position of the lipid bilayer and to distinguish between transmembrane and globular proteins. This algorithm was applied to all PDB entries and the results were collected in the PDB_TM database. By using TMDET algorithm, the PDB_TM database can be automatically updated every week, keeping it synchronized with the latest PDB updates. The PDB_TM database is available at http://www.enzim.hu/PDB_TM.

Protein arginine methylation: Cellular functions and methods of analysis.

PubMed

Pahlich, Steffen; Zakaryan, Rouzanna P; Gehring, Heinz

2006-12-01

During the last few years, new members of the growing family of protein arginine methyltransferases (PRMTs) have been identified and the role of arginine methylation in manifold cellular processes like signaling, RNA processing, transcription, and subcellular transport has been extensively investigated. In this review, we describe recent methods and findings that have yielded new insights into the cellular functions of arginine-methylated proteins, and we evaluate the currently used procedures for the detection and analysis of arginine methylation.

The Presence of ADP-Ribosylated Fe Protein of Nitrogenase in Rhodobacter capsulatus Is Correlated with Cellular Nitrogen Status

PubMed Central

Yakunin, Alexander F.; Laurinavichene, Tatyana V.; Tsygankov, Anatoly A.; Hallenbeck, Patrick C.

1999-01-01

The photosynthetic bacterium Rhodobacter capsulatus has been shown to regulate its nitrogenase by covalent modification via the reversible ADP-ribosylation of Fe protein in response to darkness or the addition of external NH4+. Here we demonstrate the presence of ADP-ribosylated Fe protein under a variety of steady-state growth conditions. We examined the modification of Fe protein and nitrogenase activity under three different growth conditions that establish different levels of cellular nitrogen: batch growth with limiting NH4+, where the nitrogen status is externally controlled; batch growth on relatively poor nitrogen sources, where the nitrogen status is internally controlled by assimilatory processes; and continuous culture. When cultures were grown to stationary phase with different limiting concentrations of NH4+, the ADP-ribosylation state of Fe protein was found to correlate with cellular nitrogen status. Additionally, actively growing cultures (grown with N2 or glutamate), which had an intermediate cellular nitrogen status, contained a portion of their Fe protein in the modified state. The correlation between cellular nitrogen status and ADP-ribosylation state was corroborated with continuous cultures grown under various degrees of nitrogen limitation. These results show that in R. capsulatus the modification system that ADP-ribosylates nitrogenase in the short term in response to abrupt changes in the environment is also capable of modifying nitrogenase in accordance with long-term cellular conditions. PMID:10094674

Toward a systems-level view of dynamic phosphorylation networks

PubMed Central

Newman, Robert H.; Zhang, Jin; Zhu, Heng

2014-01-01

To better understand how cells sense and respond to their environment, it is important to understand the organization and regulation of the phosphorylation networks that underlie most cellular signal transduction pathways. These networks, which are composed of protein kinases, protein phosphatases and their respective cellular targets, are highly dynamic. Importantly, to achieve signaling specificity, phosphorylation networks must be regulated at several levels, including at the level of protein expression, substrate recognition, and spatiotemporal modulation of enzymatic activity. Here, we briefly summarize some of the traditional methods used to study the phosphorylation status of cellular proteins before focusing our attention on several recent technological advances, such as protein microarrays, quantitative mass spectrometry, and genetically-targetable fluorescent biosensors, that are offering new insights into the organization and regulation of cellular phosphorylation networks. Together, these approaches promise to lead to a systems-level view of dynamic phosphorylation networks. PMID:25177341

Cellular proteostasis: degradation of misfolded proteins by lysosomes

PubMed Central

Jackson, Matthew P.

2016-01-01

Proteostasis refers to the regulation of the cellular concentration, folding, interactions and localization of each of the proteins that comprise the proteome. One essential element of proteostasis is the disposal of misfolded proteins by the cellular pathways of protein degradation. Lysosomes are an important site for the degradation of misfolded proteins, which are trafficked to this organelle by the pathways of macroautophagy, chaperone-mediated autophagy and endocytosis. Conversely, amyloid diseases represent a failure in proteostasis, in which proteins misfold, forming amyloid deposits that are not degraded effectively by cells. Amyloid may then exacerbate this failure by disrupting autophagy and lysosomal proteolysis. However, targeting the pathways that regulate autophagy and the biogenesis of lysosomes may present approaches that can rescue cells from the deleterious effects of amyloidogenic proteins. PMID:27744333

Genome-wide survey of DNA-binding proteins in Arabidopsis thaliana: analysis of distribution and functions.

PubMed

Malhotra, Sony; Sowdhamini, Ramanathan

2013-08-01

The interaction of proteins with their respective DNA targets is known to control many high-fidelity cellular processes. Performing a comprehensive survey of the sequenced genomes for DNA-binding proteins (DBPs) will help in understanding their distribution and the associated functions in a particular genome. Availability of fully sequenced genome of Arabidopsis thaliana enables the review of distribution of DBPs in this model plant genome. We used profiles of both structure and sequence-based DNA-binding families, derived from PDB and PFam databases, to perform the survey. This resulted in 4471 proteins, identified as DNA-binding in Arabidopsis genome, which are distributed across 300 different PFam families. Apart from several plant-specific DNA-binding families, certain RING fingers and leucine zippers also had high representation. Our search protocol helped to assign DNA-binding property to several proteins that were previously marked as unknown, putative or hypothetical in function. The distribution of Arabidopsis genes having a role in plant DNA repair were particularly studied and noted for their functional mapping. The functions observed to be overrepresented in the plant genome harbour DNA-3-methyladenine glycosylase activity, alkylbase DNA N-glycosylase activity and DNA-(apurinic or apyrimidinic site) lyase activity, suggesting their role in specialized functions such as gene regulation and DNA repair.

Phylogenetic and bioinformatic analysis of gap junction-related proteins, innexins, pannexins and connexins.

PubMed

Fushiki, Daisuke; Hamada, Yasuo; Yoshimura, Ryoichi; Endo, Yasuhisa

2010-04-01

All multi-cellular animals, including hydra, insects and vertebrates, develop gap junctions, which communicate directly with neighboring cells. Gap junctions consist of protein families called connexins in vertebrates and innexins in invertebrates. Connexins and innexins have no homology in their amino acid sequence, but both are thought to have some similar characteristics, such as a tetra-membrane-spanning structure, formation of a channel by hexamer, and transmission of small molecules (e.g. ions) to neighboring cells. Pannexins were recently identified as a homolog of innexins in vertebrate genomes. Although pannexins are thought to share the function of intercellular communication with connexins and innexins, there is little information about the relationship among these three protein families of gap junctions. We phylgenetically and bioinformatically examined these protein families and other tetra-membrane-spanning proteins using a database and three analytical softwares. The clades formed by pannexin families do not belong to the species classification but do to paralogs of each member of pannexins. Amino acid sequences of pannexins are closely related to those of innexins but less to those of connexins. These data suggest that innexins and pannexins have a common origin, but the relationship between innexins/pannexins and connexins is as slight as that of other tetra-membrane-spanning members.

Protein Corona Analysis of Silver Nanoparticles Links to Their Cellular Effects.

PubMed

Juling, Sabine; Niedzwiecka, Alicia; Böhmert, Linda; Lichtenstein, Dajana; Selve, Sören; Braeuning, Albert; Thünemann, Andreas F; Krause, Eberhard; Lampen, Alfonso

2017-11-03

The breadth of applications of nanoparticles and the access to food-associated consumer products containing nanosized materials lead to oral human exposure to such particles. In biological fluids nanoparticles dynamically interact with biomolecules and form a protein corona. Knowledge about the protein corona is of great interest for understanding the molecular effects of particles as well as their fate inside the human body. We used a mass spectrometry-based toxicoproteomics approach to elucidate mechanisms of toxicity of silver nanoparticles and to comprehensively characterize the protein corona formed around silver nanoparticles in Caco-2 human intestinal epithelial cells. Results were compared with respect to the cellular function of proteins either affected by exposure to nanoparticles or present in the protein corona. A transcriptomic data set was included in the analyses in order to obtain a combined multiomics view of nanoparticle-affected cellular processes. A relationship between corona proteins and the proteomic or transcriptomic responses was revealed, showing that differentially regulated proteins or transcripts were engaged in the same cellular signaling pathways. Protein corona analyses of nanoparticles in cells might therefore help in obtaining information about the molecular consequences of nanoparticle treatment.

Protein Analysis of Purified Respiratory Syncytial Virus Particles Reveals an Important Role for Heat Shock Protein 90 in Virus Particle Assembly*

PubMed Central

Radhakrishnan, Anuradha; Yeo, Dawn; Brown, Gaie; Myaing, Myint Zu; Iyer, Laxmi Ravi; Fleck, Roland; Tan, Boon-Huan; Aitken, Jim; Sanmun, Duangmanee; Tang, Kai; Yarwood, Andy; Brink, Jacob; Sugrue, Richard J.

2010-01-01

In this study, we used imaging and proteomics to identify the presence of virus-associated cellular proteins that may play a role in respiratory syncytial virus (RSV) maturation. Fluorescence microscopy of virus-infected cells revealed the presence of virus-induced cytoplasmic inclusion bodies and mature virus particles, the latter appearing as virus filaments. In situ electron tomography suggested that the virus filaments were complex structures that were able to package multiple copies of the virus genome. The virus particles were purified, and the protein content was analyzed by one-dimensional nano-LC MS/MS. In addition to all the major virus structural proteins, 25 cellular proteins were also detected, including proteins associated with the cortical actin network, energy pathways, and heat shock proteins (HSP70, HSC70, and HSP90). Representative actin-associated proteins, HSC70, and HSP90 were selected for further biological validation. The presence of β-actin, filamin-1, cofilin-1, HSC70, and HSP90 in the virus preparation was confirmed by immunoblotting using relevant antibodies. Immunofluorescence microscopy of infected cells stained with antibodies against relevant virus and cellular proteins confirmed the presence of these cellular proteins in the virus filaments and inclusion bodies. The relevance of HSP90 to virus infection was examined using the specific inhibitors 17-N-Allylamino-17-demethoxygeldanamycin. Although virus protein expression was largely unaffected by these drugs, we noted that the formation of virus particles was inhibited, and virus transmission was impaired, suggesting an important role for HSP90 in virus maturation. This study highlights the utility of proteomics in facilitating both our understanding of the role that cellular proteins play during RSV maturation and, by extrapolation, the identification of new potential targets for antiviral therapy. PMID:20530633

Architecture of the human interactome defines protein communities and disease networks

PubMed Central

Huttlin, Edward L.; Bruckner, Raphael J.; Paulo, Joao A.; Cannon, Joe R.; Ting, Lily; Baltier, Kurt; Colby, Greg; Gebreab, Fana; Gygi, Melanie P.; Parzen, Hannah; Szpyt, John; Tam, Stanley; Zarraga, Gabriela; Pontano-Vaites, Laura; Swarup, Sharan; White, Anne E.; Schweppe, Devin K.; Rad, Ramin; Erickson, Brian K.; Obar, Robert A.; Guruharsha, K.G.; Li, Kejie; Artavanis-Tsakonas, Spyros; Gygi, Steven P.; Harper, J. Wade

2017-01-01

The physiology of a cell can be viewed as the product of thousands of proteins acting in concert to shape the cellular response. Coordination is achieved in part through networks of protein-protein interactions that assemble functionally related proteins into complexes, organelles, and signal transduction pathways. Understanding the architecture of the human proteome has the potential to inform cellular, structural, and evolutionary mechanisms and is critical to elucidation of how genome variation contributes to disease1–3. Here, we present BioPlex 2.0 (Biophysical Interactions of ORFEOME-derived complexes), which employs robust affinity purification-mass spectrometry (AP-MS) methodology4 to elucidate protein interaction networks and co-complexes nucleated by more than 25% of protein coding genes from the human genome, and constitutes the largest such network to date. With >56,000 candidate interactions, BioPlex 2.0 contains >29,000 previously unknown co-associations and provides functional insights into hundreds of poorly characterized proteins while enhancing network-based analyses of domain associations, subcellular localization, and co-complex formation. Unsupervised Markov clustering (MCL)5 of interacting proteins identified more than 1300 protein communities representing diverse cellular activities. Genes essential for cell fitness6,7 are enriched within 53 communities representing central cellular functions. Moreover, we identified 442 communities associated with more than 2000 disease annotations, placing numerous candidate disease genes into a cellular framework. BioPlex 2.0 exceeds previous experimentally derived interaction networks in depth and breadth, and will be a valuable resource for exploring the biology of incompletely characterized proteins and for elucidating larger-scale patterns of proteome organization. PMID:28514442

Evolutionary Conservation and Emerging Functional Diversity of the Cytosolic Hsp70:J Protein Chaperone Network of Arabidopsis thaliana.

PubMed

Verma, Amit K; Diwan, Danish; Raut, Sandeep; Dobriyal, Neha; Brown, Rebecca E; Gowda, Vinita; Hines, Justin K; Sahi, Chandan

2017-06-07

Heat shock proteins of 70 kDa (Hsp70s) partner with structurally diverse Hsp40s (J proteins), generating distinct chaperone networks in various cellular compartments that perform myriad housekeeping and stress-associated functions in all organisms. Plants, being sessile, need to constantly maintain their cellular proteostasis in response to external environmental cues. In these situations, the Hsp70:J protein machines may play an important role in fine-tuning cellular protein quality control. Although ubiquitous, the functional specificity and complexity of the plant Hsp70:J protein network has not been studied. Here, we analyzed the J protein network in the cytosol of Arabidopsis thaliana and, using yeast genetics, show that the functional specificities of most plant J proteins in fundamental chaperone functions are conserved across long evolutionary timescales. Detailed phylogenetic and functional analysis revealed that increased number, regulatory differences, and neofunctionalization in J proteins together contribute to the emerging functional diversity and complexity in the Hsp70:J protein network in higher plants. Based on the data presented, we propose that higher plants have orchestrated their "chaperome," especially their J protein complement, according to their specialized cellular and physiological stipulations. Copyright © 2017 Verma et al.

Cellular automata and its applications in protein bioinformatics.

PubMed

Xiao, Xuan; Wang, Pu; Chou, Kuo-Chen

2011-09-01

With the explosion of protein sequences generated in the postgenomic era, it is highly desirable to develop high-throughput tools for rapidly and reliably identifying various attributes of uncharacterized proteins based on their sequence information alone. The knowledge thus obtained can help us timely utilize these newly found protein sequences for both basic research and drug discovery. Many bioinformatics tools have been developed by means of machine learning methods. This review is focused on the applications of a new kind of science (cellular automata) in protein bioinformatics. A cellular automaton (CA) is an open, flexible and discrete dynamic model that holds enormous potentials in modeling complex systems, in spite of the simplicity of the model itself. Researchers, scientists and practitioners from different fields have utilized cellular automata for visualizing protein sequences, investigating their evolution processes, and predicting their various attributes. Owing to its impressive power, intuitiveness and relative simplicity, the CA approach has great potential for use as a tool for bioinformatics.

Proteomic analysis of the gamma human papillomavirus type 197 E6 and E7 associated cellular proteins

PubMed Central

Grace, Miranda; Munger, Karl

2016-01-01

Gamma HPV197 was the most frequently identified HPV when human skin cancer specimens were analyzed by deep sequencing. To gain insight into the biological activities of HPV197, we investigated the cellular interactomes of HPV197 E6 and E7. HPV197 E6 protein interacts with a broad spectrum of cellular LXXLL domain proteins, including UBE3A and MAML1. HPV197 E6 also binds and inhibits the TP53 tumor suppressor and interacts with the CCR4-NOT ubiquitin ligase and deadenylation complex. Despite lacking a canonical retinoblastoma (RB1) tumor suppressor binding site, HPV197 E7 binds RB1 and activates E2F transcription. Hence, HPV197 E6 and E7 proteins interact with a similar set of cellular proteins as E6 and E7 proteins encoded by HPVs that have been linked to human carcinogenesis and/or have transforming activities in vitro. PMID:27771561

A Quantitative Microscopy Technique for Determining the Number of Specific Proteins in Cellular Compartments

PubMed Central

Mutch, Sarah A.; Gadd, Jennifer C.; Fujimoto, Bryant S.; Kensel-Hammes, Patricia; Schiro, Perry G.; Bajjalieh, Sandra M.; Chiu, Daniel T.

2013-01-01

This protocol describes a method to determine both the average number and variance of proteins in the few to tens of copies in isolated cellular compartments, such as organelles and protein complexes. Other currently available protein quantification techniques either provide an average number but lack information on the variance or are not suitable for reliably counting proteins present in the few to tens of copies. This protocol entails labeling the cellular compartment with fluorescent primary-secondary antibody complexes, TIRF (total internal reflection fluorescence) microscopy imaging of the cellular compartment, digital image analysis, and deconvolution of the fluorescence intensity data. A minimum of 2.5 days is required to complete the labeling, imaging, and analysis of a set of samples. As an illustrative example, we describe in detail the procedure used to determine the copy number of proteins in synaptic vesicles. The same procedure can be applied to other organelles or signaling complexes. PMID:22094731

MRM screening/biomarker discovery with linear ion trap MS: a library of human cancer-specific peptides.

PubMed

Yang, Xu; Lazar, Iulia M

2009-03-27

The discovery of novel protein biomarkers is essential in the clinical setting to enable early disease diagnosis and increase survivability rates. To facilitate differential expression analysis and biomarker discovery, a variety of tandem mass spectrometry (MS/MS)-based protein profiling techniques have been developed. For achieving sensitive detection and accurate quantitation, targeted MS screening approaches, such as multiple reaction monitoring (MRM), have been implemented. MCF-7 breast cancer protein cellular extracts were analyzed by 2D-strong cation exchange (SCX)/reversed phase liquid chromatography (RPLC) separations interfaced to linear ion trap MS detection. MS data were interpreted with the Sequest-based Bioworks software (Thermo Electron). In-house developed Perl-scripts were used to calculate the spectral counts and the representative fragment ions for each peptide. In this work, we report on the generation of a library of 9,677 peptides (p < 0.001), representing approximately 1,572 proteins from human breast cancer cells, that can be used for MRM/MS-based biomarker screening studies. For each protein, the library provides the number and sequence of detectable peptides, the charge state, the spectral count, the molecular weight, the parameters that characterize the quality of the tandem mass spectrum (p-value, DeltaM, Xcorr, DeltaCn, Sp, no. of matching a, b, y ions in the spectrum), the retention time, and the top 10 most intense product ions that correspond to a given peptide. Only proteins identified by at least two spectral counts are listed. The experimental distribution of protein frequencies, as a function of molecular weight, closely matched the theoretical distribution of proteins in the human proteome, as provided in the SwissProt database. The amino acid sequence coverage of the identified proteins ranged from 0.04% to 98.3%. The highest-abundance proteins in the cellular extract had a molecular weight (MW)<50,000. Preliminary experiments have demonstrated that putative biomarkers, that are not detectable by conventional data dependent MS acquisition methods in complex un-fractionated samples, can be reliable identified with the information provided in this library. Based on the spectral count, the quality of a tandem mass spectrum and the m/z values for a parent peptide and its most abundant daughter ions, MRM conditions can be selected to enable the detection of target peptides and proteins.

MRM screening/biomarker discovery with linear ion trap MS: a library of human cancer-specific peptides

PubMed Central

2009-01-01

Background The discovery of novel protein biomarkers is essential in the clinical setting to enable early disease diagnosis and increase survivability rates. To facilitate differential expression analysis and biomarker discovery, a variety of tandem mass spectrometry (MS/MS)-based protein profiling techniques have been developed. For achieving sensitive detection and accurate quantitation, targeted MS screening approaches, such as multiple reaction monitoring (MRM), have been implemented. Methods MCF-7 breast cancer protein cellular extracts were analyzed by 2D-strong cation exchange (SCX)/reversed phase liquid chromatography (RPLC) separations interfaced to linear ion trap MS detection. MS data were interpreted with the Sequest-based Bioworks software (Thermo Electron). In-house developed Perl-scripts were used to calculate the spectral counts and the representative fragment ions for each peptide. Results In this work, we report on the generation of a library of 9,677 peptides (p < 0.001), representing ~1,572 proteins from human breast cancer cells, that can be used for MRM/MS-based biomarker screening studies. For each protein, the library provides the number and sequence of detectable peptides, the charge state, the spectral count, the molecular weight, the parameters that characterize the quality of the tandem mass spectrum (p-value, DeltaM, Xcorr, DeltaCn, Sp, no. of matching a, b, y ions in the spectrum), the retention time, and the top 10 most intense product ions that correspond to a given peptide. Only proteins identified by at least two spectral counts are listed. The experimental distribution of protein frequencies, as a function of molecular weight, closely matched the theoretical distribution of proteins in the human proteome, as provided in the SwissProt database. The amino acid sequence coverage of the identified proteins ranged from 0.04% to 98.3%. The highest-abundance proteins in the cellular extract had a molecular weight (MW)<50,000. Conclusion Preliminary experiments have demonstrated that putative biomarkers, that are not detectable by conventional data dependent MS acquisition methods in complex un-fractionated samples, can be reliable identified with the information provided in this library. Based on the spectral count, the quality of a tandem mass spectrum and the m/z values for a parent peptide and its most abundant daughter ions, MRM conditions can be selected to enable the detection of target peptides and proteins. PMID:19327145

Database-independent Protein Sequencing (DiPS) Enables Full-length de Novo Protein and Antibody Sequence Determination.

PubMed

Savidor, Alon; Barzilay, Rotem; Elinger, Dalia; Yarden, Yosef; Lindzen, Moshit; Gabashvili, Alexandra; Adiv Tal, Ophir; Levin, Yishai

2017-06-01

Traditional "bottom-up" proteomic approaches use proteolytic digestion, LC-MS/MS, and database searching to elucidate peptide identities and their parent proteins. Protein sequences absent from the database cannot be identified, and even if present in the database, complete sequence coverage is rarely achieved even for the most abundant proteins in the sample. Thus, sequencing of unknown proteins such as antibodies or constituents of metaproteomes remains a challenging problem. To date, there is no available method for full-length protein sequencing, independent of a reference database, in high throughput. Here, we present Database-independent Protein Sequencing, a method for unambiguous, rapid, database-independent, full-length protein sequencing. The method is a novel combination of non-enzymatic, semi-random cleavage of the protein, LC-MS/MS analysis, peptide de novo sequencing, extraction of peptide tags, and their assembly into a consensus sequence using an algorithm named "Peptide Tag Assembler." As proof-of-concept, the method was applied to samples of three known proteins representing three size classes and to a previously un-sequenced, clinically relevant monoclonal antibody. Excluding leucine/isoleucine and glutamic acid/deamidated glutamine ambiguities, end-to-end full-length de novo sequencing was achieved with 99-100% accuracy for all benchmarking proteins and the antibody light chain. Accuracy of the sequenced antibody heavy chain, including the entire variable region, was also 100%, but there was a 23-residue gap in the constant region sequence. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

An Integrated Bioinformatics and Computational Biology Approach Identifies New BH3-Only Protein Candidates.

PubMed

Hawley, Robert G; Chen, Yuzhong; Riz, Irene; Zeng, Chen

2012-05-04

In this study, we utilized an integrated bioinformatics and computational biology approach in search of new BH3-only proteins belonging to the BCL2 family of apoptotic regulators. The BH3 (BCL2 homology 3) domain mediates specific binding interactions among various BCL2 family members. It is composed of an amphipathic α-helical region of approximately 13 residues that has only a few amino acids that are highly conserved across all members. Using a generalized motif, we performed a genome-wide search for novel BH3-containing proteins in the NCBI Consensus Coding Sequence (CCDS) database. In addition to known pro-apoptotic BH3-only proteins, 197 proteins were recovered that satisfied the search criteria. These were categorized according to α-helical content and predictive binding to BCL-xL (encoded by BCL2L1) and MCL-1, two representative anti-apoptotic BCL2 family members, using position-specific scoring matrix models. Notably, the list is enriched for proteins associated with autophagy as well as a broad spectrum of cellular stress responses such as endoplasmic reticulum stress, oxidative stress, antiviral defense, and the DNA damage response. Several potential novel BH3-containing proteins are highlighted. In particular, the analysis strongly suggests that the apoptosis inhibitor and DNA damage response regulator, AVEN, which was originally isolated as a BCL-xL-interacting protein, is a functional BH3-only protein representing a distinct subclass of BCL2 family members.

mPLR-Loc: an adaptive decision multi-label classifier based on penalized logistic regression for protein subcellular localization prediction.

PubMed

Wan, Shibiao; Mak, Man-Wai; Kung, Sun-Yuan

2015-03-15

Proteins located in appropriate cellular compartments are of paramount importance to exert their biological functions. Prediction of protein subcellular localization by computational methods is required in the post-genomic era. Recent studies have been focusing on predicting not only single-location proteins but also multi-location proteins. However, most of the existing predictors are far from effective for tackling the challenges of multi-label proteins. This article proposes an efficient multi-label predictor, namely mPLR-Loc, based on penalized logistic regression and adaptive decisions for predicting both single- and multi-location proteins. Specifically, for each query protein, mPLR-Loc exploits the information from the Gene Ontology (GO) database by using its accession number (AC) or the ACs of its homologs obtained via BLAST. The frequencies of GO occurrences are used to construct feature vectors, which are then classified by an adaptive decision-based multi-label penalized logistic regression classifier. Experimental results based on two recent stringent benchmark datasets (virus and plant) show that mPLR-Loc remarkably outperforms existing state-of-the-art multi-label predictors. In addition to being able to rapidly and accurately predict subcellular localization of single- and multi-label proteins, mPLR-Loc can also provide probabilistic confidence scores for the prediction decisions. For readers' convenience, the mPLR-Loc server is available online (http://bioinfo.eie.polyu.edu.hk/mPLRLocServer). Copyright © 2014 Elsevier Inc. All rights reserved.

Detection of dysregulated protein-association networks by high-throughput proteomics predicts cancer vulnerabilities.

PubMed

Lapek, John D; Greninger, Patricia; Morris, Robert; Amzallag, Arnaud; Pruteanu-Malinici, Iulian; Benes, Cyril H; Haas, Wilhelm

2017-10-01

The formation of protein complexes and the co-regulation of the cellular concentrations of proteins are essential mechanisms for cellular signaling and for maintaining homeostasis. Here we use isobaric-labeling multiplexed proteomics to analyze protein co-regulation and show that this allows the identification of protein-protein associations with high accuracy. We apply this 'interactome mapping by high-throughput quantitative proteome analysis' (IMAHP) method to a panel of 41 breast cancer cell lines and show that deviations of the observed protein co-regulations in specific cell lines from the consensus network affects cellular fitness. Furthermore, these aberrant interactions serve as biomarkers that predict the drug sensitivity of cell lines in screens across 195 drugs. We expect that IMAHP can be broadly used to gain insight into how changing landscapes of protein-protein associations affect the phenotype of biological systems.

Metagenomic Taxonomy-Guided Database-Searching Strategy for Improving Metaproteomic Analysis.

PubMed

Xiao, Jinqiu; Tanca, Alessandro; Jia, Ben; Yang, Runqing; Wang, Bo; Zhang, Yu; Li, Jing

2018-04-06

Metaproteomics provides a direct measure of the functional information by investigating all proteins expressed by a microbiota. However, due to the complexity and heterogeneity of microbial communities, it is very hard to construct a sequence database suitable for a metaproteomic study. Using a public database, researchers might not be able to identify proteins from poorly characterized microbial species, while a sequencing-based metagenomic database may not provide adequate coverage for all potentially expressed protein sequences. To address this challenge, we propose a metagenomic taxonomy-guided database-search strategy (MT), in which a merged database is employed, consisting of both taxonomy-guided reference protein sequences from public databases and proteins from metagenome assembly. By applying our MT strategy to a mock microbial mixture, about two times as many peptides were detected as with the metagenomic database only. According to the evaluation of the reliability of taxonomic attribution, the rate of misassignments was comparable to that obtained using an a priori matched database. We also evaluated the MT strategy with a human gut microbial sample, and we found 1.7 times as many peptides as using a standard metagenomic database. In conclusion, our MT strategy allows the construction of databases able to provide high sensitivity and precision in peptide identification in metaproteomic studies, enabling the detection of proteins from poorly characterized species within the microbiota.

Piezo Proteins: Regulators of Mechanosensation and Other Cellular Processes*

PubMed Central

Bagriantsev, Sviatoslav N.; Gracheva, Elena O.; Gallagher, Patrick G.

2014-01-01

Piezo proteins have recently been identified as ion channels mediating mechanosensory transduction in mammalian cells. Characterization of these channels has yielded important insights into mechanisms of somatosensation, as well as other mechano-associated biologic processes such as sensing of shear stress, particularly in the vasculature, and regulation of urine flow and bladder distention. Other roles for Piezo proteins have emerged, some unexpected, including participation in cellular development, volume regulation, cellular migration, proliferation, and elongation. Mutations in human Piezo proteins have been associated with a variety of disorders including hereditary xerocytosis and several syndromes with muscular contracture as a prominent feature. PMID:25305018

O-GlcNAc cycling: how a single sugar post-translational modification is changing the way we think about signaling networks.

PubMed

Slawson, Chad; Housley, Michael P; Hart, Gerald W

2006-01-01

O-GlcNAc is an ubiquitous post-translational protein modification consisting of a single N-acetlyglucosamine moiety linked to serine or threonine residues on nuclear and cytoplasmic proteins. Recent work has begun to uncover the functional roles of O-GlcNAc in cellular processes. O-GlcNAc modified proteins are involved in sensing the nutrient status of the surrounding cellular environment and adjusting the activity of cellular proteins accordingly. O-GlcNAc regulates cellular responses to hormones such as insulin, initiates a protective response to stress, modulates a cell's capacity to grow and divide, and regulates gene transcription. This review will focus on recent work involving O-GlcNAc in sensing the environment and regulating signaling cascades. (c) 2005 Wiley-Liss, Inc.

Nipah virus matrix protein: expert hacker of cellular machines.

PubMed

Watkinson, Ruth E; Lee, Benhur

2016-08-01

Nipah virus (NiV, Henipavirus) is a highly lethal emergent zoonotic paramyxovirus responsible for repeated human outbreaks of encephalitis in South East Asia. There are no approved vaccines or treatments, thus improved understanding of NiV biology is imperative. NiV matrix protein recruits a plethora of cellular machinery to scaffold and coordinate virion budding. Intriguingly, matrix also hijacks cellular trafficking and ubiquitination pathways to facilitate transient nuclear localization. While the biological significance of matrix nuclear localization for an otherwise cytoplasmic virus remains enigmatic, the molecular details have begun to be characterized, and are conserved among matrix proteins from divergent paramyxoviruses. Matrix protein appropriation of cellular machinery will be discussed in terms of its early nuclear targeting and later role in virion assembly. © 2016 Federation of European Biochemical Societies.

Merging in-silico and in vitro salivary protein complex partners using the STRING database: A tutorial.

PubMed

Crosara, Karla Tonelli Bicalho; Moffa, Eduardo Buozi; Xiao, Yizhi; Siqueira, Walter Luiz

2018-01-16

Protein-protein interaction is a common physiological mechanism for protection and actions of proteins in an organism. The identification and characterization of protein-protein interactions in different organisms is necessary to better understand their physiology and to determine their efficacy. In a previous in vitro study using mass spectrometry, we identified 43 proteins that interact with histatin 1. Six previously documented interactors were confirmed and 37 novel partners were identified. In this tutorial, we aimed to demonstrate the usefulness of the STRING database for studying protein-protein interactions. We used an in-silico approach along with the STRING database (http://string-db.org/) and successfully performed a fast simulation of a novel constructed histatin 1 protein-protein network, including both the previously known and the predicted interactors, along with our newly identified interactors. Our study highlights the advantages and importance of applying bioinformatics tools to merge in-silico tactics with experimental in vitro findings for rapid advancement of our knowledge about protein-protein interactions. Our findings also indicate that bioinformatics tools such as the STRING protein network database can help predict potential interactions between proteins and thus serve as a guide for future steps in our exploration of the Human Interactome. Our study highlights the usefulness of the STRING protein database for studying protein-protein interactions. The STRING database can collect and integrate data about known and predicted protein-protein associations from many organisms, including both direct (physical) and indirect (functional) interactions, in an easy-to-use interface. Copyright © 2017 Elsevier B.V. All rights reserved.

Mechanisms of stress-induced cellular HSP72 release: implications for exercise-induced increases in extracellular HSP72.

PubMed

Lancaster, Graeme I; Febbraio, Mark A

2005-01-01

The heat shock proteins are a family of highly conserved proteins with critical roles in maintaining cellular homeostasis and in protecting the cell from stressful conditions. While the critical intracellular roles of heat shock proteins are undisputed, evidence suggests that the cell possess the necessary machinery to actively secrete specific heat shock proteins in response to cellular stress. In this review, we firstly discuss the evidence that physical exercise induces the release of heat shock protein 72 from specific tissues in humans. Importantly, it appears as though this release is the result of an active secretory process, as opposed to non-specific processes such as cell lysis. Next we discuss recent in vitro evidence that has identified a mechanistic basis for the observation that cellular stress induces the release of a specific subset of heat shock proteins. Importantly, while the classical protein secretory pathway does not seem to be involved in the stress-induced release of HSP72, we discuss the evidence that lipid-rafts and exosomes are important mediators of the stress-induced release of HSP72.

Proteomic analysis of cellular soluble proteins from human bronchial smooth muscle cells by combining nondenaturing micro 2DE and quantitative LC-MS/MS. 2. Similarity search between protein maps for the analysis of protein complexes.

PubMed

Jin, Ya; Yuan, Qi; Zhang, Jun; Manabe, Takashi; Tan, Wen

2015-09-01

Human bronchial smooth muscle cell soluble proteins were analyzed by a combined method of nondenaturing micro 2DE, grid gel-cutting, and quantitative LC-MS/MS and a native protein map was prepared for each of the identified 4323 proteins [1]. A method to evaluate the degree of similarity between the protein maps was developed since we expected the proteins comprising a protein complex would be separated together under nondenaturing conditions. The following procedure was employed using Excel macros; (i) maps that have three or more squares with protein quantity data were selected (2328 maps), (ii) within each map, the quantity values of the squares were normalized setting the highest value to be 1.0, (iii) in comparing a map with another map, the smaller normalized quantity in two corresponding squares was taken and summed throughout the map to give an "overlap score," (iv) each map was compared against all the 2328 maps and the largest overlap score, obtained when a map was compared with itself, was set to be 1.0 thus providing 2328 "overlap factors," (v) step (iv) was repeated for all maps providing 2328 × 2328 matrix of overlap factors. From the matrix, protein pairs that showed overlap factors above 0.65 from both protein sides were selected (431 protein pairs). Each protein pair was searched in a database (UniProtKB) on complex formation and 301 protein pairs, which comprise 35 protein complexes, were found to be documented. These results demonstrated that native protein maps and their similarity search would enable simultaneous analysis of multiple protein complexes in cells. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Function of ubiquitin (Ub) specific protease 15 (USP15) in HIV-1 replication and viral protein degradation.

PubMed

Pyeon, Dohun; Timani, Khalid Amine; Gulraiz, Fahad; He, Johnny J; Park, In-Woo

2016-09-02

HIV-1 Nef is necessary and may be sufficient for HIV-1-associated AIDS pathogenicity, in that knockout of Nef alone can protect HIV-infected patients from AIDS. We therefore investigated the feasibility of physical knockout of Nef, using the host ubiquitin proteasome system in HIV-1-infected cells. Our co-immunoprecipitation analysis demonstrated that Nef interacted with ubiquitin specific protease 15 (USP15), and that USP15, which is known to stabilize cellular proteins, degraded Nef. Nef could also cause decay of USP15, although Nef-mediated degradation of USP15 was weaker than USP15-mediated Nef degradation. Direct interaction between Nef and USP15 was essential for the observed reciprocal decay of the proteins. Further, USP15 degraded not only Nef but also HIV-1 structural protein, Gag, thereby substantially inhibiting HIV-1 replication. However, Gag did not degrade USP15, indicating that the Nef and USP15 complex, in distinction to other viral proteins, play an integral role in coordinating viral protein degradation and hence HIV-1 replication. Moreover, Nef and USP15 globally suppressed ubiquitylation of cellular proteins, indicating that these proteins are major determinants for the stability of cellular as well as viral proteins. Taken together, these data indicate that Nef and USP15 are vital in regulating degradation of viral and cellular proteins and thus HIV-1 replication, and specific degradation of viral, not cellular proteins, by USP15 points to USP15 as a candidate therapeutic agent to combat AIDS by eliminating viral proteins from the infected cells via USP15-mediated proteosomal degradation. Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.

Distal Renal Tubules Are Deficient in Aggresome Formation and Autophagy upon Aldosterone Administration

PubMed Central

Cheema, Muhammad Umar; Damkier, Helle Hasager; Nielsen, Jakob; Poulsen, Ebbe Toftgaard; Enghild, Jan J.; Fenton, Robert A.; Praetorius, Jeppe

2014-01-01

Prolonged elevations of plasma aldosterone levels are associated with renal pathogenesis. We hypothesized that renal distress could be imposed by an augmented aldosterone-induced protein turnover challenging cellular protein degradation systems of the renal tubular cells. Cellular accumulation of specific protein aggregates in rat kidneys was assessed after 7 days of aldosterone administration. Aldosterone induced intracellular accumulation of 60 s ribosomal protein L22 in protein aggregates, specifically in the distal convoluted tubules. The mineralocorticoid receptor inhibitor spironolactone abolished aldosterone-induced accumulation of these aggregates. The aldosterone-induced protein aggregates also contained proteasome 20 s subunits. The partial de-ubiquitinase ataxin-3 was not localized to the distal renal tubule protein aggregates, and the aggregates only modestly colocalized with aggresome transfer proteins dynactin p62 and histone deacetylase 6. Intracellular protein aggregation in distal renal tubules did not lead to development of classical juxta-nuclear aggresomes or to autophagosome formation. Finally, aldosterone treatment induced foci in renal cortex of epithelial vimentin expression and a loss of E-cadherin expression, as signs of cellular stress. The cellular changes occurred within high, but physiological aldosterone concentrations. We conclude that aldosterone induces protein accumulation in distal renal tubules; these aggregates are not cleared by autophagy that may lead to early renal tubular damage. PMID:25000288

Resolving the homology—function relationship through comparative genomics of membrane-trafficking machinery and parasite cell biology

PubMed Central

Klinger, Christen M.; Ramirez-Macias, Inmaculada; Herman, Emily K.; Turkewitz, Aaron P.; Field, Mark C.; Dacks, Joel B.

2016-01-01

With advances in DNA sequencing technology, it is increasingly common and tractable to informatically look for genes of interest in the genomic databases of parasitic organisms and infer cellular states. Assignment of a putative gene function based on homology to functionally characterized genes in other organisms, though powerful, relies on the implicit assumption of functional homology, i.e. that orthology indicates conserved function. Eukaryotes reveal a dazzling array of cellular features and structural organization, suggesting a concomitant diversity in their underlying molecular machinery. Significantly, examples of novel functions for pre-existing or new paralogues are not uncommon. Do these examples undermine the basic assumption of functional homology, especially in parasitic protists, which are often highly derived? Here we examine the extent to which functional homology exists between organisms spanning the eukaryotic lineage. By comparing membrane trafficking proteins between parasitic protists and traditional model organisms, where direct functional evidence is available, we find that function is indeed largely conserved between orthologues, albeit with significant adaptation arising from the unique biological features within each lineage. PMID:27444378

Oxidative stress and protein aggregation during biological aging.

PubMed

Squier, T C

2001-09-01

Biological aging is a fundamental process that represents the major risk factor with respect to the development of cancer, neurodegenerative, and cardiovascular diseases in vertebrates. It is, therefore, evident that the molecular mechanisms of aging are fundamental to understand many disease processes. In this regard, the oxidation and nitration of intracellular proteins and the formation of protein aggregates have been suggested to underlie the loss of cellular function and the reduced ability of senescent animals to withstand physiological stresses. Since oxidatively modified proteins are thermodynamically unstable and assume partially unfolded tertiary structures that readily form aggregates, it is likely that oxidized proteins are intermediates in the formation of amyloid fibrils. It is, therefore, of interest to identify oxidatively sensitive protein targets that may play a protective role through their ability to down-regulate energy metabolism and the consequent generation of reactive oxygen species (ROS). In this respect, the maintenance of cellular calcium gradients represents a major energetic expense, which links alterations in intracellular calcium levels to ATP utilization and the associated generation of ROS through respiratory control mechanisms. The selective oxidation or nitration of the calcium regulatory proteins calmodulin and Ca-ATPase that occurs in vivo during aging and under conditions of oxidative stress may represent an adaptive response to oxidative stress that functions to down-regulate energy metabolism and the associated generation of ROS. Since these calcium regulatory proteins are also preferentially oxidized or nitrated under in vitro conditions, these results suggest an enhanced sensitivity of these critical calcium regulatory proteins, which modulate signal transduction processes and intracellular energy metabolism, to conditions of oxidative stress. Thus, the selective oxidation of critical signal transduction proteins probably represents a regulatory mechanism that functions to minimize the generation of ROS through respiratory control mechanisms. The reduction of the rate of ROS generation, in turn, will promote cellular survival under conditions of oxidative stress, when reactive oxygen and nitrogen species overwhelm cellular antioxidant defense systems, by minimizing the non-selective oxidation of a range of biomolecules. Since protein aggregation occurs if protein repair and degradative systems are unable to act upon oxidized proteins and restore cellular function, the reduction of the oxidative load on the cell by the down-regulation of the electron transport chain functions to minimize protein aggregation. Thus, ROS function as signaling molecules that fine-tune cellular metabolism through the selective oxidation or nitration of calcium regulatory proteins in order to minimize wide-spread oxidative damage and protein aggregation. Oxidative damage to cellular proteins, the loss of calcium homeostasis and protein aggregation contribute to the formation of amyloid deposits that accumulate during biological aging. Critical to understand the relationship between these processes and biological aging is the identification of oxidatively sensitive proteins that modulate energy utilization and the associated generation of ROS. In this latter respect, oxidative modifications to the calcium regulatory proteins calmodulin (CaM) and the sarco/endoplasmic reticulum Ca-ATPase (SERCA) function to down-regulate ATP utilization and the associated generation of ROS associated with replenishing intracellular ATP through oxidative phosphorylation. Reductions in the rate of ROS generation, in turn, will minimize protein oxidation and facilitate intracellular repair and degradative systems that function to eliminate damaged and partially unfolded proteins. Since the rates of protein repair or degradation compete with the rate of protein aggregation, the modulation of intracellular calcium concentrations and energy metabolism through the selective oxidation or nitration of critical signal transduction proteins (i.e. CaM or SERCA) is thought to maintain cellular function by minimizing protein aggregation and amyloid formation. Age-dependent increases in the rate of ROS generation or declines in cellular repair or degradation mechanisms will increase the oxidative load on the cell, resulting in corresponding increases in the concentrations of oxidized proteins and the associated formation of amyloid.

Regulation of cell function by methionine oxidation and reduction

PubMed Central

Hoshi, Toshinori; Heinemann, Stefan H

2001-01-01

Reactive oxygen species (ROS) are generated during normal cellular activity and may exist in excess in some pathophysiological conditions, such as inflammation or reperfusion injury. These molecules oxidize a variety of cellular constituents, but sulfur-containing amino acid residues are especially susceptible. While reversible cysteine oxidation and reduction is part of well-established signalling systems, the oxidation and the enzymatically catalysed reduction of methionine is just emerging as a novel molecular mechanism for cellular regulation. Here we discuss how the oxidation of methionine to methionine sulfoxide in signalling proteins such as ion channels affects the function of these target proteins. Methionine sulfoxide reductase, which reduces methionine sulfoxide to methionine in a thioredoxin-dependent manner, is therefore not only an enzyme important for the repair of age- or degenerative disease-related protein modifications. It is also a potential missing link in the post-translational modification cycle involved in the specific oxidation and reduction of methionine residues in cellular signalling proteins, which may give rise to activity-dependent plastic changes in cellular excitability. PMID:11179387

Phenylbutyric acid induces the cellular senescence through an Akt/p21{sup WAF1} signaling pathway

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kim, Hag Dong; Jang, Chang-Young; Choe, Jeong Min

2012-06-01

Highlights: Black-Right-Pointing-Pointer Phenylbutyric acid induces cellular senescence. Black-Right-Pointing-Pointer Phenylbutyric acid activates Akt kinase. Black-Right-Pointing-Pointer The knockdown of PERK also can induce cellular senescence. Black-Right-Pointing-Pointer Akt/p21{sup WAF1} pathway activates in PERK knockdown induced cellular senescence. -- Abstract: It has been well known that three sentinel proteins - PERK, ATF6 and IRE1 - initiate the unfolded protein response (UPR) in the presence of misfolded or unfolded proteins in the ER. Recent studies have demonstrated that upregulation of UPR in cancer cells is required to survive and proliferate. Here, we showed that long exposure to 4-phenylbutyric acid (PBA), a chemical chaperone that canmore » reduce retention of unfolded and misfolded proteins in ER, induced cellular senescence in cancer cells such as MCF7 and HT1080. In addition, we found that treatment with PBA activates Akt, which results in p21{sup WAF1} induction. Interestingly, the depletion of PERK but not ATF6 and IRE1 also induces cellular senescence, which was rescued by additional depletion of Akt. This suggests that Akt pathway is downstream of PERK in PBA induced cellular senescence. Taken together, these results show that PBA induces cellular senescence via activation of the Akt/p21{sup WAF1} pathway by PERK inhibition.« less

Prions: Beyond a Single Protein

PubMed Central

Das, Alvin S.

2016-01-01

SUMMARY Since the term protein was first coined in 1838 and protein was discovered to be the essential component of fibrin and albumin, all cellular proteins were presumed to play beneficial roles in plants and mammals. However, in 1967, Griffith proposed that proteins could be infectious pathogens and postulated their involvement in scrapie, a universally fatal transmissible spongiform encephalopathy in goats and sheep. Nevertheless, this novel hypothesis had not been evidenced until 1982, when Prusiner and coworkers purified infectious particles from scrapie-infected hamster brains and demonstrated that they consisted of a specific protein that he called a “prion.” Unprecedentedly, the infectious prion pathogen is actually derived from its endogenous cellular form in the central nervous system. Unlike other infectious agents, such as bacteria, viruses, and fungi, prions do not contain genetic materials such as DNA or RNA. The unique traits and genetic information of prions are believed to be encoded within the conformational structure and posttranslational modifications of the proteins. Remarkably, prion-like behavior has been recently observed in other cellular proteins—not only in pathogenic roles but also serving physiological functions. The significance of these fascinating developments in prion biology is far beyond the scope of a single cellular protein and its related disease. PMID:27226089

The protein expression landscape of the Arabidopsis root

PubMed Central

Petricka, Jalean J.; Schauer, Monica A.; Megraw, Molly; Breakfield, Natalie W.; Thompson, J. Will; Georgiev, Stoyan; Soderblom, Erik J.; Ohler, Uwe; Moseley, Martin Arthur; Grossniklaus, Ueli; Benfey, Philip N.

2012-01-01

Because proteins are the major functional components of cells, knowledge of their cellular localization is crucial to gaining an understanding of the biology of multicellular organisms. We have generated a protein expression map of the Arabidopsis root providing the identity and cell type-specific localization of nearly 2,000 proteins. Grouping proteins into functional categories revealed unique cellular functions and identified cell type-specific biomarkers. Cellular colocalization provided support for numerous protein–protein interactions. With a binary comparison, we found that RNA and protein expression profiles are weakly correlated. We then performed peak integration at cell type-specific resolution and found an improved correlation with transcriptome data using continuous values. We performed GeLC-MS/MS (in-gel tryptic digestion followed by liquid chromatography-tandem mass spectrometry) proteomic experiments on mutants with ectopic and no root hairs, providing complementary proteomic data. Finally, among our root hair-specific proteins we identified two unique regulators of root hair development. PMID:22447775

The Protein Disease Database of human body fluids: II. Computer methods and data issues.

PubMed

Lemkin, P F; Orr, G A; Goldstein, M P; Creed, G J; Myrick, J E; Merril, C R

1995-01-01

The Protein Disease Database (PDD) is a relational database of proteins and diseases. With this database it is possible to screen for quantitative protein abnormalities associated with disease states. These quantitative relationships use data drawn from the peer-reviewed biomedical literature. Assays may also include those observed in high-resolution electrophoretic gels that offer the potential to quantitate many proteins in a single test as well as data gathered by enzymatic or immunologic assays. We are using the Internet World Wide Web (WWW) and the Web browser paradigm as an access method for wide distribution and querying of the Protein Disease Database. The WWW hypertext transfer protocol and its Common Gateway Interface make it possible to build powerful graphical user interfaces that can support easy-to-use data retrieval using query specification forms or images. The details of these interactions are totally transparent to the users of these forms. Using a client-server SQL relational database, user query access, initial data entry and database maintenance are all performed over the Internet with a Web browser. We discuss the underlying design issues, mapping mechanisms and assumptions that we used in constructing the system, data entry, access to the database server, security, and synthesis of derived two-dimensional gel image maps and hypertext documents resulting from SQL database searches.

Domain fusion analysis by applying relational algebra to protein sequence and domain databases

PubMed Central

Truong, Kevin; Ikura, Mitsuhiko

2003-01-01

Background Domain fusion analysis is a useful method to predict functionally linked proteins that may be involved in direct protein-protein interactions or in the same metabolic or signaling pathway. As separate domain databases like BLOCKS, PROSITE, Pfam, SMART, PRINTS-S, ProDom, TIGRFAMs, and amalgamated domain databases like InterPro continue to grow in size and quality, a computational method to perform domain fusion analysis that leverages on these efforts will become increasingly powerful. Results This paper proposes a computational method employing relational algebra to find domain fusions in protein sequence databases. The feasibility of this method was illustrated on the SWISS-PROT+TrEMBL sequence database using domain predictions from the Pfam HMM (hidden Markov model) database. We identified 235 and 189 putative functionally linked protein partners in H. sapiens and S. cerevisiae, respectively. From scientific literature, we were able to confirm many of these functional linkages, while the remainder offer testable experimental hypothesis. Results can be viewed at . Conclusion As the analysis can be computed quickly on any relational database that supports standard SQL (structured query language), it can be dynamically updated along with the sequence and domain databases, thereby improving the quality of predictions over time. PMID:12734020

Cellular oxido-reductive proteins of Chlamydomonas reinhardtii control the biosynthesis of silver nanoparticles

PubMed Central

2011-01-01

Background Elucidation of molecular mechanism of silver nanoparticles (SNPs) biosynthesis is important to control its size, shape and monodispersity. The evaluation of molecular mechanism of biosynthesis of SNPs is of prime importance for the commercialization and methodology development for controlling the shape and size (uniform distribution) of SNPs. The unicellular algae Chlamydomonas reinhardtii was exploited as a model system to elucidate the role of cellular proteins in SNPs biosynthesis. Results The C. reinhardtii cell free extract (in vitro) and in vivo cells mediated synthesis of silver nanoparticles reveals SNPs of size range 5 ± 1 to 15 ± 2 nm and 5 ± 1 to 35 ± 5 nm respectively. In vivo biosynthesized SNPs were localized in the peripheral cytoplasm and at one side of flagella root, the site of pathway of ATP transport and its synthesis related enzymes. This provides an evidence for the involvement of oxidoreductive proteins in biosynthesis and stabilization of SNPs. Alteration in size distribution and decrease of synthesis rate of SNPs in protein-depleted fractions confirmed the involvement of cellular proteins in SNPs biosynthesis. Spectroscopic and SDS-PAGE analysis indicate the association of various proteins on C. reinhardtii mediated in vivo and in vitro biosynthesized SNPs. We have identified various cellular proteins associated with biosynthesized (in vivo and in vitro) SNPs by using MALDI-MS-MS, like ATP synthase, superoxide dismutase, carbonic anhydrase, ferredoxin-NADP+ reductase, histone etc. However, these proteins were not associated on the incubation of pre-synthesized silver nanoparticles in vitro. Conclusion Present study provides the indication of involvement of molecular machinery and various cellular proteins in the biosynthesis of silver nanoparticles. In this report, the study is mainly focused towards understanding the role of diverse cellular protein in the synthesis and capping of silver nanoparticles using C. reinhardtii as a model system. PMID:22152042

Trichoderma G protein-coupled receptors: functional characterisation of a cAMP receptor-like protein from Trichoderma atroviride.

PubMed

Brunner, Kurt; Omann, Markus; Pucher, Marion E; Delic, Marizela; Lehner, Sylvia M; Domnanich, Patrick; Kratochwill, Klaus; Druzhinina, Irina; Denk, Dagmar; Zeilinger, Susanne

2008-12-01

Galpha subunits act to regulate vegetative growth, conidiation, and the mycoparasitic response in Trichoderma atroviride. To extend our knowledge on G protein signalling, we analysed G protein-coupled receptors (GPCRs). As the genome sequence of T. atroviride is not publicly available yet, we carried out an in silico exploration of the genome database of the close relative T. reesei. Twenty genes encoding putative GPCRs distributed over eight classes and additional 35 proteins similar to the Magnaporthe grisea PTH11 receptor were identified. Subsequently, four T. atroviride GPCR-encoding genes were isolated and affiliated to the cAMP receptor-like family by phylogenetic and topological analyses. All four genes showed lowest expression on glycerol and highest mRNA levels upon carbon starvation. Transcription of gpr3 and gpr4 responded to exogenously added cAMP and the shift from liquid to solid media. gpr3 mRNA levels also responded to the presence of fungal hyphae or cellulose membranes. Further characterisation of mutants bearing a gpr1-silencing construct revealed that Gpr1 is essential for vegetative growth, conidiation and conidial germination. Four genes encoding the first GPCRs described in Trichoderma were isolated and their expression characterized. At least one of these GPCRs is important for several cellular processes, supporting the fundamental role of G protein signalling in this fungus.

Papillomavirus E6 oncoproteins

PubMed Central

Vande Pol, Scott B.; Klingelhutz, Aloysius J.

2013-01-01

Papillomaviruses induce benign and malignant epithelial tumors, and the viral E6 oncoprotein is essential for full transformation. E6 contributes to transformation by associating with cellular proteins, docking on specific acidic LXXLL peptide motifs found on the associated cellular proteins. This review examines insights from recent studies of human and animal E6 proteins that determine the three-dimensional structure of E6 when bound to acidic LXXLL peptides. The structure of E6 is related to recent advances in the purification and identification of E6 associated protein complexes. These E6 protein-complexes, together with other proteins that bind to E6, alter a broad array of biological outcomes including modulation of cell survival, cellular transcription, host cell differentiation, growth factor dependence, DNA damage responses, and cell cycle progression. PMID:23711382

TOPDOM: database of conservatively located domains and motifs in proteins.

PubMed

Varga, Julia; Dobson, László; Tusnády, Gábor E

2016-09-01

The TOPDOM database-originally created as a collection of domains and motifs located consistently on the same side of the membranes in α-helical transmembrane proteins-has been updated and extended by taking into consideration consistently localized domains and motifs in globular proteins, too. By taking advantage of the recently developed CCTOP algorithm to determine the type of a protein and predict topology in case of transmembrane proteins, and by applying a thorough search for domains and motifs as well as utilizing the most up-to-date version of all source databases, we managed to reach a 6-fold increase in the size of the whole database and a 2-fold increase in the number of transmembrane proteins. TOPDOM database is available at http://topdom.enzim.hu The webpage utilizes the common Apache, PHP5 and MySQL software to provide the user interface for accessing and searching the database. The database itself is generated on a high performance computer. tusnady.gabor@ttk.mta.hu Supplementary data are available at Bioinformatics online. © The Author 2016. Published by Oxford University Press.

COMBREX-DB: an experiment centered database of protein function: knowledge, predictions and knowledge gaps.

PubMed

Chang, Yi-Chien; Hu, Zhenjun; Rachlin, John; Anton, Brian P; Kasif, Simon; Roberts, Richard J; Steffen, Martin

2016-01-04

The COMBREX database (COMBREX-DB; combrex.bu.edu) is an online repository of information related to (i) experimentally determined protein function, (ii) predicted protein function, (iii) relationships among proteins of unknown function and various types of experimental data, including molecular function, protein structure, and associated phenotypes. The database was created as part of the novel COMBREX (COMputational BRidges to EXperiments) effort aimed at accelerating the rate of gene function validation. It currently holds information on ∼ 3.3 million known and predicted proteins from over 1000 completely sequenced bacterial and archaeal genomes. The database also contains a prototype recommendation system for helping users identify those proteins whose experimental determination of function would be most informative for predicting function for other proteins within protein families. The emphasis on documenting experimental evidence for function predictions, and the prioritization of uncharacterized proteins for experimental testing distinguish COMBREX from other publicly available microbial genomics resources. This article describes updates to COMBREX-DB since an initial description in the 2011 NAR Database Issue. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

The HUPO PSI's molecular interaction format--a community standard for the representation of protein interaction data.

PubMed

Hermjakob, Henning; Montecchi-Palazzi, Luisa; Bader, Gary; Wojcik, Jérôme; Salwinski, Lukasz; Ceol, Arnaud; Moore, Susan; Orchard, Sandra; Sarkans, Ugis; von Mering, Christian; Roechert, Bernd; Poux, Sylvain; Jung, Eva; Mersch, Henning; Kersey, Paul; Lappe, Michael; Li, Yixue; Zeng, Rong; Rana, Debashis; Nikolski, Macha; Husi, Holger; Brun, Christine; Shanker, K; Grant, Seth G N; Sander, Chris; Bork, Peer; Zhu, Weimin; Pandey, Akhilesh; Brazma, Alvis; Jacq, Bernard; Vidal, Marc; Sherman, David; Legrain, Pierre; Cesareni, Gianni; Xenarios, Ioannis; Eisenberg, David; Steipe, Boris; Hogue, Chris; Apweiler, Rolf

2004-02-01

A major goal of proteomics is the complete description of the protein interaction network underlying cell physiology. A large number of small scale and, more recently, large-scale experiments have contributed to expanding our understanding of the nature of the interaction network. However, the necessary data integration across experiments is currently hampered by the fragmentation of publicly available protein interaction data, which exists in different formats in databases, on authors' websites or sometimes only in print publications. Here, we propose a community standard data model for the representation and exchange of protein interaction data. This data model has been jointly developed by members of the Proteomics Standards Initiative (PSI), a work group of the Human Proteome Organization (HUPO), and is supported by major protein interaction data providers, in particular the Biomolecular Interaction Network Database (BIND), Cellzome (Heidelberg, Germany), the Database of Interacting Proteins (DIP), Dana Farber Cancer Institute (Boston, MA, USA), the Human Protein Reference Database (HPRD), Hybrigenics (Paris, France), the European Bioinformatics Institute's (EMBL-EBI, Hinxton, UK) IntAct, the Molecular Interactions (MINT, Rome, Italy) database, the Protein-Protein Interaction Database (PPID, Edinburgh, UK) and the Search Tool for the Retrieval of Interacting Genes/Proteins (STRING, EMBL, Heidelberg, Germany).

Functional phylogenomics analysis of bacteria and archaea using consistent genome annotation with UniFam

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chai, Juanjuan; Kora, Guruprasad; Ahn, Tae-Hyuk

2014-10-09

To supply some background, phylogenetic studies have provided detailed knowledge on the evolutionary mechanisms of genes and species in Bacteria and Archaea. However, the evolution of cellular functions, represented by metabolic pathways and biological processes, has not been systematically characterized. Many clades in the prokaryotic tree of life have now been covered by sequenced genomes in GenBank. This enables a large-scale functional phylogenomics study of many computationally inferred cellular functions across all sequenced prokaryotes. Our results show a total of 14,727 GenBank prokaryotic genomes were re-annotated using a new protein family database, UniFam, to obtain consistent functional annotations for accuratemore » comparison. The functional profile of a genome was represented by the biological process Gene Ontology (GO) terms in its annotation. The GO term enrichment analysis differentiated the functional profiles between selected archaeal taxa. 706 prokaryotic metabolic pathways were inferred from these genomes using Pathway Tools and MetaCyc. The consistency between the distribution of metabolic pathways in the genomes and the phylogenetic tree of the genomes was measured using parsimony scores and retention indices. The ancestral functional profiles at the internal nodes of the phylogenetic tree were reconstructed to track the gains and losses of metabolic pathways in evolutionary history. In conclusion, our functional phylogenomics analysis shows divergent functional profiles of taxa and clades. Such function-phylogeny correlation stems from a set of clade-specific cellular functions with low parsimony scores. On the other hand, many cellular functions are sparsely dispersed across many clades with high parsimony scores. These different types of cellular functions have distinct evolutionary patterns reconstructed from the prokaryotic tree.« less

Transcriptome-wide analysis of WRKY transcription factors in wheat and their leaf rust responsive expression profiling.

PubMed

Satapathy, Lopamudra; Singh, Dharmendra; Ranjan, Prashant; Kumar, Dhananjay; Kumar, Manish; Prabhu, Kumble Vinod; Mukhopadhyay, Kunal

2014-12-01

WRKY, a plant-specific transcription factor family, has important roles in pathogen defense, abiotic cues and phytohormone signaling, yet little is known about their roles and molecular mechanism of function in response to rust diseases in wheat. We identified 100 TaWRKY sequences using wheat Expressed Sequence Tag database of which 22 WRKY sequences were novel. Identified proteins were characterized based on their zinc finger motifs and phylogenetic analysis clustered them into six clades consisting of class IIc and class III WRKY proteins. Functional annotation revealed major functions in metabolic and cellular processes in control plants; whereas response to stimuli, signaling and defense in pathogen inoculated plants, their major molecular function being binding to DNA. Tag-based expression analysis of the identified genes revealed differential expression between mock and Puccinia triticina inoculated wheat near isogenic lines. Gene expression was also performed with six rust-related microarray experiments at Gene Expression Omnibus database. TaWRKY10, 15, 17 and 56 were common in both tag-based and microarray-based differential expression analysis and could be representing rust specific WRKY genes. The obtained results will bestow insight into the functional characterization of WRKY transcription factors responsive to leaf rust pathogenesis that can be used as candidate genes in molecular breeding programs to improve biotic stress tolerance in wheat.

The yeast two hybrid system in a screen for proteins interacting with axolotl (Ambystoma mexicanum) Msx1 during early limb regeneration.

PubMed

Abuqarn, Mehtap; Allmeling, Christina; Amshoff, Inga; Menger, Bjoern; Nasser, Inas; Vogt, Peter M; Reimers, Kerstin

2011-07-01

Urodele amphibians are exceptional in their ability to regenerate complex body structures such as limbs. Limb regeneration depends on a process called dedifferentiation. Under an inductive wound epidermis terminally differentiated cells transform to pluripotent progenitor cells that coordinately proliferate and eventually redifferentiate to form the new appendage. Recent studies have developed molecular models integrating a set of genes that might have important functions in the control of regenerative cellular plasticity. Among them is Msx1, which induced dedifferentiation in mammalian myotubes in vitro. Herein, we screened for interaction partners of axolotl Msx1 using a yeast two hybrid system. A two hybrid cDNA library of 5-day-old wound epidermis and underlying tissue containing more than 2×10⁶ cDNAs was constructed and used in the screen. 34 resulting cDNA clones were isolated and sequenced. We then compared sequences of the isolated clones to annotated EST contigs of the Salamander EST database (BLASTn) to identify presumptive orthologs. We subsequently searched all no-hit clone sequences against non redundant NCBI sequence databases using BLASTx. It is the first time, that the yeast two hybrid system was adapted to the axolotl animal model and successfully used in a screen for proteins interacting with Msx1 in the context of amphibian limb regeneration. 2011 Elsevier B.V. All rights reserved.

Gene Expression Profiling Reveals Functional Specialization along the Intestinal Tract of a Carnivorous Teleostean Fish (Dicentrarchus labrax)

PubMed Central

Calduch-Giner, Josep A.; Sitjà-Bobadilla, Ariadna; Pérez-Sánchez, Jaume

2016-01-01

High-quality sequencing reads from the intestine of European sea bass were assembled, annotated by similarity against protein reference databases and combined with nucleotide sequences from public and private databases. After redundancy filtering, 24,906 non-redundant annotated sequences encoding 15,367 different gene descriptions were obtained. These annotated sequences were used to design a custom, high-density oligo-microarray (8 × 15 K) for the transcriptomic profiling of anterior (AI), middle (MI), and posterior (PI) intestinal segments. Similar molecular signatures were found for AI and MI segments, which were combined in a single group (AI-MI) whereas the PI outstood separately, with more than 1900 differentially expressed genes with a fold-change cutoff of 2. Functional analysis revealed that molecular and cellular functions related to feed digestion and nutrient absorption and transport were over-represented in AI-MI segments. By contrast, the initiation and establishment of immune defense mechanisms became especially relevant in PI, although the microarray expression profiling validated by qPCR indicated that these functional changes are gradual from anterior to posterior intestinal segments. This functional divergence occurred in association with spatial transcriptional changes in nutrient transporters and the mucosal chemosensing system via G protein-coupled receptors. These findings contribute to identify key indicators of gut functions and to compare different fish feeding strategies and immune defense mechanisms acquired along the evolution of teleosts. PMID:27610085

Gene Expression Profiling Reveals Functional Specialization along the Intestinal Tract of a Carnivorous Teleostean Fish (Dicentrarchus labrax).

PubMed

Calduch-Giner, Josep A; Sitjà-Bobadilla, Ariadna; Pérez-Sánchez, Jaume

2016-01-01

High-quality sequencing reads from the intestine of European sea bass were assembled, annotated by similarity against protein reference databases and combined with nucleotide sequences from public and private databases. After redundancy filtering, 24,906 non-redundant annotated sequences encoding 15,367 different gene descriptions were obtained. These annotated sequences were used to design a custom, high-density oligo-microarray (8 × 15 K) for the transcriptomic profiling of anterior (AI), middle (MI), and posterior (PI) intestinal segments. Similar molecular signatures were found for AI and MI segments, which were combined in a single group (AI-MI) whereas the PI outstood separately, with more than 1900 differentially expressed genes with a fold-change cutoff of 2. Functional analysis revealed that molecular and cellular functions related to feed digestion and nutrient absorption and transport were over-represented in AI-MI segments. By contrast, the initiation and establishment of immune defense mechanisms became especially relevant in PI, although the microarray expression profiling validated by qPCR indicated that these functional changes are gradual from anterior to posterior intestinal segments. This functional divergence occurred in association with spatial transcriptional changes in nutrient transporters and the mucosal chemosensing system via G protein-coupled receptors. These findings contribute to identify key indicators of gut functions and to compare different fish feeding strategies and immune defense mechanisms acquired along the evolution of teleosts.

Variation of the chemical reactivity of Thermus thermophilus HB8 ribosomal proteins as a function of pH.

PubMed

Running, William E; Reilly, James P

2010-10-01

Ribosomes occupy a central position in cellular metabolism, converting stored genetic information into active cellular machinery. Ribosomal proteins modulate both the intrinsic function of the ribosome and its interaction with other cellular complexes, such as chaperonins or the signal recognition particle. Chemical modification of proteins combined with mass spectrometric detection of the extent and position of covalent modifications is a rapid, sensitive method for the study of protein structure and flexibility. By altering the pH of the solution, we have induced non-denaturing changes in the structure of bacterial ribosomal proteins and detected these conformational changes by covalent labeling. Changes in ribosomal protein modification across a pH range from 6.6 to 8.3 are unique to each protein, and correlate with their structural environment in the ribosome. Lysine residues whose extent of modification increases as a function of increasing pH are on the surface of proteins, but in close proximity either to glutamate and aspartate residues, or to rRNA backbone phosphates. Increasing pH disrupts tertiary and quaternary interactions mediated by hydrogen bonding or ionic interactions, and regions of protein structure whose conformations are sensitive to these changes are of potential importance in modulating the flexibility of the ribosome or its interaction with other cellular complexes.

Metaproteome analysis of endodontic infections in association with different clinical conditions.

PubMed

Provenzano, José Claudio; Siqueira, José F; Rôças, Isabela N; Domingues, Romênia R; Paes Leme, Adriana F; Silva, Márcia R S

2013-01-01

Analysis of the metaproteome of microbial communities is important to provide an insight of community physiology and pathogenicity. This study evaluated the metaproteome of endodontic infections associated with acute apical abscesses and asymptomatic apical periodontitis lesions. Proteins persisting or expressed after root canal treatment were also evaluated. Finally, human proteins associated with these infections were identified. Samples were taken from root canals of teeth with asymptomatic apical periodontitis before and after chemomechanical treatment using either NaOCl or chlorhexidine as the irrigant. Samples from abscesses were taken by aspiration of the purulent exudate. Clinical samples were processed for analysis of the exoproteome by using two complementary mass spectrometry platforms: nanoflow liquid chromatography coupled with linear ion trap quadrupole Velos Orbitrap and liquid chromatography-quadrupole time-of-flight. A total of 308 proteins of microbial origin were identified. The number of proteins in abscesses was higher than in asymptomatic cases. In canals irrigated with chlorhexidine, the number of identified proteins decreased substantially, while in the NaOCl group the number of proteins increased. The large majority of microbial proteins found in endodontic samples were related to metabolic and housekeeping processes, including protein synthesis, energy metabolism and DNA processes. Moreover, several other proteins related to pathogenicity and resistance/survival were found, including proteins involved with adhesion, biofilm formation and antibiotic resistance, stress proteins, exotoxins, invasins, proteases and endopeptidases (mostly in abscesses), and an archaeal protein linked to methane production. The majority of human proteins detected were related to cellular processes and metabolism, as well as immune defense. Interrogation of the metaproteome of endodontic microbial communities provides information on the physiology and pathogenicity of the community at the time of sampling. There is a growing need for expanded and more curated protein databases that permit more accurate identifications of proteins in metaproteomic studies.

A Circular Dichroism Reference Database for Membrane Proteins

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wallace,B.; Wien, F.; Stone, T.

2006-01-01

Membrane proteins are a major product of most genomes and the target of a large number of current pharmaceuticals, yet little information exists on their structures because of the difficulty of crystallising them; hence for the most part they have been excluded from structural genomics programme targets. Furthermore, even methods such as circular dichroism (CD) spectroscopy which seek to define secondary structure have not been fully exploited because of technical limitations to their interpretation for membrane embedded proteins. Empirical analyses of circular dichroism (CD) spectra are valuable for providing information on secondary structures of proteins. However, the accuracy of themore » results depends on the appropriateness of the reference databases used in the analyses. Membrane proteins have different spectral characteristics than do soluble proteins as a result of the low dielectric constants of membrane bilayers relative to those of aqueous solutions (Chen & Wallace (1997) Biophys. Chem. 65:65-74). To date, no CD reference database exists exclusively for the analysis of membrane proteins, and hence empirical analyses based on current reference databases derived from soluble proteins are not adequate for accurate analyses of membrane protein secondary structures (Wallace et al (2003) Prot. Sci. 12:875-884). We have therefore created a new reference database of CD spectra of integral membrane proteins whose crystal structures have been determined. To date it contains more than 20 proteins, and spans the range of secondary structures from mostly helical to mostly sheet proteins. This reference database should enable more accurate secondary structure determinations of membrane embedded proteins and will become one of the reference database options in the CD calculation server DICHROWEB (Whitmore & Wallace (2004) NAR 32:W668-673).« less

Tandem mass spectrometry for the detection of plant pathogenic fungi and the effects of database composition on protein inferences.

PubMed

Padliya, Neerav D; Garrett, Wesley M; Campbell, Kimberly B; Tabb, David L; Cooper, Bret

2007-11-01

LC-MS/MS has demonstrated potential for detecting plant pathogens. Unlike PCR or ELISA, LC-MS/MS does not require pathogen-specific reagents for the detection of pathogen-specific proteins and peptides. However, the MS/MS approach we and others have explored does require a protein sequence reference database and database-search software to interpret tandem mass spectra. To evaluate the limitations of database composition on pathogen identification, we analyzed proteins from cultured Ustilago maydis, Phytophthora sojae, Fusarium graminearum, and Rhizoctonia solani by LC-MS/MS. When the search database did not contain sequences for a target pathogen, or contained sequences to related pathogens, target pathogen spectra were reliably matched to protein sequences from nontarget organisms, giving an illusion that proteins from nontarget organisms were identified. Our analysis demonstrates that when database-search software is used as part of the identification process, a paradox exists whereby additional sequences needed to detect a wide variety of possible organisms may lead to more cross-species protein matches and misidentification of pathogens.

BIOZON: a system for unification, management and analysis of heterogeneous biological data.

PubMed

Birkland, Aaron; Yona, Golan

2006-02-15

Integration of heterogeneous data types is a challenging problem, especially in biology, where the number of databases and data types increase rapidly. Amongst the problems that one has to face are integrity, consistency, redundancy, connectivity, expressiveness and updatability. Here we present a system (Biozon) that addresses these problems, and offers biologists a new knowledge resource to navigate through and explore. Biozon unifies multiple biological databases consisting of a variety of data types (such as DNA sequences, proteins, interactions and cellular pathways). It is fundamentally different from previous efforts as it uses a single extensive and tightly connected graph schema wrapped with hierarchical ontology of documents and relations. Beyond warehousing existing data, Biozon computes and stores novel derived data, such as similarity relationships and functional predictions. The integration of similarity data allows propagation of knowledge through inference and fuzzy searches. Sophisticated methods of query that span multiple data types were implemented and first-of-a-kind biological ranking systems were explored and integrated. The Biozon system is an extensive knowledge resource of heterogeneous biological data. Currently, it holds more than 100 million biological documents and 6.5 billion relations between them. The database is accessible through an advanced web interface that supports complex queries, "fuzzy" searches, data materialization and more, online at http://biozon.org.

Network portal: a database for storage, analysis and visualization of biological networks

PubMed Central

Turkarslan, Serdar; Wurtmann, Elisabeth J.; Wu, Wei-Ju; Jiang, Ning; Bare, J. Christopher; Foley, Karen; Reiss, David J.; Novichkov, Pavel; Baliga, Nitin S.

2014-01-01

The ease of generating high-throughput data has enabled investigations into organismal complexity at the systems level through the inference of networks of interactions among the various cellular components (genes, RNAs, proteins and metabolites). The wider scientific community, however, currently has limited access to tools for network inference, visualization and analysis because these tasks often require advanced computational knowledge and expensive computing resources. We have designed the network portal (http://networks.systemsbiology.net) to serve as a modular database for the integration of user uploaded and public data, with inference algorithms and tools for the storage, visualization and analysis of biological networks. The portal is fully integrated into the Gaggle framework to seamlessly exchange data with desktop and web applications and to allow the user to create, save and modify workspaces, and it includes social networking capabilities for collaborative projects. While the current release of the database contains networks for 13 prokaryotic organisms from diverse phylogenetic clades (4678 co-regulated gene modules, 3466 regulators and 9291 cis-regulatory motifs), it will be rapidly populated with prokaryotic and eukaryotic organisms as relevant data become available in public repositories and through user input. The modular architecture, simple data formats and open API support community development of the portal. PMID:24271392

UUCD: a family-based database of ubiquitin and ubiquitin-like conjugation.

PubMed

Gao, Tianshun; Liu, Zexian; Wang, Yongbo; Cheng, Han; Yang, Qing; Guo, Anyuan; Ren, Jian; Xue, Yu

2013-01-01

In this work, we developed a family-based database of UUCD (http://uucd.biocuckoo.org) for ubiquitin and ubiquitin-like conjugation, which is one of the most important post-translational modifications responsible for regulating a variety of cellular processes, through a similar E1 (ubiquitin-activating enzyme)-E2 (ubiquitin-conjugating enzyme)-E3 (ubiquitin-protein ligase) enzyme thioester cascade. Although extensive experimental efforts have been taken, an integrative data resource is still not available. From the scientific literature, 26 E1s, 105 E2s, 1003 E3s and 148 deubiquitination enzymes (DUBs) were collected and classified into 1, 3, 19 and 7 families, respectively. To computationally characterize potential enzymes in eukaryotes, we constructed 1, 1, 15 and 6 hidden Markov model (HMM) profiles for E1s, E2s, E3s and DUBs at the family level, separately. Moreover, the ortholog searches were conducted for E3 and DUB families without HMM profiles. Then the UUCD database was developed with 738 E1s, 2937 E2s, 46 631 E3s and 6647 DUBs of 70 eukaryotic species. The detailed annotations and classifications were also provided. The online service of UUCD was implemented in PHP + MySQL + JavaScript + Perl.

PROFESS: a PROtein Function, Evolution, Structure and Sequence database

PubMed Central

Triplet, Thomas; Shortridge, Matthew D.; Griep, Mark A.; Stark, Jaime L.; Powers, Robert; Revesz, Peter

2010-01-01

The proliferation of biological databases and the easy access enabled by the Internet is having a beneficial impact on biological sciences and transforming the way research is conducted. There are ∼1100 molecular biology databases dispersed throughout the Internet. To assist in the functional, structural and evolutionary analysis of the abundant number of novel proteins continually identified from whole-genome sequencing, we introduce the PROFESS (PROtein Function, Evolution, Structure and Sequence) database. Our database is designed to be versatile and expandable and will not confine analysis to a pre-existing set of data relationships. A fundamental component of this approach is the development of an intuitive query system that incorporates a variety of similarity functions capable of generating data relationships not conceived during the creation of the database. The utility of PROFESS is demonstrated by the analysis of the structural drift of homologous proteins and the identification of potential pancreatic cancer therapeutic targets based on the observation of protein–protein interaction networks. Database URL: http://cse.unl.edu/∼profess/ PMID:20624718

Preparation of BFV Gag antiserum and preliminary study on cellular distribution of BFV.

PubMed

Wang, Jian; Guo, Hong-yan; Jia, Rui; Xu, Xuan; Tan, Juan; Geng, Yun-qi; Qiao, Wen-tao

2010-04-01

Viruses (e.g. Human immunodeficiency virus, Human simplex virus and Prototype foamy virus) are obligate intracellular parasites and therefore depend on the cellular machinery for cellular trafficking. Bovine foamy virus (BFV) is a member of the Spumaretrovirinae subfamily of Retroviruses, however, details of its cellular trafficking remain unknown. In this study, we cloned the BFV gag gene into prokaryotic expression vector pET28a and purified the denaturalized Gag protein. The protein was used to immunize BALB/c mouse to produce antiserum, which could specifically recognize the BFV Gag protein in BFV-infected cells through western blot assay. Additionally, these results demonstrated that both the optimal and suboptimal cleavage of Gag protein occur in BFV-infected cells. Subsequently, the Gag antiserum was used to investigate subcellular localization of BFV. In immunofluorescence microscopy assays, colocalization microtubules (MTs) and assembling viral particles were clearly observed, which implied that BFV may transport along cellular MTs in host cells. Furthermore, MTs-depolymerizing assay indicated MTs were required for the efficient replication of BFV. In conclusion, our study suggests that BFV has evolved the mechanism to hijack the cellular cytoskeleton for its replication.

SynechoNET: integrated protein-protein interaction database of a model cyanobacterium Synechocystis sp. PCC 6803.

PubMed

Kim, Woo-Yeon; Kang, Sungsoo; Kim, Byoung-Chul; Oh, Jeehyun; Cho, Seongwoong; Bhak, Jong; Choi, Jong-Soon

2008-01-01

Cyanobacteria are model organisms for studying photosynthesis, carbon and nitrogen assimilation, evolution of plant plastids, and adaptability to environmental stresses. Despite many studies on cyanobacteria, there is no web-based database of their regulatory and signaling protein-protein interaction networks to date. We report a database and website SynechoNET that provides predicted protein-protein interactions. SynechoNET shows cyanobacterial domain-domain interactions as well as their protein-level interactions using the model cyanobacterium, Synechocystis sp. PCC 6803. It predicts the protein-protein interactions using public interaction databases that contain mutually complementary and redundant data. Furthermore, SynechoNET provides information on transmembrane topology, signal peptide, and domain structure in order to support the analysis of regulatory membrane proteins. Such biological information can be queried and visualized in user-friendly web interfaces that include the interactive network viewer and search pages by keyword and functional category. SynechoNET is an integrated protein-protein interaction database designed to analyze regulatory membrane proteins in cyanobacteria. It provides a platform for biologists to extend the genomic data of cyanobacteria by predicting interaction partners, membrane association, and membrane topology of Synechocystis proteins. SynechoNET is freely available at http://synechocystis.org/ or directly at http://bioportal.kobic.kr/SynechoNET/.

PACSY, a relational database management system for protein structure and chemical shift analysis.

PubMed

Lee, Woonghee; Yu, Wookyung; Kim, Suhkmann; Chang, Iksoo; Lee, Weontae; Markley, John L

2012-10-01

PACSY (Protein structure And Chemical Shift NMR spectroscopY) is a relational database management system that integrates information from the Protein Data Bank, the Biological Magnetic Resonance Data Bank, and the Structural Classification of Proteins database. PACSY provides three-dimensional coordinates and chemical shifts of atoms along with derived information such as torsion angles, solvent accessible surface areas, and hydrophobicity scales. PACSY consists of six relational table types linked to one another for coherence by key identification numbers. Database queries are enabled by advanced search functions supported by an RDBMS server such as MySQL or PostgreSQL. PACSY enables users to search for combinations of information from different database sources in support of their research. Two software packages, PACSY Maker for database creation and PACSY Analyzer for database analysis, are available from http://pacsy.nmrfam.wisc.edu.

MIPS: a database for protein sequences and complete genomes.

PubMed Central

Mewes, H W; Hani, J; Pfeiffer, F; Frishman, D

1998-01-01

The MIPS group [Munich Information Center for Protein Sequences of the German National Center for Environment and Health (GSF)] at the Max-Planck-Institute for Biochemistry, Martinsried near Munich, Germany, is involved in a number of data collection activities, including a comprehensive database of the yeast genome, a database reflecting the progress in sequencing the Arabidopsis thaliana genome, the systematic analysis of other small genomes and the collection of protein sequence data within the framework of the PIR-International Protein Sequence Database (described elsewhere in this volume). Through its WWW server (http://www.mips.biochem.mpg.de ) MIPS provides access to a variety of generic databases, including a database of protein families as well as automatically generated data by the systematic application of sequence analysis algorithms. The yeast genome sequence and its related information was also compiled on CD-ROM to provide dynamic interactive access to the 16 chromosomes of the first eukaryotic genome unraveled. PMID:9399795

Protein Structure in Context: The Molecular Landscape of Angiogenesis

ERIC Educational Resources Information Center

Span, Elise A.; Goodsell, David S.; Ramchandran, Ramani; Franzen, Margaret A.; Herman, Tim; Sem, Daniel S.

2013-01-01

A team of students, educators, and researchers has developed new materials to teach cell signaling within its cellular context. Two nontraditional modalities are employed: physical models, to explore the atomic details of several of the proteins in the angiogenesis signaling cascade, and illustrations of the proteins in their cellular environment,…

ARCPHdb: A comprehensive protein database for SF1 and SF2 helicase from archaea.

PubMed

Moukhtar, Mirna; Chaar, Wafi; Abdel-Razzak, Ziad; Khalil, Mohamad; Taha, Samir; Chamieh, Hala

2017-01-01

Superfamily 1 and Superfamily 2 helicases, two of the largest helicase protein families, play vital roles in many biological processes including replication, transcription and translation. Study of helicase proteins in the model microorganisms of archaea have largely contributed to the understanding of their function, architecture and assembly. Based on a large phylogenomics approach, we have identified and classified all SF1 and SF2 protein families in ninety five sequenced archaea genomes. Here we developed an online webserver linked to a specialized protein database named ARCPHdb to provide access for SF1 and SF2 helicase families from archaea. ARCPHdb was implemented using MySQL relational database. Web interfaces were developed using Netbeans. Data were stored according to UniProt accession numbers, NCBI Ref Seq ID, PDB IDs and Entrez Databases. A user-friendly interactive web interface has been developed to browse, search and download archaeal helicase protein sequences, their available 3D structure models, and related documentation available in the literature provided by ARCPHdb. The database provides direct links to matching external databases. The ARCPHdb is the first online database to compile all protein information on SF1 and SF2 helicase from archaea in one platform. This database provides essential resource information for all researchers interested in the field. Copyright © 2016 Elsevier Ltd. All rights reserved.

Rice proteome database: a step toward functional analysis of the rice genome.

PubMed

Komatsu, Setsuko

2005-09-01

The technique of proteome analysis using two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) has the power to monitor global changes that occur in the protein complement of tissues and subcellular compartments. In this study, the proteins of rice were cataloged, a rice proteome database was constructed, and a functional characterization of some of the identified proteins was undertaken. Proteins extracted from various tissues and subcellular compartments in rice were separated by 2D-PAGE and an image analyzer was used to construct a display of the proteins. The Rice Proteome Database contains 23 reference maps based on 2D-PAGE of proteins from various rice tissues and subcellular compartments. These reference maps comprise 13129 identified proteins, and the amino acid sequences of 5092 proteins are entered in the database. Major proteins involved in growth or stress responses were identified using the proteome approach. Some of these proteins, including a beta-tubulin, calreticulin, and ribulose-1,5-bisphosphate carboxylase/oxygenase activase in rice, have unexpected functions. The information obtained from the Rice Proteome Database will aid in cloning the genes for and predicting the function of unknown proteins.

[Expression of Dengue virus type 2 nonstructural protein 3 and isolation of host proteins interacting with it].

PubMed

Weng, Daihui; Lei, Yingfeng; Dong, Yangchao; Han, Peijun; Ye, Chuantao; Yang, Jing; Wang, Yuan; Yin, Wen

2015-12-01

To construct the plasmid expressing the fusion protein of Dengue virus type 2 (DENV2) nonstructural protein 3 (NS3) with affinity tag, and isolate the cellular proteins interacting with NS3 protein using tandem affinity purification (TAP) assay. Primers for amplifying NS3 gene were designed according to the sequence of DENV2 genome and chemically synthesized. The NS3 fragments, after amplified by PCR with DENV2 cDNA as template, were digested and cloned into the mammalian eukaryotic expression vector pCI-SF with the tandem affinity tag (FLAG-StrepII). The recombinant pCI-NS3-SF was transiently transformed by Lipofectamine(TM) 2000 into HEK293T cells, and the expression of the fusion protein was confirmed by Western blotting. Cellular proteins that interacted with NS3 were isolated and purified by TAP assay. The eukaryotic expression vector expressing NS3 protein was successfully constructed. The host proteins interacting with NS3 protein were isolated by TAP system. TAP is an efficient method to isolate the cellular proteins interacting with DENV2 NS3.

A calmodulin binding protein from Arabidopsis is induced by ethylene and contains a DNA-binding motif

NASA Technical Reports Server (NTRS)

Reddy, A. S.; Reddy, V. S.; Golovkin, M.

2000-01-01

Calmodulin (CaM), a key calcium sensor in all eukaryotes, regulates diverse cellular processes by interacting with other proteins. To isolate CaM binding proteins involved in ethylene signal transduction, we screened an expression library prepared from ethylene-treated Arabidopsis seedlings with 35S-labeled CaM. A cDNA clone, EICBP (Ethylene-Induced CaM Binding Protein), encoding a protein that interacts with activated CaM was isolated in this screening. The CaM binding domain in EICBP was mapped to the C-terminus of the protein. These results indicate that calcium, through CaM, could regulate the activity of EICBP. The EICBP is expressed in different tissues and its expression in seedlings is induced by ethylene. The EICBP contains, in addition to a CaM binding domain, several features that are typical of transcription factors. These include a DNA-binding domain at the N terminus, an acidic region at the C terminus, and nuclear localization signals. In database searches a partial cDNA (CG-1) encoding a DNA-binding motif from parsley and an ethylene up-regulated partial cDNA from tomato (ER66) showed significant similarity to EICBP. In addition, five hypothetical proteins in the Arabidopsis genome also showed a very high sequence similarity with EICBP, indicating that there are several EICBP-related proteins in Arabidopsis. The structural features of EICBP are conserved in all EICBP-related proteins in Arabidopsis, suggesting that they may constitute a new family of DNA binding proteins and are likely to be involved in modulating gene expression in the presence of ethylene.

Biomolecular interactions modulate macromolecular structure and dynamics in atomistic model of a bacterial cytoplasm.

PubMed

Yu, Isseki; Mori, Takaharu; Ando, Tadashi; Harada, Ryuhei; Jung, Jaewoon; Sugita, Yuji; Feig, Michael

2016-11-01

Biological macromolecules function in highly crowded cellular environments. The structure and dynamics of proteins and nucleic acids are well characterized in vitro, but in vivo crowding effects remain unclear. Using molecular dynamics simulations of a comprehensive atomistic model cytoplasm we found that protein-protein interactions may destabilize native protein structures, whereas metabolite interactions may induce more compact states due to electrostatic screening. Protein-protein interactions also resulted in significant variations in reduced macromolecular diffusion under crowded conditions, while metabolites exhibited significant two-dimensional surface diffusion and altered protein-ligand binding that may reduce the effective concentration of metabolites and ligands in vivo. Metabolic enzymes showed weak non-specific association in cellular environments attributed to solvation and entropic effects. These effects are expected to have broad implications for the in vivo functioning of biomolecules. This work is a first step towards physically realistic in silico whole-cell models that connect molecular with cellular biology.

Dual Coordination of Post Translational Modifications in Human Protein Networks

PubMed Central

Woodsmith, Jonathan; Kamburov, Atanas; Stelzl, Ulrich

2013-01-01

Post-translational modifications (PTMs) regulate protein activity, stability and interaction profiles and are critical for cellular functioning. Further regulation is gained through PTM interplay whereby modifications modulate the occurrence of other PTMs or act in combination. Integration of global acetylation, ubiquitination and tyrosine or serine/threonine phosphorylation datasets with protein interaction data identified hundreds of protein complexes that selectively accumulate each PTM, indicating coordinated targeting of specific molecular functions. A second layer of PTM coordination exists in these complexes, mediated by PTM integration (PTMi) spots. PTMi spots represent very dense modification patterns in disordered protein regions and showed an equally high mutation rate as functional protein domains in cancer, inferring equivocal importance for cellular functioning. Systematic PTMi spot identification highlighted more than 300 candidate proteins for combinatorial PTM regulation. This study reveals two global PTM coordination mechanisms and emphasizes dataset integration as requisite in proteomic PTM studies to better predict modification impact on cellular signaling. PMID:23505349

Cellular Strategies for Regulating Functional and Nonfunctional Protein Aggregation

PubMed Central

Gsponer, Jörg; Babu, M. Madan

2012-01-01

Summary Growing evidence suggests that aggregation-prone proteins are both harmful and functional for a cell. How do cellular systems balance the detrimental and beneficial effect of protein aggregation? We reveal that aggregation-prone proteins are subject to differential transcriptional, translational, and degradation control compared to nonaggregation-prone proteins, which leads to their decreased synthesis, low abundance, and high turnover. Genetic modulators that enhance the aggregation phenotype are enriched in genes that influence expression homeostasis. Moreover, genes encoding aggregation-prone proteins are more likely to be harmful when overexpressed. The trends are evolutionarily conserved and suggest a strategy whereby cellular mechanisms specifically modulate the availability of aggregation-prone proteins to (1) keep concentrations below the critical ones required for aggregation and (2) shift the equilibrium between the monomeric and oligomeric/aggregate form, as explained by Le Chatelier’s principle. This strategy may prevent formation of undesirable aggregates and keep functional assemblies/aggregates under control. PMID:23168257

In silico analysis of fragile histidine triad involved in regression of carcinoma.

PubMed

Rasheed, Muhammad Asif; Tariq, Fatima; Afzal, Sara; Mannanv, Shazia

2017-04-01

Hepatocellular carcinoma (HCCa) is a primary malignancy of the liver. Many different proteins are involved in HCCa including insulin growth factor (IGF) II , signal transducers and activators of transcription (STAT) 3, STAT4, mothers against decapentaplegic homolog 4 (SMAD 4), fragile histidine triad (FHIT) and selective internal radiation therapy (SIRT) etc. The present study is based on the bioinformatics analysis of FHIT protein in order to understand the proteomics aspect and improvement of the diagnosis of the disease based on the protein. Different information related to protein were gathered from different databases, including National Centre for Biotechnology Information (NCBI) Gene, Protein and Online Mendelian Inheritance in Man (OMIM) databases, Uniprot database, String database and Kyoto Encyclopedia of Genes and Genomes (KEGG) database. Moreover, the structure of the protein and evaluation of the quality of the structure were included from Easy modeler programme. Hence, this analysis not only helped to gather information related to the protein at one place, but also analysed the structure and quality of the protein to conclude that the protein has a role in carcinoma.

Cyclosporin A associated helicase-like protein facilitates the association of hepatitis C virus RNA polymerase with its cellular cyclophilin B.

PubMed

Morohashi, Kengo; Sahara, Hiroeki; Watashi, Koichi; Iwabata, Kazuki; Sunoki, Takashi; Kuramochi, Kouji; Takakusagi, Kaori; Miyashita, Hiroki; Sato, Noriyuki; Tanabe, Atsushi; Shimotohno, Kunitada; Kobayashi, Susumu; Sakaguchi, Kengo; Sugawara, Fumio

2011-04-29

Cyclosporin A (CsA) is well known as an immunosuppressive drug useful for allogeneic transplantation. It has been reported that CsA inhibits hepatitis C virus (HCV) genome replication, which indicates that cellular targets of CsA regulate the viral replication. However, the regulation mechanisms of HCV replication governed by CsA target proteins have not been fully understood. Here we show a chemical biology approach that elucidates a novel mechanism of HCV replication. We developed a phage display screening to investigate compound-peptide interaction and identified a novel cellular target molecule of CsA. This protein, named CsA associated helicase-like protein (CAHL), possessed RNA-dependent ATPase activity that was negated by treatment with CsA. The downregulation of CAHL in the cells resulted in a decrease of HCV genome replication. CAHL formed a complex with HCV-derived RNA polymerase NS5B and host-derived cyclophilin B (CyPB), known as a cellular cofactor for HCV replication, to regulate NS5B-CyPB interaction. We found a cellular factor, CAHL, as CsA associated helicase-like protein, which would form trimer complex with CyPB and NS5B of HCV. The strategy using a chemical compound and identifying its target molecule by our phage display analysis is useful to reveal a novel mechanism underlying cellular and viral physiology.

RBFOX2 protein domains and cellular activities.

PubMed

Arya, Anurada D; Wilson, David I; Baralle, Diana; Raponi, Michaela

2014-08-01

RBFOX2 (RNA-binding protein, Fox-1 homologue 2)/RBM9 (RNA-binding-motif protein 9)/RTA (repressor of tamoxifen action)/HNRBP2 (hexaribonucleotide-binding protein 2) encodes an RNA-binding protein involved in tissue specific alternative splicing regulation and steroid receptors transcriptional activity. Its ability to regulate specific splicing profiles depending on context has been related to different expression levels of the RBFOX2 protein itself and that of other splicing regulatory proteins involved in the shared modulation of specific genes splicing. However, this cannot be the sole explanation as to why RBFOX2 plays a widespread role in numerous cellular mechanisms from development to cell survival dependent on cell/tissue type. RBFOX2 isoforms with altered protein domains exist. In the present article, we describe the main RBFOX2 protein domains, their importance in the context of splicing and transcriptional regulation and we propose that RBFOX2 isoform distribution may play a fundamental role in RBFOX2-specific cellular effects.

Reconstituting protein interaction networks using parameter-dependent domain-domain interactions

PubMed Central

2013-01-01

Background We can describe protein-protein interactions (PPIs) as sets of distinct domain-domain interactions (DDIs) that mediate the physical interactions between proteins. Experimental data confirm that DDIs are more consistent than their corresponding PPIs, lending support to the notion that analyses of DDIs may improve our understanding of PPIs and lead to further insights into cellular function, disease, and evolution. However, currently available experimental DDI data cover only a small fraction of all existing PPIs and, in the absence of structural data, determining which particular DDI mediates any given PPI is a challenge. Results We present two contributions to the field of domain interaction analysis. First, we introduce a novel computational strategy to merge domain annotation data from multiple databases. We show that when we merged yeast domain annotations from six annotation databases we increased the average number of domains per protein from 1.05 to 2.44, bringing it closer to the estimated average value of 3. Second, we introduce a novel computational method, parameter-dependent DDI selection (PADDS), which, given a set of PPIs, extracts a small set of domain pairs that can reconstruct the original set of protein interactions, while attempting to minimize false positives. Based on a set of PPIs from multiple organisms, our method extracted 27% more experimentally detected DDIs than existing computational approaches. Conclusions We have provided a method to merge domain annotation data from multiple sources, ensuring large and consistent domain annotation for any given organism. Moreover, we provided a method to extract a small set of DDIs from the underlying set of PPIs and we showed that, in contrast to existing approaches, our method was not biased towards DDIs with low or high occurrence counts. Finally, we used these two methods to highlight the influence of the underlying annotation density on the characteristics of extracted DDIs. Although increased annotations greatly expanded the possible DDIs, the lack of knowledge of the true biological false positive interactions still prevents an unambiguous assignment of domain interactions responsible for all protein network interactions. Executable files and examples are given at: http://www.bhsai.org/downloads/padds/ PMID:23651452

Selective recruitment of nuclear factors to productively replicating herpes simplex virus genomes.

PubMed

Dembowski, Jill A; DeLuca, Neal A

2015-05-01

Much of the HSV-1 life cycle is carried out in the cell nucleus, including the expression, replication, repair, and packaging of viral genomes. Viral proteins, as well as cellular factors, play essential roles in these processes. Isolation of proteins on nascent DNA (iPOND) was developed to label and purify cellular replication forks. We adapted aspects of this method to label viral genomes to both image, and purify replicating HSV-1 genomes for the identification of associated proteins. Many viral and cellular factors were enriched on viral genomes, including factors that mediate DNA replication, repair, chromatin remodeling, transcription, and RNA processing. As infection proceeded, packaging and structural components were enriched to a greater extent. Among the more abundant proteins that copurified with genomes were the viral transcription factor ICP4 and the replication protein ICP8. Furthermore, all seven viral replication proteins were enriched on viral genomes, along with cellular PCNA and topoisomerases, while other cellular replication proteins were not detected. The chromatin-remodeling complexes present on viral genomes included the INO80, SWI/SNF, NURD, and FACT complexes, which may prevent chromatinization of the genome. Consistent with this conclusion, histones were not readily recovered with purified viral genomes, and imaging studies revealed an underrepresentation of histones on viral genomes. RNA polymerase II, the mediator complex, TFIID, TFIIH, and several other transcriptional activators and repressors were also affinity purified with viral DNA. The presence of INO80, NURD, SWI/SNF, mediator, TFIID, and TFIIH components is consistent with previous studies in which these complexes copurified with ICP4. Therefore, ICP4 is likely involved in the recruitment of these key cellular chromatin remodeling and transcription factors to viral genomes. Taken together, iPOND is a valuable method for the study of viral genome dynamics during infection and provides a comprehensive view of how HSV-1 selectively utilizes cellular resources.

Understanding the cancer cell phenotype beyond the limitations of current omics analyses.

PubMed

Moreno-Sánchez, Rafael; Saavedra, Emma; Gallardo-Pérez, Juan Carlos; Rumjanek, Franklin D; Rodríguez-Enríquez, Sara

2016-01-01

Efforts to understand the mechanistic principles driving cancer metabolism and proliferation have been lately governed by genomic, transcriptomic and proteomic studies. This paper analyzes the caveats of these approaches. As molecular biology's central dogma proposes a unidirectional flux of information from genes to mRNA to proteins, it has frequently been assumed that monitoring the changes in the gene sequences and in mRNA and protein contents is sufficient to explain complex cellular processes. Such a stance commonly disregards that post-translational modifications can alter the protein function/activity and also that regulatory mechanisms enter into action, to coordinate the protein activities of pathways/cellular processes, in order to keep the cellular homeostasis. Hence, the actual protein activities (as enzymes/transporters/receptors) and their regulatory mechanisms ultimately dictate the final outcomes of a pathway/cellular process. In this regard, it is here documented that the mRNA levels of many metabolic enzymes and transcriptional factors have no correlation with the respective protein contents and activities. The validity of current clinical mRNA-based tests and proposed metabolite biomarkers for cancer detection/prognosis is also discussed. Therefore, it is proposed that, to achieve a thorough understanding of the modifications undergone by proliferating cancer cells, it is mandatory to experimentally analyze the cellular processes at the functional level. This could be achieved (a) locally, by examining the actual protein activities in the cell and their kinetic properties (or at least kinetically characterize the most controlling steps of the pathway/cellular process); (b) systemically, by analyzing the main fluxes of the pathway/cellular process, and how they are modulated by metabolites, all which should contribute to comprehending the regulatory mechanisms that have been altered in cancer cells. By adopting a more holistic approach it may become possible to improve the design of therapeutic strategies that would target cancer cells more specifically. © 2015 FEBS.

Chlamydia trachomatis can protect host cells against apoptosis in the absence of cellular Inhibitor of Apoptosis Proteins and Mcl-1.

PubMed

Ying, Songmin; Christian, Jan G; Paschen, Stefan A; Häcker, Georg

2008-01-01

Infection with Chlamydia protects mammalian host cells against apoptosis. Hypotheses have been proposed to explain this molecularly, including the up-regulation of host anti-apoptotic proteins such as cellular Inhibitor of Apoptosis Protein (IAP) 2 and the Bcl-2 protein Mcl-1. To test for the importance of these proteins, we used mouse embryonic fibroblasts from gene-targeted mice that were deficient in cIAP1, cIAP2, cIAP1/cIAP2, XIAP, or Mcl-1. Infection with Chlamydia trachomatis protected all cells equally well against apoptosis, which was induced either with tumour necrosis factor/cycloheximide (IAP-knock-out cells) or staurosporine (Mcl-1-knock-out). Therefore, these cellular anti-apoptotic proteins are not essential for apoptosis-protection by C. trachomatis.

FunCoup 3.0: database of genome-wide functional coupling networks

PubMed Central

Schmitt, Thomas; Ogris, Christoph; Sonnhammer, Erik L. L.

2014-01-01

We present an update of the FunCoup database (http://FunCoup.sbc.su.se) of functional couplings, or functional associations, between genes and gene products. Identifying these functional couplings is an important step in the understanding of higher level mechanisms performed by complex cellular processes. FunCoup distinguishes between four classes of couplings: participation in the same signaling cascade, participation in the same metabolic process, co-membership in a protein complex and physical interaction. For each of these four classes, several types of experimental and statistical evidence are combined by Bayesian integration to predict genome-wide functional coupling networks. The FunCoup framework has been completely re-implemented to allow for more frequent future updates. It contains many improvements, such as a regularization procedure to automatically downweight redundant evidences and a novel method to incorporate phylogenetic profile similarity. Several datasets have been updated and new data have been added in FunCoup 3.0. Furthermore, we have developed a new Web site, which provides powerful tools to explore the predicted networks and to retrieve detailed information about the data underlying each prediction. PMID:24185702

FunCoup 3.0: database of genome-wide functional coupling networks.

PubMed

Schmitt, Thomas; Ogris, Christoph; Sonnhammer, Erik L L

2014-01-01

We present an update of the FunCoup database (http://FunCoup.sbc.su.se) of functional couplings, or functional associations, between genes and gene products. Identifying these functional couplings is an important step in the understanding of higher level mechanisms performed by complex cellular processes. FunCoup distinguishes between four classes of couplings: participation in the same signaling cascade, participation in the same metabolic process, co-membership in a protein complex and physical interaction. For each of these four classes, several types of experimental and statistical evidence are combined by Bayesian integration to predict genome-wide functional coupling networks. The FunCoup framework has been completely re-implemented to allow for more frequent future updates. It contains many improvements, such as a regularization procedure to automatically downweight redundant evidences and a novel method to incorporate phylogenetic profile similarity. Several datasets have been updated and new data have been added in FunCoup 3.0. Furthermore, we have developed a new Web site, which provides powerful tools to explore the predicted networks and to retrieve detailed information about the data underlying each prediction.

The Protein-DNA Interface database

PubMed Central

2010-01-01

The Protein-DNA Interface database (PDIdb) is a repository containing relevant structural information of Protein-DNA complexes solved by X-ray crystallography and available at the Protein Data Bank. The database includes a simple functional classification of the protein-DNA complexes that consists of three hierarchical levels: Class, Type and Subtype. This classification has been defined and manually curated by humans based on the information gathered from several sources that include PDB, PubMed, CATH, SCOP and COPS. The current version of the database contains only structures with resolution of 2.5 Å or higher, accounting for a total of 922 entries. The major aim of this database is to contribute to the understanding of the main rules that underlie the molecular recognition process between DNA and proteins. To this end, the database is focused on each specific atomic interface rather than on the separated binding partners. Therefore, each entry in this database consists of a single and independent protein-DNA interface. We hope that PDIdb will be useful to many researchers working in fields such as the prediction of transcription factor binding sites in DNA, the study of specificity determinants that mediate enzyme recognition events, engineering and design of new DNA binding proteins with distinct binding specificity and affinity, among others. Finally, due to its friendly and easy-to-use web interface, we hope that PDIdb will also serve educational and teaching purposes. PMID:20482798

The Protein-DNA Interface database.

PubMed

Norambuena, Tomás; Melo, Francisco

2010-05-18

The Protein-DNA Interface database (PDIdb) is a repository containing relevant structural information of Protein-DNA complexes solved by X-ray crystallography and available at the Protein Data Bank. The database includes a simple functional classification of the protein-DNA complexes that consists of three hierarchical levels: Class, Type and Subtype. This classification has been defined and manually curated by humans based on the information gathered from several sources that include PDB, PubMed, CATH, SCOP and COPS. The current version of the database contains only structures with resolution of 2.5 A or higher, accounting for a total of 922 entries. The major aim of this database is to contribute to the understanding of the main rules that underlie the molecular recognition process between DNA and proteins. To this end, the database is focused on each specific atomic interface rather than on the separated binding partners. Therefore, each entry in this database consists of a single and independent protein-DNA interface.We hope that PDIdb will be useful to many researchers working in fields such as the prediction of transcription factor binding sites in DNA, the study of specificity determinants that mediate enzyme recognition events, engineering and design of new DNA binding proteins with distinct binding specificity and affinity, among others. Finally, due to its friendly and easy-to-use web interface, we hope that PDIdb will also serve educational and teaching purposes.

The Universal Protein Resource (UniProt): an expanding universe of protein information.

PubMed

Wu, Cathy H; Apweiler, Rolf; Bairoch, Amos; Natale, Darren A; Barker, Winona C; Boeckmann, Brigitte; Ferro, Serenella; Gasteiger, Elisabeth; Huang, Hongzhan; Lopez, Rodrigo; Magrane, Michele; Martin, Maria J; Mazumder, Raja; O'Donovan, Claire; Redaschi, Nicole; Suzek, Baris

2006-01-01

The Universal Protein Resource (UniProt) provides a central resource on protein sequences and functional annotation with three database components, each addressing a key need in protein bioinformatics. The UniProt Knowledgebase (UniProtKB), comprising the manually annotated UniProtKB/Swiss-Prot section and the automatically annotated UniProtKB/TrEMBL section, is the preeminent storehouse of protein annotation. The extensive cross-references, functional and feature annotations and literature-based evidence attribution enable scientists to analyse proteins and query across databases. The UniProt Reference Clusters (UniRef) speed similarity searches via sequence space compression by merging sequences that are 100% (UniRef100), 90% (UniRef90) or 50% (UniRef50) identical. Finally, the UniProt Archive (UniParc) stores all publicly available protein sequences, containing the history of sequence data with links to the source databases. UniProt databases continue to grow in size and in availability of information. Recent and upcoming changes to database contents, formats, controlled vocabularies and services are described. New download availability includes all major releases of UniProtKB, sequence collections by taxonomic division and complete proteomes. A bibliography mapping service has been added, and an ID mapping service will be available soon. UniProt databases can be accessed online at http://www.uniprot.org or downloaded at ftp://ftp.uniprot.org/pub/databases/.

The identification of cellular targets of 17β estradiol using a lytic (T7) cDNA phage display approach.

PubMed

Van Dorst, Bieke; Mehta, Jaytry; Rouah-Martin, Elsa; De Coen, Wim; Blust, Ronny; Robbens, Johan

2011-02-01

To unravel the mechanism of action of chemical compounds, it is crucial to know their cellular targets. A novel in vitro tool that can be used as a fast, simple and cost effective alternative is cDNA phage display. This tool is used in our study to select cellular targets of 17β estradiol (E2). It was possible to select two potential cellular targets of E2 out of the T7 Select™ Human Breast cDNA phage library. The selected cellular targets, autophagy/beclin-1 regulator 1 (beclin 1) and ATP synthase F(0) subunit 6 (ATP6) have so far been unknown as binding proteins of E2. To confirm the E2 binding properties of these selected proteins, surface plasmon resonance (SPR) was used. With SPR the K(d) values were determined to be 0.178±0.031 and 0.401±0.142 nM for the ATP6 phage and beclin 1 phage, respectively. These K(d) values in the low nM range verify that the selected cellular proteins are indeed binding proteins for E2. The selection and identification of these two potential cellular targets of E2, can enhance our current understanding of its mechanism of action. This illustrates the potential of lytic (T7) cDNA phage display in toxicology, to provide important information about cellular targets of chemical compounds. Copyright © 2010 Elsevier Ltd. All rights reserved.

Domain fusion analysis by applying relational algebra to protein sequence and domain databases.

PubMed

Truong, Kevin; Ikura, Mitsuhiko

2003-05-06

Domain fusion analysis is a useful method to predict functionally linked proteins that may be involved in direct protein-protein interactions or in the same metabolic or signaling pathway. As separate domain databases like BLOCKS, PROSITE, Pfam, SMART, PRINTS-S, ProDom, TIGRFAMs, and amalgamated domain databases like InterPro continue to grow in size and quality, a computational method to perform domain fusion analysis that leverages on these efforts will become increasingly powerful. This paper proposes a computational method employing relational algebra to find domain fusions in protein sequence databases. The feasibility of this method was illustrated on the SWISS-PROT+TrEMBL sequence database using domain predictions from the Pfam HMM (hidden Markov model) database. We identified 235 and 189 putative functionally linked protein partners in H. sapiens and S. cerevisiae, respectively. From scientific literature, we were able to confirm many of these functional linkages, while the remainder offer testable experimental hypothesis. Results can be viewed at http://calcium.uhnres.utoronto.ca/pi. As the analysis can be computed quickly on any relational database that supports standard SQL (structured query language), it can be dynamically updated along with the sequence and domain databases, thereby improving the quality of predictions over time.

Protein Bioinformatics Databases and Resources

PubMed Central

Chen, Chuming; Huang, Hongzhan; Wu, Cathy H.

2017-01-01

Many publicly available data repositories and resources have been developed to support protein related information management, data-driven hypothesis generation and biological knowledge discovery. To help researchers quickly find the appropriate protein related informatics resources, we present a comprehensive review (with categorization and description) of major protein bioinformatics databases in this chapter. We also discuss the challenges and opportunities for developing next-generation protein bioinformatics databases and resources to support data integration and data analytics in the Big Data era. PMID:28150231

Expression of psoriasis-associated fatty acid-binding protein in senescent human dermal microvascular endothelial cells.

PubMed

Ha, Moon Kyung; Chung, Kee Yang; Lee, Ju Hee; Bang, Dongsik; Park, Yoon Kee; Lee, Kwang Hoon

2004-09-01

Aging is associated with the progressive pathophysiologic modification of endothelial cells. In vitro endothelial cell senescence is accompanied by proliferative activity failure and by perturbations in gene and protein expressions. Moreover, this cellular senescence in culture has been proposed to reflect processes that occur in aging organisms. In order to observe the changing patterns of protein expression in senescent human dermal microvascular endothelial cells (HDMECs), proteins obtained from both early- and late-passaged HDMECs were separated by two-dimensional electrophoresis, visualized by silver staining, and quantified by image processing. Proteins of interest were extracted by in-gel digestion with trypsin and quantified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS), by searching the National Center for Biotechnology Information protein-sequence database. More than 2000 spots were detected by 2D electrophoresis within a linear pH range of 3-10. Twenty-two major differentially expressed spots were observed in serially passaged HDMECs and identified with high confidence by MALDI-TOF-MS. One of these spots was found to be a 14-15 kDa psoriasis-associated fatty acid-binding protein (PA-FABP) with high affinity for long-chain fatty acids. The expression of PA-FABP was confirmed to be elevated in senescent HDMECs (passage 20) by fluorescence-activated cell sorting (FACS), confocal laser microscopy, and by immunohistochemistry in aged human skin tissue. Our results suggest that the overexpression of FABP in cultured senescent HDMECs is closely related to skin aging.

Transcriptomic insights on the ABC transporter gene family in the salmon louse Caligus rogercresseyi.

PubMed

Valenzuela-Muñoz, Valentina; Sturm, Armin; Gallardo-Escárate, Cristian

2015-04-09

ATP-binding cassette (ABC) protein family encode for membrane proteins involved in the transport of various biomolecules through the cellular membrane. These proteins have been identified in all taxa and present important physiological functions, including the process of insecticide detoxification in arthropods. For that reason the ectoparasite Caligus rogercresseyi represents a model species for understanding the molecular underpinnings involved in insecticide drug resistance. llumina sequencing was performed using sea lice exposed to 2 and 3 ppb of deltamethrin and azamethiphos. Contigs obtained from de novo assembly were annotated by Blastx. RNA-Seq analysis was performed and validated by qPCR analysis. From the transcriptome database of C. rogercresseyi, 57 putative members of ABC protein sequences were identified and phylogenetically classified into the eight subfamilies described for ABC transporters in arthropods. Transcriptomic profiles for ABC proteins subfamilies were evaluated throughout C. rogercresseyi development. Moreover, RNA-Seq analysis was performed for adult male and female salmon lice exposed to the delousing drugs azamethiphos and deltamethrin. High transcript levels of the ABCB and ABCC subfamilies were evidenced. Furthermore, SNPs mining was carried out for the ABC proteins sequences, revealing pivotal genomic information. The present study gives a comprehensive transcriptome analysis of ABC proteins from C. rogercresseyi, providing relevant information about transporter roles during ontogeny and in relation to delousing drug responses in salmon lice. This genomic information represents a valuable tool for pest management in the Chilean salmon aquaculture industry.

Modeling and simulating networks of interdependent protein interactions.

PubMed

Stöcker, Bianca K; Köster, Johannes; Zamir, Eli; Rahmann, Sven

2018-05-21

Protein interactions are fundamental building blocks of biochemical reaction systems underlying cellular functions. The complexity and functionality of these systems emerge not only from the protein interactions themselves but also from the dependencies between these interactions, as generated by allosteric effects or mutual exclusion due to steric hindrance. Therefore, formal models for integrating and utilizing information about interaction dependencies are of high interest. Here, we describe an approach for endowing protein networks with interaction dependencies using propositional logic, thereby obtaining constrained protein interaction networks ("constrained networks"). The construction of these networks is based on public interaction databases as well as text-mined information about interaction dependencies. We present an efficient data structure and algorithm to simulate protein complex formation in constrained networks. The efficiency of the model allows fast simulation and facilitates the analysis of many proteins in large networks. In addition, this approach enables the simulation of perturbation effects, such as knockout of single or multiple proteins and changes of protein concentrations. We illustrate how our model can be used to analyze a constrained human adhesome protein network, which is responsible for the formation of diverse and dynamic cell-matrix adhesion sites. By comparing protein complex formation under known interaction dependencies versus without dependencies, we investigate how these dependencies shape the resulting repertoire of protein complexes. Furthermore, our model enables investigating how the interplay of network topology with interaction dependencies influences the propagation of perturbation effects across a large biochemical system. Our simulation software CPINSim (for Constrained Protein Interaction Network Simulator) is available under the MIT license at http://github.com/BiancaStoecker/cpinsim and as a Bioconda package (https://bioconda.github.io).

PACSY, a relational database management system for protein structure and chemical shift analysis

PubMed Central

Lee, Woonghee; Yu, Wookyung; Kim, Suhkmann; Chang, Iksoo

2012-01-01

PACSY (Protein structure And Chemical Shift NMR spectroscopY) is a relational database management system that integrates information from the Protein Data Bank, the Biological Magnetic Resonance Data Bank, and the Structural Classification of Proteins database. PACSY provides three-dimensional coordinates and chemical shifts of atoms along with derived information such as torsion angles, solvent accessible surface areas, and hydrophobicity scales. PACSY consists of six relational table types linked to one another for coherence by key identification numbers. Database queries are enabled by advanced search functions supported by an RDBMS server such as MySQL or PostgreSQL. PACSY enables users to search for combinations of information from different database sources in support of their research. Two software packages, PACSY Maker for database creation and PACSY Analyzer for database analysis, are available from http://pacsy.nmrfam.wisc.edu. PMID:22903636

RNAimmuno: A database of the nonspecific immunological effects of RNA interference and microRNA reagents

PubMed Central

Olejniczak, Marta; Galka-Marciniak, Paulina; Polak, Katarzyna; Fligier, Andrzej; Krzyzosiak, Wlodzimierz J.

2012-01-01

The RNAimmuno database was created to provide easy access to information regarding the nonspecific effects generated in cells by RNA interference triggers and microRNA regulators. Various RNAi and microRNA reagents, which differ in length and structure, often cause non-sequence-specific immune responses, in addition to triggering the intended sequence-specific effects. The activation of the cellular sensors of foreign RNA or DNA may lead to the induction of type I interferon and proinflammatory cytokine release. Subsequent changes in the cellular transcriptome and proteome may result in adverse effects, including cell death during therapeutic treatments or the misinterpretation of experimental results in research applications. The manually curated RNAimmuno database gathers the majority of the published data regarding the immunological side effects that are caused in investigated cell lines, tissues, and model organisms by different reagents. The database is accessible at http://rnaimmuno.ibch.poznan.pl and may be helpful in the further application and development of RNAi- and microRNA-based technologies. PMID:22411954

RNAimmuno: a database of the nonspecific immunological effects of RNA interference and microRNA reagents.

PubMed

Olejniczak, Marta; Galka-Marciniak, Paulina; Polak, Katarzyna; Fligier, Andrzej; Krzyzosiak, Wlodzimierz J

2012-05-01

The RNAimmuno database was created to provide easy access to information regarding the nonspecific effects generated in cells by RNA interference triggers and microRNA regulators. Various RNAi and microRNA reagents, which differ in length and structure, often cause non-sequence-specific immune responses, in addition to triggering the intended sequence-specific effects. The activation of the cellular sensors of foreign RNA or DNA may lead to the induction of type I interferon and proinflammatory cytokine release. Subsequent changes in the cellular transcriptome and proteome may result in adverse effects, including cell death during therapeutic treatments or the misinterpretation of experimental results in research applications. The manually curated RNAimmuno database gathers the majority of the published data regarding the immunological side effects that are caused in investigated cell lines, tissues, and model organisms by different reagents. The database is accessible at http://rnaimmuno.ibch.poznan.pl and may be helpful in the further application and development of RNAi- and microRNA-based technologies.

A protein relational database and protein family knowledge bases to facilitate structure-based design analyses.

PubMed

Mobilio, Dominick; Walker, Gary; Brooijmans, Natasja; Nilakantan, Ramaswamy; Denny, R Aldrin; Dejoannis, Jason; Feyfant, Eric; Kowticwar, Rupesh K; Mankala, Jyoti; Palli, Satish; Punyamantula, Sairam; Tatipally, Maneesh; John, Reji K; Humblet, Christine

2010-08-01

The Protein Data Bank is the most comprehensive source of experimental macromolecular structures. It can, however, be difficult at times to locate relevant structures with the Protein Data Bank search interface. This is particularly true when searching for complexes containing specific interactions between protein and ligand atoms. Moreover, searching within a family of proteins can be tedious. For example, one cannot search for some conserved residue as residue numbers vary across structures. We describe herein three databases, Protein Relational Database, Kinase Knowledge Base, and Matrix Metalloproteinase Knowledge Base, containing protein structures from the Protein Data Bank. In Protein Relational Database, atom-atom distances between protein and ligand have been precalculated allowing for millisecond retrieval based on atom identity and distance constraints. Ring centroids, centroid-centroid and centroid-atom distances and angles have also been included permitting queries for pi-stacking interactions and other structural motifs involving rings. Other geometric features can be searched through the inclusion of residue pair and triplet distances. In Kinase Knowledge Base and Matrix Metalloproteinase Knowledge Base, the catalytic domains have been aligned into common residue numbering schemes. Thus, by searching across Protein Relational Database and Kinase Knowledge Base, one can easily retrieve structures wherein, for example, a ligand of interest is making contact with the gatekeeper residue.

Close the Textbook & Open "The Cell: An Image Library"

ERIC Educational Resources Information Center

Saunders, Cheston; Taylor, Amy

2014-01-01

Many students leave the biology classroom with misconceptions centered on cellular structure. This article presents an activity in which students utilize images from an online database called "The Cell: An Image Library" (http://www.cellimagelibrary. org/) to gain a greater understanding of the diversity of cellular structure and the…

Growth factors and myometrium: biological effects in uterine fibroid and possible clinical implications

PubMed Central

Ciarmela, Pasquapina; Islam, Md. Soriful; Reis, Fernando M.; Gray, Peter C.; Bloise, Enrrico; Petraglia, Felice; Vale, Wylie; Castellucci, Mario

2011-01-01

BACKGROUND Growth factors are proteins secreted by a number of cell types that are capable of modulating cellular growth, proliferation and cellular differentiation. It is well accepted that uterine cellular events such as proliferation and differentiation are regulated by sex steroids and their actions in target tissues are mediated by local production of growth factors acting through paracrine and/or autocrine mechanisms. Myometrial mass is ultimately modified in pregnancy as well as in tumour conditions such as leiomyoma and leiomyosarcoma. Leiomyomas, also known as fibroids, are benign tumours of the uterus, considered to be one of the most frequent causes of infertility in reproductive years in women. METHODS For this review, we searched the database MEDLINE and Google Scholar for articles with content related to growth factors acting on myometrium; the findings are hereby reviewed and discussed. RESULTS Different growth factors such as epidermal growth factor (EGF), transforming growth factor-α (TGF-α), heparin-binding EGF (HB-EGF), acidic fibroblast growth factor (aFGF), basic fibroblast growth factor (bFGF), vascular endothelial growth factor (VEGF), insulin-like growth factor (IGF), platelet-derived growth factor (PDGF) and TGF-β perform actions in myometrium and in leiomyomas. In addition to these growth factors, activin and myostatin have been recently identified in myometrium and leiomyoma. CONCLUSIONS Growth factors play an important role in the mechanisms involved in myometrial patho-physiology. PMID:21788281

Interaction between core protein of classical swine fever virus with cellular IQGAP1 proetin appears essential for virulence in swine

USDA-ARS?s Scientific Manuscript database

Here we show that IQGAP1, a cellular protein that plays a pivotal role as a regulator of the cytoskeleton affecting cell adhesion, polarization and migration, interacts with Classical Swine Fever Virus (CSFV) Core protein. Sequence analyses identified a defined set of residues within CSFV Core prote...

Differential Activation of Cellular DNA Damage Responses by Replication-Defective and Replication-Competent Adenovirus Mutants

PubMed Central

Prakash, Anand; Jayaram, Sumithra

2012-01-01

Adenovirus (Ad) mutants that lack early region 4 (E4) activate the phosphorylation of cellular DNA damage response proteins. In wild-type Ad type 5 (Ad5) infections, E1b and E4 proteins target the cellular DNA repair protein Mre11 for redistribution and degradation, thereby interfering with its ability to activate phosphorylation cascades important during DNA repair. The characteristics of Ad infection that activate cellular DNA repair processes are not yet well understood. We investigated the activation of DNA damage responses by a replication-defective Ad vector (AdRSVβgal) that lacks E1 and fails to produce the immediate-early E1a protein. E1a is important for activating early gene expression from the other viral early transcription units, including E4. AdRSVβgal can deliver its genome to the cell, but it is subsequently deficient for viral early gene expression and DNA replication. We studied the ability of AdRSVβgal-infected cells to induce cellular DNA damage responses. AdRSVβgal infection does activate formation of foci containing the Mdc1 protein. However, AdRSVβgal fails to activate phosphorylation of the damage response proteins Nbs1 and Chk1. We found that viral DNA replication is important for Nbs1 phosphorylation, suggesting that this step in the viral life cycle may provide an important trigger for activating at least some DNA repair proteins. PMID:23015708

Piezo proteins: regulators of mechanosensation and other cellular processes.

PubMed

Bagriantsev, Sviatoslav N; Gracheva, Elena O; Gallagher, Patrick G

2014-11-14

Piezo proteins have recently been identified as ion channels mediating mechanosensory transduction in mammalian cells. Characterization of these channels has yielded important insights into mechanisms of somatosensation, as well as other mechano-associated biologic processes such as sensing of shear stress, particularly in the vasculature, and regulation of urine flow and bladder distention. Other roles for Piezo proteins have emerged, some unexpected, including participation in cellular development, volume regulation, cellular migration, proliferation, and elongation. Mutations in human Piezo proteins have been associated with a variety of disorders including hereditary xerocytosis and several syndromes with muscular contracture as a prominent feature. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

In silico screening of small molecule libraries using the dengue virus envelope E protein has identified compounds with antiviral activity against multiple flaviviruses.

PubMed

Kampmann, Thorsten; Yennamalli, Ragothaman; Campbell, Phillipa; Stoermer, Martin J; Fairlie, David P; Kobe, Bostjan; Young, Paul R

2009-12-01

The flaviviruses comprise a large group of related viruses, many of which pose a significant global human health threat, most notably the dengue viruses (DENV), West Nile virus (WNV) and yellow fever virus (YFV). Flaviviruses enter host cells via fusion of the viral and cellular membranes, a process mediated by the major viral envelope protein E as it undergoes a low pH induced conformational change in the endosomal compartment of the host cell. This essential entry stage in the flavivirus life cycle provides an attractive target for the development of antiviral agents. We performed an in silico docking screen of the Maybridge chemical database within a previously described ligand binding pocket in the dengue E protein structure that is thought to play a key role in the conformational transitions that lead to membrane fusion. The biological activity of selected compounds identified from this screen revealed low micromolar antiviral potency against dengue virus for two of the compounds. Our results also provide the first evidence that compounds selected to bind to this ligand binding site on the flavivirus E protein abrogate fusion activity. Interestingly, one of these compounds also has antiviral activity against both WNV (kunjin strain) and YFV.

JASSA: a comprehensive tool for prediction of SUMOylation sites and SIMs.

PubMed

Beauclair, Guillaume; Bridier-Nahmias, Antoine; Zagury, Jean-François; Saïb, Ali; Zamborlini, Alessia

2015-11-01

Post-translational modification by the Small Ubiquitin-like Modifier (SUMO) proteins, a process termed SUMOylation, is involved in many fundamental cellular processes. SUMO proteins are conjugated to a protein substrate, creating an interface for the recruitment of cofactors harboring SUMO-interacting motifs (SIMs). Mapping both SUMO-conjugation sites and SIMs is required to study the functional consequence of SUMOylation. To define the best candidate sites for experimental validation we designed JASSA, a Joint Analyzer of SUMOylation site and SIMs. JASSA is a predictor that uses a scoring system based on a Position Frequency Matrix derived from the alignment of experimental SUMOylation sites or SIMs. Compared with existing web-tools, JASSA displays on par or better performances. Novel features were implemented towards a better evaluation of the prediction, including identification of database hits matching the query sequence and representation of candidate sites within the secondary structural elements and/or the 3D fold of the protein of interest, retrievable from deposited PDB files. JASSA is freely accessible at http://www.jassa.fr/. Website is implemented in PHP and MySQL, with all major browsers supported. guillaume.beauclair@inserm.fr Supplementary data are available at Bioinformatics online. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Proteomic study reveals a co-occurrence of gallic acid-induced apoptosis and glycolysis in B16F10 melanoma cells.

PubMed

Liu, Cheng; Lin, Jen-Jie; Yang, Zih-Yan; Tsai, Chi-Chu; Hsu, Jue-Liang; Wu, Yu-Jen

2014-12-03

Gallic acid (GA) has long been associated with a wide range of biological activities. In this study, its antitumor effect against B16F10 melanoma cells was demonstrated by MTT assay, cell migration assay, wound-healing assay, and flow cytometric analysis. GA with a concentration >200 μM shows apoptotic activity toward B16F10 cells. According to Western blotting data, overexpressions of cleaved forms of caspase-9, caspase-3, and PARP-1 and pro-apoptotic Bax and Bad, accompanied by underexpressed anti-apoptotic Bcl-2 and Bcl-xL indicate that GA induces B16F10 cell apoptosis via mitochondrial pathway. The 2-DE based comparative proteomics was further employed in B16F10 cells with and without GA treatment for a large-scale protein expression profiling. A total of 41 differential protein spots were quantified, and their identities were characterized using LC-MS/MS analysis and database matching. In addition to some regulated proteins that were associated with apoptosis, interestingly, some identified proteins involved in glycolysis such as glucokinase, α-enolase, aldolase, pyruvate kinase, and GAPDH were simultaneously up-regulated, which reveals that the GA-induced cellular apoptosis in B16 melanoma cells is associated with metabolic glycolysis.

Effective screening strategy using ensembled pharmacophore models combined with cascade docking: application to p53-MDM2 interaction inhibitors.

PubMed

Xue, Xin; Wei, Jin-Lian; Xu, Li-Li; Xi, Mei-Yang; Xu, Xiao-Li; Liu, Fang; Guo, Xiao-Ke; Wang, Lei; Zhang, Xiao-Jin; Zhang, Ming-Ye; Lu, Meng-Chen; Sun, Hao-Peng; You, Qi-Dong

2013-10-28

Protein-protein interactions (PPIs) play a crucial role in cellular function and form the backbone of almost all biochemical processes. In recent years, protein-protein interaction inhibitors (PPIIs) have represented a treasure trove of potential new drug targets. Unfortunately, there are few successful drugs of PPIIs on the market. Structure-based pharmacophore (SBP) combined with docking has been demonstrated as a useful Virtual Screening (VS) strategy in drug development projects. However, the combination of target complexity and poor binding affinity prediction has thwarted the application of this strategy in the discovery of PPIIs. Here we report an effective VS strategy on p53-MDM2 PPI. First, we built a SBP model based on p53-MDM2 complex cocrystal structures. The model was then simplified by using a Receptor-Ligand complex-based pharmacophore model considering the critical binding features between MDM2 and its small molecular inhibitors. Cascade docking was subsequently applied to improve the hit rate. Based on this strategy, we performed VS on NCI and SPECS databases and successfully discovered 6 novel compounds from 15 hits with the best, compound 1 (NSC 5359), K(i) = 180 ± 50 nM. These compounds can serve as lead compounds for further optimization.

Comparative proteomics of human endothelial cell caveolae and rafts using two-dimensional gel electrophoresis and mass spectrometry.

PubMed

Sprenger, Richard R; Speijer, Dave; Back, Jaap Willem; De Koster, Chris G; Pannekoek, Hans; Horrevoets, Anton J G

2004-01-01

The human endothelial cell plasma membrane harbors two subdomains of similar lipid composition, caveolae and rafts, both crucially involved in various essential cellular processes like transcytosis, signal transduction and cholesterol homeostasis. Caveolin-enriched membranes, isolated by either cationic silica or buoyant density methods, were explored by comparing large series of two-dimensional (2-D) maps and subsequent identification of over 100 protein spots by matrix-assisted laser desorption/ionization (MALDI) peptide mass fingerprinting. Improved representation and identification of membrane proteins and valuable information on various post-translational modifications was achieved by the presented optimized procedures for solubilization, destaining and database searching/computing. Whereas the cationic silica purification yielded predominantly known endoplasmic reticulum residents, the cold-detergent method yielded a large number of known caveolae residents, including caveolin-1. Thus, a large part of this subproteome was established, including known (trans-)membrane, signal transduction and glycosyl phosphatidylinositol (GPI)-anchored proteins. Several predicted proteins from the human genome were isolated for the first time from biological samples, including SGRP58, SLP-2, C8ORF2, and XRP-2. These findings and various optimized procedures can serve as a reference to study the differential composition of endothelial cell caveolae and rafts, known to be involved in pathologies like cancer and cardiovascular disease.

THGS: a web-based database of Transmembrane Helices in Genome Sequences

PubMed Central

Fernando, S. A.; Selvarani, P.; Das, Soma; Kumar, Ch. Kiran; Mondal, Sukanta; Ramakumar, S.; Sekar, K.

2004-01-01

Transmembrane Helices in Genome Sequences (THGS) is an interactive web-based database, developed to search the transmembrane helices in the user-interested gene sequences available in the Genome Database (GDB). The proposed database has provision to search sequence motifs in transmembrane and globular proteins. In addition, the motif can be searched in the other sequence databases (Swiss-Prot and PIR) or in the macromolecular structure database, Protein Data Bank (PDB). Further, the 3D structure of the corresponding queried motif, if it is available in the solved protein structures deposited in the Protein Data Bank, can also be visualized using the widely used graphics package RASMOL. All the sequence databases used in the present work are updated frequently and hence the results produced are up to date. The database THGS is freely available via the world wide web and can be accessed at http://pranag.physics.iisc.ernet.in/thgs/ or http://144.16.71.10/thgs/. PMID:14681375

Cross-reactivity of anti-HIV-1 T cell immune responses among the major HIV-1 clades in HIV-1-positive individuals from 4 continents.

PubMed

Coplan, Paul M; Gupta, Swati B; Dubey, Sheri A; Pitisuttithum, Punnee; Nikas, Alex; Mbewe, Bernard; Vardas, Efthyia; Schechter, Mauro; Kallas, Esper G; Freed, Dan C; Fu, Tong-Ming; Mast, Christopher T; Puthavathana, Pilaipan; Kublin, James; Brown Collins, Kelly; Chisi, John; Pendame, Richard; Thaler, Scott J; Gray, Glenda; Mcintyre, James; Straus, Walter L; Condra, Jon H; Mehrotra, Devan V; Guess, Harry A; Emini, Emilio A; Shiver, John W

2005-05-01

The genetic diversity of human immunodeficiency virus type 1 (HIV-1) raises the question of whether vaccines that include a component to elicit antiviral T cell immunity based on a single viral genetic clade could provide cellular immune protection against divergent HIV-1 clades. Therefore, we quantified the cross-clade reactivity, among unvaccinated individuals, of anti-HIV-1 T cell responses to the infecting HIV-1 clade relative to other major circulating clades. Cellular immune responses to HIV-1 clades A, B, and C were compared by standardized interferon- gamma enzyme-linked immunospot assays among 250 unvaccinated individuals, infected with diverse HIV-1 clades, from Brazil, Malawi, South Africa, Thailand, and the United States. Cross-clade reactivity was evaluated by use of the ratio of responses to heterologous versus homologous (infecting) clades of HIV-1. Cellular immune responses were predominantly focused on viral Gag and Nef proteins. Cross-clade reactivity of cellular immune responses to HIV-1 clade A, B, and C proteins was substantial for Nef proteins (ratio, 0.97 [95% confidence interval, 0.89-1.05]) and lower for Gag proteins (ratio, 0.67 [95% confidence interval, 0.62-0.73]). The difference in cross-clade reactivity to Nef and Gag proteins was significant (P<.0001). Cross-clade reactivity of cellular immune responses can be substantial but varies by viral protein.

The Novel Fission Yeast Protein Pal1p Interacts with Hip1-related Sla2p/End4p and Is Involved in Cellular Morphogenesis

PubMed Central

Ge, Wanzhong; Chew, Ting Gang; Wachtler, Volker; Naqvi, Suniti N.; Balasubramanian, Mohan K.

2005-01-01

The establishment and maintenance of characteristic cellular morphologies is a fundamental property of all cells. Here we describe Schizosaccharomyces pombe Pal1p, a protein important for maintenance of cylindrical cellular morphology. Pal1p is a novel membrane-associated protein that localizes to the growing tips of interphase cells and to the division site in cells undergoing cytokinesis in an F-actin- and microtubule-independent manner. Cells deleted for pal1 display morphological defects, characterized by the occurrence of spherical and pear-shaped cells with an abnormal cell wall. Pal1p physically interacts and displays overlapping localization with the Huntingtin-interacting-protein (Hip1)-related protein Sla2p/End4p, which is also required for establishment of cylindrical cellular morphology. Sla2p is important for efficient localization of Pal1p to the sites of polarized growth and appears to function upstream of Pal1p. Interestingly, spherical pal1Δ mutants polarize to establish a pearlike morphology before mitosis in a manner dependent on the kelch-repeat protein Tea1p and the cell cycle inhibitory kinase Wee1p. Thus, overlapping mechanisms involving Pal1p, Tea1p, and Sla2p contribute to the establishment of cylindrical cellular morphology, which is important for proper spatial regulation of cytokinesis. PMID:15975911

The novel fission yeast protein Pal1p interacts with Hip1-related Sla2p/End4p and is involved in cellular morphogenesis.

PubMed

Ge, Wanzhong; Chew, Ting Gang; Wachtler, Volker; Naqvi, Suniti N; Balasubramanian, Mohan K

2005-09-01

The establishment and maintenance of characteristic cellular morphologies is a fundamental property of all cells. Here we describe Schizosaccharomyces pombe Pal1p, a protein important for maintenance of cylindrical cellular morphology. Pal1p is a novel membrane-associated protein that localizes to the growing tips of interphase cells and to the division site in cells undergoing cytokinesis in an F-actin- and microtubule-independent manner. Cells deleted for pal1 display morphological defects, characterized by the occurrence of spherical and pear-shaped cells with an abnormal cell wall. Pal1p physically interacts and displays overlapping localization with the Huntingtin-interacting-protein (Hip1)-related protein Sla2p/End4p, which is also required for establishment of cylindrical cellular morphology. Sla2p is important for efficient localization of Pal1p to the sites of polarized growth and appears to function upstream of Pal1p. Interestingly, spherical pal1Delta mutants polarize to establish a pearlike morphology before mitosis in a manner dependent on the kelch-repeat protein Tea1p and the cell cycle inhibitory kinase Wee1p. Thus, overlapping mechanisms involving Pal1p, Tea1p, and Sla2p contribute to the establishment of cylindrical cellular morphology, which is important for proper spatial regulation of cytokinesis.

Sys-BodyFluid: a systematical database for human body fluid proteome research

PubMed Central

Li, Su-Jun; Peng, Mao; Li, Hong; Liu, Bo-Shu; Wang, Chuan; Wu, Jia-Rui; Li, Yi-Xue; Zeng, Rong

2009-01-01

Recently, body fluids have widely become an important target for proteomic research and proteomic study has produced more and more body fluid related protein data. A database is needed to collect and analyze these proteome data. Thus, we developed this web-based body fluid proteome database Sys-BodyFluid. It contains eleven kinds of body fluid proteomes, including plasma/serum, urine, cerebrospinal fluid, saliva, bronchoalveolar lavage fluid, synovial fluid, nipple aspirate fluid, tear fluid, seminal fluid, human milk and amniotic fluid. Over 10 000 proteins are presented in the Sys-BodyFluid. Sys-BodyFluid provides the detailed protein annotations, including protein description, Gene Ontology, domain information, protein sequence and involved pathways. These proteome data can be retrieved by using protein name, protein accession number and sequence similarity. In addition, users can query between these different body fluids to get the different proteins identification information. Sys-BodyFluid database can facilitate the body fluid proteomics and disease proteomics research as a reference database. It is available at http://www.biosino.org/bodyfluid/. PMID:18978022

Sys-BodyFluid: a systematical database for human body fluid proteome research.

PubMed

Li, Su-Jun; Peng, Mao; Li, Hong; Liu, Bo-Shu; Wang, Chuan; Wu, Jia-Rui; Li, Yi-Xue; Zeng, Rong

2009-01-01

Recently, body fluids have widely become an important target for proteomic research and proteomic study has produced more and more body fluid related protein data. A database is needed to collect and analyze these proteome data. Thus, we developed this web-based body fluid proteome database Sys-BodyFluid. It contains eleven kinds of body fluid proteomes, including plasma/serum, urine, cerebrospinal fluid, saliva, bronchoalveolar lavage fluid, synovial fluid, nipple aspirate fluid, tear fluid, seminal fluid, human milk and amniotic fluid. Over 10,000 proteins are presented in the Sys-BodyFluid. Sys-BodyFluid provides the detailed protein annotations, including protein description, Gene Ontology, domain information, protein sequence and involved pathways. These proteome data can be retrieved by using protein name, protein accession number and sequence similarity. In addition, users can query between these different body fluids to get the different proteins identification information. Sys-BodyFluid database can facilitate the body fluid proteomics and disease proteomics research as a reference database. It is available at http://www.biosino.org/bodyfluid/.

Proteomic Analysis of Matched Formalin-Fixed, Paraffin-Embedded Specimens in Patients with Advanced Serous Ovarian Carcinoma

PubMed Central

Smith, Ashlee L.; Sun, Mai; Bhargava, Rohit; Stewart, Nicolas A.; Flint, Melanie S.; Bigbee, William L.; Krivak, Thomas C.; Strange, Mary A.; Cooper, Kristine L.; Zorn, Kristin K.

2013-01-01

Objective: The biology of high grade serous ovarian carcinoma (HGSOC) is poorly understood. Little has been reported on intratumoral homogeneity or heterogeneity of primary HGSOC tumors and their metastases. We evaluated the global protein expression profiles of paired primary and metastatic HGSOC from formalin-fixed, paraffin-embedded (FFPE) tissue samples. Methods: After IRB approval, six patients with advanced HGSOC were identified with tumor in both ovaries at initial surgery. Laser capture microdissection (LCM) was used to extract tumor for protein digestion. Peptides were extracted and analyzed by reversed-phase liquid chromatography coupled to a linear ion trap mass spectrometer. Tandem mass spectra were searched against the UniProt human protein database. Differences in protein abundance between samples were assessed and analyzed by Ingenuity Pathway Analysis software. Immunohistochemistry (IHC) for select proteins from the original and an additional validation set of five patients was performed. Results: Unsupervised clustering of the abundance profiles placed the paired specimens adjacent to each other. IHC H-score analysis of the validation set revealed a strong correlation between paired samples for all proteins. For the similarly expressed proteins, the estimated correlation coefficients in two of three experimental samples and all validation samples were statistically significant (p < 0.05). The estimated correlation coefficients in the experimental sample proteins classified as differentially expressed were not statistically significant. Conclusion: A global proteomic screen of primary HGSOC tumors and their metastatic lesions identifies tumoral homogeneity and heterogeneity and provides preliminary insight into these protein profiles and the cellular pathways they constitute. PMID:28250404

2′-5′-Oligoadenylate Synthetase-Like Protein Inhibits Respiratory Syncytial Virus Replication and Is Targeted by the Viral Nonstructural Protein 1

PubMed Central

Dhar, Jayeeta; Cuevas, Rolando A.; Goswami, Ramansu; Zhu, Jianzhong

2015-01-01

2′-5′-Oligoadenylate synthetase-like protein (OASL) is an interferon-inducible antiviral protein. Here we describe differential inhibitory activities of human OASL and the two mouse OASL homologs against respiratory syncytial virus (RSV) replication. Interestingly, nonstructural protein 1 (NS1) of RSV promoted proteasome-dependent degradation of specific OASL isoforms. We conclude that OASL acts as a cellular antiviral protein and that RSV NS1 suppresses this function to evade cellular innate immunity and allow virus growth. PMID:26178980

AAA+ Machines of Protein Destruction in Mycobacteria.

PubMed

Alhuwaider, Adnan Ali H; Dougan, David A

2017-01-01

The bacterial cytosol is a complex mixture of macromolecules (proteins, DNA, and RNA), which collectively are responsible for an enormous array of cellular tasks. Proteins are central to most, if not all, of these tasks and as such their maintenance (commonly referred to as protein homeostasis or proteostasis) is vital for cell survival during normal and stressful conditions. The two key aspects of protein homeostasis are, (i) the correct folding and assembly of proteins (coupled with their delivery to the correct cellular location) and (ii) the timely removal of unwanted or damaged proteins from the cell, which are performed by molecular chaperones and proteases, respectively. A major class of proteins that contribute to both of these tasks are the AAA+ (ATPases associated with a variety of cellular activities) protein superfamily. Although much is known about the structure of these machines and how they function in the model Gram-negative bacterium Escherichia coli , we are only just beginning to discover the molecular details of these machines and how they function in mycobacteria. Here we review the different AAA+ machines, that contribute to proteostasis in mycobacteria. Primarily we will focus on the recent advances in the structure and function of AAA+ proteases, the substrates they recognize and the cellular pathways they control. Finally, we will discuss the recent developments related to these machines as novel drug targets.

Proteomic characterization of an isolated fraction of synthetic proteasome inhibitor (PSI)-induced inclusions in PC12 cells might offer clues to aggresomes as a cellular defensive response against proteasome inhibition by PSI

PubMed Central

2010-01-01

Background Cooperation of constituents of the ubiquitin proteasome system (UPS) with chaperone proteins in degrading proteins mediate a wide range of cellular processes, such as synaptic function and neurotransmission, gene transcription, protein trafficking, mitochondrial function and metabolism, antioxidant defence mechanisms, and apoptotic signal transduction. It is supposed that constituents of the UPS and chaperone proteins are recruited into aggresomes where aberrant and potentially cytotoxic proteins may be sequestered in an inactive form. Results To determinate the proteomic pattern of synthetic proteasome inhibitor (PSI)-induced inclusions in PC12 cells after proteasome inhibition by PSI, we analyzed a fraction of PSI-induced inclusions. A proteomic feature of the isolated fraction was characterized by identification of fifty six proteins including twenty previously reported protein components of Lewy bodies, twenty eight newly identified proteins and eight unknown proteins. These proteins, most of which were recognized as a profile of proteins within cellular processes mediated by the UPS, a profile of constituents of the UPS and a profile of chaperone proteins, are classed into at least nine accepted categories. In addition, prolyl-4-hydroxylase beta polypeptide, an endoplasmic reticulum member of the protein disulfide isomerase family, was validated in the developmental process of PSI-induced inclusions in the cells. Conclusions It is speculated that proteomic characterization of an isolated fraction of PSI-induced inclusions in PC12 cells might offer clues to appearance of aggresomes serving as a cellular defensive response against proteasome inhibition. PMID:20704702

Proteome analysis of the plant pathogen Xylella fastidiosa reveals major cellular and extracellular proteins and a peculiar codon bias distribution.

PubMed

Smolka, Marcus Bustamante; Martins-de-Souza, Daniel; Martins, Daniel; Winck, Flavia Vischi; Santoro, Carlos Eduardo; Castellari, Rafael Ramos; Ferrari, Fernanda; Brum, Itaraju Junior; Galembeck, Eduardo; Della Coletta Filho, Helvécio; Machado, Marcos Antonio; Marangoni, Sergio; Novello, Jose Camillo

2003-02-01

The bacteria Xylella fastidiosa is the causative agent of a number of economically important crop diseases, including citrus variegated chlorosis. Although its complete genome is already sequenced, X. fastidiosa is very poorly characterized by biochemical approaches at the protein level. In an initial effort to characterize protein expression in X. fastidiosa we used one- and two-dimensional gel electrophoresis and mass spectrometry to identify the products of 142 genes present in a whole cell extract and in an extracellular fraction of the citrus isolated strain 9a5c. Of particular interest for the study of pathogenesis are adhesion and secreted proteins. Homologs to proteins from three different adhesion systems (type IV fimbriae, mrk pili and hsf surface fibrils) were found to be coexpressed, the last two being detected only as multimeric complexes in the high molecular weight region of one-dimensional electrophoresis gels. Using a procedure to extract secreted proteins as well as proteins weakly attached to the cell surface we identified 30 different proteins including toxins, adhesion related proteins, antioxidant enzymes, different types of proteases and 16 hypothetical proteins. These data suggest that the intercellular space of X. fastidiosa colonies is a multifunctional microenvironment containing proteins related to in vivo bacterial survival and pathogenesis. A codon usage analysis of the most expressed proteins from the whole cell extract revealed a low biased distribution, which we propose is related to the slow growing nature of X. fastidiosa. A database of the X. fastidiosa proteome was developed and can be accessed via the internet (URL: www.proteome.ibi.unicamp.br).

Yeast prions are useful for studying protein chaperones and protein quality control.

PubMed

Masison, Daniel C; Reidy, Michael

2015-01-01

Protein chaperones help proteins adopt and maintain native conformations and play vital roles in cellular processes where proteins are partially folded. They comprise a major part of the cellular protein quality control system that protects the integrity of the proteome. Many disorders are caused when proteins misfold despite this protection. Yeast prions are fibrous amyloid aggregates of misfolded proteins. The normal action of chaperones on yeast prions breaks the fibers into pieces, which results in prion replication. Because this process is necessary for propagation of yeast prions, even small differences in activity of many chaperones noticeably affect prion phenotypes. Several other factors involved in protein processing also influence formation, propagation or elimination of prions in yeast. Thus, in much the same way that the dependency of viruses on cellular functions has allowed us to learn much about cell biology, the dependency of yeast prions on chaperones presents a unique and sensitive way to monitor the functions and interactions of many components of the cell's protein quality control system. Our recent work illustrates the utility of this system for identifying and defining chaperone machinery interactions.

De Novo Transcriptome Analysis of Allium cepa L. (Onion) Bulb to Identify Allergens and Epitopes

PubMed Central

Rajkumar, Hemalatha; Ramagoni, Ramesh Kumar; Anchoju, Vijayendra Chary; Vankudavath, Raju Naik; Syed, Arshi Uz Zaman

2015-01-01

Allium cepa (onion) is a diploid plant with one of the largest nuclear genomes among all diploids. Onion is an example of an under-researched crop which has a complex heterozygous genome. There are no allergenic proteins and genomic data available for onions. This study was conducted to establish a transcriptome catalogue of onion bulb that will enable us to study onion related genes involved in medicinal use and allergies. Transcriptome dataset generated from onion bulb using the Illumina HiSeq 2000 technology showed a total of 99,074,309 high quality raw reads (~20 Gb). Based on sequence homology onion genes were categorized into 49 different functional groups. Most of the genes however, were classified under 'unknown' in all three gene ontology categories. Of the categorized genes, 61.2% showed metabolic functions followed by cellular components such as binding, cellular processes; catalytic activity and cell part. With BLASTx top hit analysis, a total of 2,511 homologous allergenic sequences were found, which had 37–100% similarity with 46 different types of allergens existing in the database. From the 46 contigs or allergens, 521 B-cell linear epitopes were identified using BepiPred linear epitope prediction tool. This is the first comprehensive insight into the transcriptome of onion bulb tissue using the NGS technology, which can be used to map IgE epitopes and prediction of structures and functions of various proteins. PMID:26284934

Toxic influence of organophosphate, carbamate, and organochlorine pesticides on cellular metabolism of lipids, proteins, and carbohydrates: a systematic review.

PubMed

Karami-Mohajeri, Somayyeh; Abdollahi, Mohammad

2011-09-01

Pesticides, including organophosphate (OP), organochlorine (OC), and carbamate (CB) compounds, are widely used in agricultural and indoor purposes. OP and CB act as acetyl cholinesterase (AChE) inhibitors that affect lots of organs such as peripheral and central nervous systems, muscles, liver, pancreas, and brain, whereas OC are neurotoxic involved in alteration of ion channels. There are several reports about metabolic disorders, hyperglycemia, and also oxidative stress in acute and chronic exposures to pesticides that are linked with diabetes and other metabolic disorders. In this respect, there are several in vitro and in vivo but few clinical studies about mechanism underlying these effects. Bibliographic databases were searched for the years 1963-2010 and resulted in 1652 articles. After elimination of duplicates or irrelevant papers, 204 papers were included and reviewed. Results indicated that OP and CB impair the enzymatic pathways involved in metabolism of carbohydrates, fats and protein within cytoplasm, mitochondria, and proxisomes. It is believed that OP and CB show this effect through inhibition of AChE or affecting target organs directly. OC mostly affect lipid metabolism in the adipose tissues and change glucose pathway in other cells. As a shared mechanism, all OP, CB and OC induce cellular oxidative stress via affecting mitochondrial function and therefore disrupt neuronal and hormonal status of the body. Establishing proper epidemiological studies to explore exact relationships between exposure levels to these pesticides and rate of resulted metabolic disorders in human will be helpful.

Principal component analysis on molecular descriptors as an alternative point of view in the search of new Hsp90 inhibitors.

PubMed

Lauria, Antonino; Ippolito, Mario; Almerico, Anna Maria

2009-10-01

Inhibiting a protein that regulates multiple signal transduction pathways in cancer cells is an attractive goal for cancer therapy. Heat shock protein 90 (Hsp90) is one of the most promising molecular targets for such an approach. In fact, Hsp90 is a ubiquitous molecular chaperone protein that is involved in folding, activating and assembling of many key mediators of signal transduction, cellular growth, differentiation, stress-response and apoptothic pathways. With the aim to analyze which molecular descriptors have the higher importance in the binding interactions of these classes, we first performed molecular docking experiments on the 187 Hsp90 inhibitors included in the BindingDB, a public database of measured binding affinities. Further, for each frozen conformation obtained from the docking, a set of 250 molecular descriptors was calculated, and the resulting Structure/Descriptors matrix was submitted to Principal Component Analysis. From the factor scores it emerged a good clusterization among similar compounds both in terms of structural class and activity spectrum, while examination of the loadings of the first two factors also allowed to study the classes of descriptors which mainly contribute to each one.

Exploring the cross talk between ER stress and inflammation in age-related macular degeneration.

PubMed

Kheitan, Samira; Minuchehr, Zarrin; Soheili, Zahra-Soheila

2017-01-01

Increasing evidence demonstrates that inflammation and endoplasmic reticulum (ER) stress is implicated in the development and progression of age-related macular degeneration (AMD), a multifactorial neurodegenerative disease. However the cross talk between these cellular mechanisms has not been clearly and fully understood. The present study investigates a possible intersection between ER stress and inflammation in AMD. In this study, we recruited two collections of involved protein markers to retrieve their interaction information from IMEx-curated databases, which are the most well- known protein-protein interaction collections, allowing us to design an intersection network for AMD that is unprecedented. In order to find expression activated subnetworks, we utilized AMD expression profiles in our network. In addition, we studied topological characteristics of the most expressed active subnetworks to identify the hubs. With regard to topological quantifications and expressional activity, we reported a list of the most pivotal hubs which are potentially applicable as probable therapeutic targets. Furthermore, we introduced MAPK signaling pathway as a significantly involved pathway in the association between ER stress and inflammation, leading to promising new directions in discovering AMD formation mechanisms and possible treatments.

Exploring the cross talk between ER stress and inflammation in age-related macular degeneration

PubMed Central

Kheitan, Samira; Soheili, Zahra-Soheila

2017-01-01

Increasing evidence demonstrates that inflammation and endoplasmic reticulum (ER) stress is implicated in the development and progression of age-related macular degeneration (AMD), a multifactorial neurodegenerative disease. However the cross talk between these cellular mechanisms has not been clearly and fully understood. The present study investigates a possible intersection between ER stress and inflammation in AMD. In this study, we recruited two collections of involved protein markers to retrieve their interaction information from IMEx-curated databases, which are the most well- known protein-protein interaction collections, allowing us to design an intersection network for AMD that is unprecedented. In order to find expression activated subnetworks, we utilized AMD expression profiles in our network. In addition, we studied topological characteristics of the most expressed active subnetworks to identify the hubs. With regard to topological quantifications and expressional activity, we reported a list of the most pivotal hubs which are potentially applicable as probable therapeutic targets. Furthermore, we introduced MAPK signaling pathway as a significantly involved pathway in the association between ER stress and inflammation, leading to promising new directions in discovering AMD formation mechanisms and possible treatments. PMID:28742151

Hole hopping through tyrosine/tryptophan chains protects proteins from oxidative damage

PubMed Central

Gray, Harry B.; Winkler, Jay R.

2015-01-01

Living organisms have adapted to atmospheric dioxygen by exploiting its oxidizing power while protecting themselves against toxic side effects. Reactive oxygen and nitrogen species formed during oxidative stress, as well as high-potential reactive intermediates formed during enzymatic catalysis, could rapidly and irreversibly damage polypeptides were protective mechanisms not available. Chains of redox-active tyrosine and tryptophan residues can transport potentially damaging oxidizing equivalents (holes) away from fragile active sites and toward protein surfaces where they can be scavenged by cellular reductants. Precise positioning of these chains is required to provide effective protection without inhibiting normal function. A search of the structural database reveals that about one third of all proteins contain Tyr/Trp chains composed of three or more residues. Although these chains are distributed among all enzyme classes, they appear with greatest frequency in the oxidoreductases and hydrolases. Consistent with a redox-protective role, approximately half of the dioxygen-using oxidoreductases have Tyr/Trp chain lengths ≥3 residues. Among the hydrolases, long Tyr/Trp chains appear almost exclusively in the glycoside hydrolases. These chains likely are important for substrate binding and positioning, but a secondary redox role also is a possibility. PMID:26195784

Structural genomics reveals EVE as a new ASCH/PUA-related domain

PubMed Central

Bertonati, Claudia; Punta, Marco; Fischer, Markus; Yachdav, Guy; Forouhar, Farhad; Zhou, Weihong; Kuzin, Alexander P.; Seetharaman, Jayaraman; Abashidze, Mariam; Ramelot, Theresa A.; Kennedy, Michael A.; Cort, John R.; Belachew, Adam; Hunt, John F.; Tong, Liang; Montelione, Gaetano T.; Rost, Burkhard

2014-01-01

Summary We report on several proteins recently solved by structural genomics consortia, in particular by the Northeast Structural Genomics consortium (NESG). The proteins considered in this study differ substantially in their sequences but they share a similar structural core, characterized by a pseudobarrel five-stranded beta sheet. This core corresponds to the PUA domain-like architecture in the SCOP database. By connecting sequence information with structural knowledge, we characterize a new subgroup of these proteins that we propose to be distinctly different from previously described PUA domain-like domains such as PUA proper or ASCH. We refer to these newly defined domains as EVE. Although EVE may have retained the ability of PUA domains to bind RNA, the available experimental and computational data suggests that both the details of its molecular function and its cellular function differ from those of other PUA domain-like domains. This study of EVE and its relatives illustrates how the combination of structure and genomics creates new insights by connecting a cornucopia of structures that map to the same evolutionary potential. Primary sequence information alone would have not been sufficient to reveal these evolutionary links. PMID:19191354

Structural Genomics Reveals EVE as a New ASCH/PUA-Related Domain

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bertonati, C.; Punta, M; Fischer, M

2008-01-01

We report on several proteins recently solved by structural genomics consortia, in particular by the Northeast Structural Genomics consortium (NESG). The proteins considered in this study differ substantially in their sequences but they share a similar structural core, characterized by a pseudobarrel five-stranded beta sheet. This core corresponds to the PUA domain-like architecture in the SCOP database. By connecting sequence information with structural knowledge, we characterize a new subgroup of these proteins that we propose to be distinctly different from previously described PUA domain-like domains such as PUA proper or ASCH. We refer to these newly defined domains as EVE.more » Although EVE may have retained the ability of PUA domains to bind RNA, the available experimental and computational data suggests that both the details of its molecular function and its cellular function differ from those of other PUA domain-like domains. This study of EVE and its relatives illustrates how the combination of structure and genomics creates new insights by connecting a cornucopia of structures that map to the same evolutionary potential. Primary sequence information alone would have not been sufficient to reveal these evolutionary links.« less

PARPs database: A LIMS systems for protein-protein interaction data mining or laboratory information management system

PubMed Central

Droit, Arnaud; Hunter, Joanna M; Rouleau, Michèle; Ethier, Chantal; Picard-Cloutier, Aude; Bourgais, David; Poirier, Guy G

2007-01-01

Background In the "post-genome" era, mass spectrometry (MS) has become an important method for the analysis of proteins and the rapid advancement of this technique, in combination with other proteomics methods, results in an increasing amount of proteome data. This data must be archived and analysed using specialized bioinformatics tools. Description We herein describe "PARPs database," a data analysis and management pipeline for liquid chromatography tandem mass spectrometry (LC-MS/MS) proteomics. PARPs database is a web-based tool whose features include experiment annotation, protein database searching, protein sequence management, as well as data-mining of the peptides and proteins identified. Conclusion Using this pipeline, we have successfully identified several interactions of biological significance between PARP-1 and other proteins, namely RFC-1, 2, 3, 4 and 5. PMID:18093328

The 2015 Nucleic Acids Research Database Issue and molecular biology database collection.

PubMed

Galperin, Michael Y; Rigden, Daniel J; Fernández-Suárez, Xosé M

2015-01-01

The 2015 Nucleic Acids Research Database Issue contains 172 papers that include descriptions of 56 new molecular biology databases, and updates on 115 databases whose descriptions have been previously published in NAR or other journals. Following the classification that has been introduced last year in order to simplify navigation of the entire issue, these articles are divided into eight subject categories. This year's highlights include RNAcentral, an international community portal to various databases on noncoding RNA; ValidatorDB, a validation database for protein structures and their ligands; SASBDB, a primary repository for small-angle scattering data of various macromolecular complexes; MoonProt, a database of 'moonlighting' proteins, and two new databases of protein-protein and other macromolecular complexes, ComPPI and the Complex Portal. This issue also includes an unusually high number of cancer-related databases and other databases dedicated to genomic basics of disease and potential drugs and drug targets. The size of NAR online Molecular Biology Database Collection, http://www.oxfordjournals.org/nar/database/a/, remained approximately the same, following the addition of 74 new resources and removal of 77 obsolete web sites. The entire Database Issue is freely available online on the Nucleic Acids Research web site (http://nar.oxfordjournals.org/). Published by Oxford University Press on behalf of Nucleic Acids Research 2014. This work is written by (a) US Government employee(s) and is in the public domain in the US.

The Histone Database: an integrated resource for histones and histone fold-containing proteins

PubMed Central

Mariño-Ramírez, Leonardo; Levine, Kevin M.; Morales, Mario; Zhang, Suiyuan; Moreland, R. Travis; Baxevanis, Andreas D.; Landsman, David

2011-01-01

Eukaryotic chromatin is composed of DNA and protein components—core histones—that act to compactly pack the DNA into nucleosomes, the fundamental building blocks of chromatin. These nucleosomes are connected to adjacent nucleosomes by linker histones. Nucleosomes are highly dynamic and, through various core histone post-translational modifications and incorporation of diverse histone variants, can serve as epigenetic marks to control processes such as gene expression and recombination. The Histone Sequence Database is a curated collection of sequences and structures of histones and non-histone proteins containing histone folds, assembled from major public databases. Here, we report a substantial increase in the number of sequences and taxonomic coverage for histone and histone fold-containing proteins available in the database. Additionally, the database now contains an expanded dataset that includes archaeal histone sequences. The database also provides comprehensive multiple sequence alignments for each of the four core histones (H2A, H2B, H3 and H4), the linker histones (H1/H5) and the archaeal histones. The database also includes current information on solved histone fold-containing structures. The Histone Sequence Database is an inclusive resource for the analysis of chromatin structure and function focused on histones and histone fold-containing proteins. Database URL: The Histone Sequence Database is freely available and can be accessed at http://research.nhgri.nih.gov/histones/. PMID:22025671

Exosome proteomics reveals transcriptional regulator proteins with potential to mediate downstream pathways.

PubMed

Ung, Timothy H; Madsen, Helen J; Hellwinkel, Justin E; Lencioni, Alex M; Graner, Michael W

2014-11-01

Exosomes are virus-sized, membrane-enclosed vesicles with origins in the cellular endosomal system, but are released extracellularly. As a population, these tiny vesicles carry relatively enormous amounts of information in their protein, lipid and nucleic acid content, and the vesicles can have profound impacts on recipient cells. This review employs publically-available data combined with gene ontology applications to propose a novel concept, that exosomes transport transcriptional and translational machinery that may have direct impacts on gene expression in recipient cells. Here, we examine the previously published proteomic contents of medulloblastoma-derived exosomes, focusing on transcriptional regulators; we found that there are numerous proteins that may have potential roles in transcriptional and translational regulation with putative influence on downstream, cancer-related pathways. We expanded this search to all of the proteins in the Vesiclepedia database; using gene ontology approaches, we see that these regulatory factors are implicated in many of the processes involved in cancer initiation and progression. This information suggests that some of the effects of exosomes on recipient cells may be due to the delivery of protein factors that can directly and fundamentally change the transcriptional landscape of the cells. Within a tumor environment, this has potential to tilt the advantage towards the cancer. © 2014 The Authors. Cancer Science published by Wiley Publishing Asia Pty Ltd on behalf of Japanese Cancer Association.

Proteomic methods for analysis of S-nitrosation⋄

PubMed Central

Kettenhofen, Nicholas; Broniowska, Katarzyna; Keszler, Agnes; Zhang, Yanhong; Hogg, Neil

2007-01-01

This review discusses proteomic methods to detect and identify S-nitrosated proteins. Protein S-nitrosation, the post-translational modification of thiol residues to form S-nitrosothiols, has been suggested to be a mechanism of cellular redox signaling by which nitric oxide can alter cellular function through modification of protein thiol residues. It has become apparent that methods that will detect and identify low levels of S-nitrosated protein in complex protein mixtures are required in order to fully appreciate the range, extent and selectivity of this modification in both physiological and pathological conditions. While many advances have been made in the detection of either total cellular S-nitrosation or individual S-nitrosothiols, proteomic methods for the detection of S-nitrosation are in relative infancy. This review will discuss the major methods that have been used for the proteomic analysis of protein S-nitrosation and discuss the pros and cons of this methodology. PMID:17360249

Computational design of a red fluorophore ligase for site-specific protein labeling in living cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liu, Daniel S.; Nivon, Lucas G.; Richter, Florian

In this study, chemical fluorophores offer tremendous size and photophysical advantages over fluorescent proteins but are much more challenging to target to specific cellular proteins. Here, we used Rosetta-based computation to design a fluorophore ligase that accepts the red dye resorufin, starting from Escherichia coli lipoic acid ligase. X-ray crystallography showed that the design closely matched the experimental structure. Resorufin ligase catalyzed the site-specific and covalent attachment of resorufin to various cellular proteins genetically fused to a 13-aa recognition peptide in multiple mammalian cell lines and in primary cultured neurons. We used resorufin ligase to perform superresolution imaging of themore » intermediate filament protein vimentin by stimulated emission depletion and electron microscopies. This work illustrates the power of Rosetta for major redesign of enzyme specificity and introduces a tool for minimally invasive, highly specific imaging of cellular proteins by both conventional and superresolution microscopies.« less

Exploiting the biomolecular corona: pre-coating of nanoparticles enables controlled cellular interactions.

PubMed

Simon, Johanna; Müller, Laura K; Kokkinopoulou, Maria; Lieberwirth, Ingo; Morsbach, Svenja; Landfester, Katharina; Mailänder, Volker

2018-06-14

Formation of the biomolecular corona ultimately determines the successful application of nanoparticles in vivo. Adsorption of biomolecules such as proteins is an inevitable process that takes place instantaneously upon contact with physiological fluid (e.g. blood). Therefore, strategies are needed to control this process in order to improve the properties of the nanoparticles and to allow targeted drug delivery. Here, we show that the design of the protein corona by a pre-formed protein corona with tailored properties enables targeted cellular interactions. Nanoparticles were pre-coated with immunoglobulin depleted plasma to create and design a protein corona that reduces cellular uptake by immune cells. It was proven that a pre-formed protein corona remains stable even after nanoparticles were re-introduced to plasma. This opens up the great potential to exploit protein corona formation, which will significantly influence the development of novel nanomaterials.

Computational design of a red fluorophore ligase for site-specific protein labeling in living cells

DOE PAGES

Liu, Daniel S.; Nivon, Lucas G.; Richter, Florian; ...

2014-10-13

In this study, chemical fluorophores offer tremendous size and photophysical advantages over fluorescent proteins but are much more challenging to target to specific cellular proteins. Here, we used Rosetta-based computation to design a fluorophore ligase that accepts the red dye resorufin, starting from Escherichia coli lipoic acid ligase. X-ray crystallography showed that the design closely matched the experimental structure. Resorufin ligase catalyzed the site-specific and covalent attachment of resorufin to various cellular proteins genetically fused to a 13-aa recognition peptide in multiple mammalian cell lines and in primary cultured neurons. We used resorufin ligase to perform superresolution imaging of themore » intermediate filament protein vimentin by stimulated emission depletion and electron microscopies. This work illustrates the power of Rosetta for major redesign of enzyme specificity and introduces a tool for minimally invasive, highly specific imaging of cellular proteins by both conventional and superresolution microscopies.« less

General Protein Diffusion Barriers create Compartments within Bacterial Cells

PubMed Central

Schlimpert, Susan; Klein, Eric A.; Briegel, Ariane; Hughes, Velocity; Kahnt, Jörg; Bolte, Kathrin; Maier, Uwe G.; Brun, Yves V.; Jensen, Grant J.; Gitai, Zemer; Thanbichler, Martin

2013-01-01

SUMMARY In eukaryotes, the differentiation of cellular extensions such as cilia or neuronal axons depends on the partitioning of proteins to distinct plasma membrane domains by specialized diffusion barriers. However, examples of this compartmentalization strategy are still missing for prokaryotes, although complex cellular architectures are widespread among this group of organisms. This study reveals the existence of a protein-mediated membrane diffusion barrier in the stalked bacterium Caulobacter crescentus. We show that the Caulobacter cell envelope is compartmentalized by macromolecular complexes that prevent the exchange of both membrane and soluble proteins between the polar stalk extension and the cell body. The barrier structures span the cross-sectional area of the stalk and comprise at least four proteins that assemble in a cell cycle-dependent manner. Their presence is critical for cellular fitness, as they minimize the effective cell volume, allowing faster adaptation to environmental changes that require de novo synthesis of envelope proteins. PMID:23201141

Role of naturally occurring osmolytes in protein folding and stability.

PubMed

Kumar, Raj

2009-11-01

Osmolytes are typically accumulated in the intracellular environment at relatively high concentrations when cells/tissues are subjected to stress conditions. Osmolytes are common in a variety of organisms, including microorganisms, plants, and animals. They enhance thermodynamic stability of proteins by providing natively folded conformations without perturbing other cellular processes. By burying the backbone into the core of folded proteins, osmolytes can provide significant stability to proteins. Two properties of osmolytes are particularly important: (i) their ability to impart increased thermodynamic stability to folded proteins; and (ii) their compatibility in the intracellular environment at high concentrations. Under physiological conditions, the cellular compositions of osmolytes may vary significantly. This may lead to different protein folding pathways utilized in cells depending upon the intracellular environment. Proper understanding of the role of osmolytes in cell regulation should allow predicting the action of osmolytes on macromolecular interactions in stressed and crowded environments typical of cellular conditions.

The effects of glutathione depletion on thermotolerance and heat stress protein synthesis.

PubMed Central

Russo, A.; Mitchell, J. B.; McPherson, S.

1984-01-01

The effects of cellular glutathione depletion by buthionine sulfoximine on the development of thermotolerance and synthesis of heat stress protein was studied. Cellular glutathione levels were found to increase rapidly following an acute heat treatment of either 12 min at 45.5 degrees C or 1 h at 43 degrees C and remain elevated for prolonged periods. Glutathione depletion and prevention of glutathione synthesis by buthionine sulfoximine resulted in inhibition of the development of thermotolerance and a decrease in total protein as well as specific heat stress proteins. While the degree of inhibition of thermotolerance was similar for both glutathione depletion protocols, inhibition in heat stress protein synthesis was greater when glutathione was depleted to low levels prior to heating. The possible role of glutathione and the cellular redox state to thermotolerance and synthesis of heat stress protein is discussed. Images Figure 2 PMID:6733022

The electric dipole moment of DNA-binding HU protein calculated by the use of an NMR database.

PubMed

Takashima, S; Yamaoka, K

1999-08-30

Electric birefringence measurements indicated the presence of a large permanent dipole moment in HU protein-DNA complex. In order to substantiate this observation, numerical computation of the dipole moment of HU protein homodimer was carried out by using NMR protein databases. The dipole moments of globular proteins have hitherto been calculated with X-ray databases and NMR data have never been used before. The advantages of NMR databases are: (a) NMR data are obtained, unlike X-ray databases, using protein solutions. Accordingly, this method eliminates the bothersome question as to the possible alteration of the protein structure due to the transition from the crystalline state to the solution state. This question is particularly important for proteins such as HU protein which has some degree of internal flexibility; (b) the three-dimensional coordinates of hydrogen atoms in protein molecules can be determined with a sufficient resolution and this enables the N-H as well as C = O bond moments to be calculated. Since the NMR database of HU protein from Bacillus stearothermophilus consists of 25 models, the surface charge as well as the core dipole moments were computed for each of these structures. The results of these calculations show that the net permanent dipole moments of HU protein homodimer is approximately 500-530 D (1 D = 3.33 x 10(-30) Cm) at pH 7.5 and 600-630 D at the isoelectric point (pH 10.5). These permanent dipole moments are unusually large for a small protein of the size of 19.5 kDa. Nevertheless, the result of numerical calculations is compatible with the electro-optical observation, confirming a very large dipole moment in this protein.

Protein structure in context: The molecular landscape of angiogenesis

PubMed Central

Span, Elise A.; Goodsell, David S.; Ramchandran, Ramani; Franzen, Margaret; Herman, Timothy; Sem, Daniel S.

2014-01-01

A team of students, educators, and researchers has developed new materials to teach cell signaling within its cellular context. Two non-traditional modalities are employed: physical models, to explore the atomic details of several of the proteins in the angiogenesis signaling cascade, and illustrations of the proteins in their cellular environment, to give an intuitive understanding of the cellular context of the pathway. The experiences of the team underscore the utility of these types of materials as an effective mode for fostering students’ understanding of the molecular world, and the scientific method used to define it. PMID:23868376

MoonProt: a database for proteins that are known to moonlight

PubMed Central

Mani, Mathew; Chen, Chang; Amblee, Vaishak; Liu, Haipeng; Mathur, Tanu; Zwicke, Grant; Zabad, Shadi; Patel, Bansi; Thakkar, Jagravi; Jeffery, Constance J.

2015-01-01

Moonlighting proteins comprise a class of multifunctional proteins in which a single polypeptide chain performs multiple biochemical functions that are not due to gene fusions, multiple RNA splice variants or pleiotropic effects. The known moonlighting proteins perform a variety of diverse functions in many different cell types and species, and information about their structures and functions is scattered in many publications. We have constructed the manually curated, searchable, internet-based MoonProt Database (http://www.moonlightingproteins.org) with information about the over 200 proteins that have been experimentally verified to be moonlighting proteins. The availability of this organized information provides a more complete picture of what is currently known about moonlighting proteins. The database will also aid researchers in other fields, including determining the functions of genes identified in genome sequencing projects, interpreting data from proteomics projects and annotating protein sequence and structural databases. In addition, information about the structures and functions of moonlighting proteins can be helpful in understanding how novel protein functional sites evolved on an ancient protein scaffold, which can also help in the design of proteins with novel functions. PMID:25324305

Ubiquitylation-dependent regulation of NEIL1 by Mule and TRIM26 is required for the cellular DNA damage response.

PubMed

Edmonds, Matthew J; Carter, Rachel J; Nickson, Catherine M; Williams, Sarah C; Parsons, Jason L

2017-01-25

Endonuclease VIII-like protein 1 (NEIL1) is a DNA glycosylase involved in initiating the base excision repair pathway, the major cellular mechanism for repairing DNA base damage. Here, we have purified the major E3 ubiquitin ligases from human cells responsible for regulation of NEIL1 by ubiquitylation. Interestingly, we have identified two enzymes that catalyse NEIL1 polyubiquitylation, Mcl-1 ubiquitin ligase E3 (Mule) and tripartite motif 26 (TRIM26). We demonstrate that these enzymes are capable of polyubiquitylating NEIL1 in vitro, and that both catalyse ubiquitylation of NEIL1 within the same C-terminal lysine residues. An siRNA-mediated knockdown of Mule or TRIM26 leads to stabilisation of NEIL1, demonstrating that these enzymes are important in regulating cellular NEIL1 steady state protein levels. Similarly, a mutant NEIL1 protein lacking residues for ubiquitylation is more stable than the wild type protein in vivo We also demonstrate that cellular NEIL1 protein is induced in response to ionising radiation (IR), although this occurs specifically in a Mule-dependent manner. Finally we show that stabilisation of NEIL1, particularly following TRIM26 siRNA, contributes to cellular resistance to IR. This highlights the importance of Mule and TRIM26 in maintaining steady state levels of NEIL1, but also those required for the cellular DNA damage response. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

Cyclosporin A Associated Helicase-Like Protein Facilitates the Association of Hepatitis C Virus RNA Polymerase with Its Cellular Cyclophilin B

PubMed Central

Sahara, Hiroeki; Iwabata, Kazuki; Sunoki, Takashi; Kuramochi, Kouji; Takakusagi, Kaori; Miyashita, Hiroki; Sato, Noriyuki; Tanabe, Atsushi; Shimotohno, Kunitada; Kobayashi, Susumu; Sakaguchi, Kengo; Sugawara, Fumio

2011-01-01

Background Cyclosporin A (CsA) is well known as an immunosuppressive drug useful for allogeneic transplantation. It has been reported that CsA inhibits hepatitis C virus (HCV) genome replication, which indicates that cellular targets of CsA regulate the viral replication. However, the regulation mechanisms of HCV replication governed by CsA target proteins have not been fully understood. Principal Findings Here we show a chemical biology approach that elucidates a novel mechanism of HCV replication. We developed a phage display screening to investigate compound-peptide interaction and identified a novel cellular target molecule of CsA. This protein, named CsA associated helicase-like protein (CAHL), possessed RNA-dependent ATPase activity that was negated by treatment with CsA. The downregulation of CAHL in the cells resulted in a decrease of HCV genome replication. CAHL formed a complex with HCV-derived RNA polymerase NS5B and host-derived cyclophilin B (CyPB), known as a cellular cofactor for HCV replication, to regulate NS5B-CyPB interaction. Conclusions We found a cellular factor, CAHL, as CsA associated helicase-like protein, which would form trimer complex with CyPB and NS5B of HCV. The strategy using a chemical compound and identifying its target molecule by our phage display analysis is useful to reveal a novel mechanism underlying cellular and viral physiology. PMID:21559518

Slight temperature changes affect protein affinity and cellular uptake/toxicity of nanoparticles

NASA Astrophysics Data System (ADS)

Mahmoudi, Morteza; Shokrgozar, Mohammad A.; Behzadi, Shahed

2013-03-01

It is known that what the cell actually ``sees'' at the nanoscale is an outer shell formed of `protein corona' on the surface of nanoparticles (NPs). The amount and composition of various proteins on the corona are strongly dependent on the biophysicochemical properties of NPs, which have been extensively studied. However, the effect of a small variation in temperature, due to the human circadian rhythm, on the composition of the protein corona and the affinity of various proteins to the surface of NPs, was ignored. Here, the effect of temperature on the composition of protein corona and the affinity of various proteins to the surface of NPs and, subsequently, cell responses to the protein coated NPs are probed. The results confirmed that cellular entrance, dispersion, and toxicity of NPs are strongly diverse with slight body temperature changes. This new finding can help scientists to maximise NP entrance to specific cells/organs with lower toxicity by adjusting the cellular/organ temperature.It is known that what the cell actually ``sees'' at the nanoscale is an outer shell formed of `protein corona' on the surface of nanoparticles (NPs). The amount and composition of various proteins on the corona are strongly dependent on the biophysicochemical properties of NPs, which have been extensively studied. However, the effect of a small variation in temperature, due to the human circadian rhythm, on the composition of the protein corona and the affinity of various proteins to the surface of NPs, was ignored. Here, the effect of temperature on the composition of protein corona and the affinity of various proteins to the surface of NPs and, subsequently, cell responses to the protein coated NPs are probed. The results confirmed that cellular entrance, dispersion, and toxicity of NPs are strongly diverse with slight body temperature changes. This new finding can help scientists to maximise NP entrance to specific cells/organs with lower toxicity by adjusting the cellular/organ temperature. Electronic supplementary information (ESI) available. See DOI: 10.1039/c3nr32551b

Impact of silica nanoparticle surface chemistry on protein corona formation and consequential interactions with biological cells.

PubMed

Kurtz-Chalot, Andréa; Villiers, Christian; Pourchez, Jérémie; Boudard, Delphine; Martini, Matteo; Marche, Patrice N; Cottier, Michèle; Forest, Valérie

2017-06-01

Nanoparticles (NP) physico-chemical features greatly influence NP/cell interactions. NP surface functionalization is often used to improve NP biocompatibility or to enhance cellular uptake. But in biological media, the formation of a protein corona adds a level of complexity. The aim of this study was to investigate in vitro the influence of NP surface functionalization on their cellular uptake and the biological response induced. 50nm fluorescent silica NP were functionalized either with amine or carboxylic groups, in presence or in absence of polyethylene glycol (PEG). NP were incubated with macrophages, cellular uptake and cellular response were assessed in terms of cytotoxicity, pro-inflammatory response and oxidative stress. The NP protein corona was also characterized by protein mass spectroscopy. Results showed that NP uptake was enhanced in absence of PEG, while NP adsorption at the cell membrane was fostered by an initial positively charged NP surface. NP toxicity was not correlated with NP uptake. NP surface functionalization also influenced the formation of the protein corona as the profile of protein binding differed among the NP types. Copyright © 2017 Elsevier B.V. All rights reserved.

Raman microscopy of bladder cancer cells expressing green fluorescent protein

NASA Astrophysics Data System (ADS)

Mandair, Gurjit S.; Han, Amy L.; Keller, Evan T.; Morris, Michael D.

2016-11-01

Gene engineering is a commonly used tool in cellular biology to determine changes in function or expression of downstream targets. However, the impact of genetic modulation on biochemical effects is less frequently evaluated. The aim of this study is to use Raman microscopy to assess the biochemical effects of gene silencing on T24 and UMUC-13 bladder cancer cell lines. Cellular biochemical information related to nucleic acid and lipogenic components was obtained from deconvolved Raman spectra. We show that the green fluorescence protein (GFP), the chromophore that served as a fluorescent reporter for gene silencing, could also be detected by Raman microscopy. Only the gene-silenced UMUC-13 cell lines exhibited low-to-moderate GFP fluorescence as determined by fluorescence imaging and Raman spectroscopic studies. Moreover, we show that gene silencing and cell phenotype had a greater effect on nucleic acid and lipogenic components with minimal interference from GFP expression. Gene silencing was also found to perturb cellular protein secondary structure in which the amount of disorderd protein increased at the expense of more ordered protein. Overall, our study identified the spectral signature for cellular GFP expression and elucidated the effects of gene silencing on cancer cell biochemistry and protein secondary structure.

Metal ions induced heat shock protein response by elevating superoxide anion level in HeLa cells transformed by HSE-SEAP reporter gene.

PubMed

Yu, Zhanjiang; Yang, Xiaoda; Wang, Kui

2006-06-01

The aim of this work is to define the relationship between heat shock protein (HSP) and reactive oxygen species (ROS) in the cells exposed to different concentrations of metal ions, and to evaluate a new method for tracing the dynamic levels of cellular reactive oxygen species using a HSE-SEAP reporter gene. The expression of heat shock protein was measured using a secreted alkaline phosphatase (SEAP) reporter gene transformed into HeLa cell strain, the levels of superoxide anion (O(2)(-)) and hydrogen peroxide (H(2)O(2)) were determined by NBT reduction assay and DCFH staining flow cytometry (FCM), respectively. The experimental results demonstrated that the expression of heat shock protein induced by metal ions was linearly related to the cellular superoxide anion level before cytotoxic effects were observed, but not related to the cellular hydrogen peroxide level. The experimental results suggested that metal ions might induce heat shock protein by elevating cellular superoxide anion level, and thus the expression of heat shock protein indicated by the HSE-SEAP reporter gene can be an effective model for monitoring the dynamic level of superoxide anion and early metal-induced oxidative stress/cytotoxicity.

A comprehensive and scalable database search system for metaproteomics.

PubMed

Chatterjee, Sandip; Stupp, Gregory S; Park, Sung Kyu Robin; Ducom, Jean-Christophe; Yates, John R; Su, Andrew I; Wolan, Dennis W

2016-08-16

Mass spectrometry-based shotgun proteomics experiments rely on accurate matching of experimental spectra against a database of protein sequences. Existing computational analysis methods are limited in the size of their sequence databases, which severely restricts the proteomic sequencing depth and functional analysis of highly complex samples. The growing amount of public high-throughput sequencing data will only exacerbate this problem. We designed a broadly applicable metaproteomic analysis method (ComPIL) that addresses protein database size limitations. Our approach to overcome this significant limitation in metaproteomics was to design a scalable set of sequence databases assembled for optimal library querying speeds. ComPIL was integrated with a modified version of the search engine ProLuCID (termed "Blazmass") to permit rapid matching of experimental spectra. Proof-of-principle analysis of human HEK293 lysate with a ComPIL database derived from high-quality genomic libraries was able to detect nearly all of the same peptides as a search with a human database (~500x fewer peptides in the database), with a small reduction in sensitivity. We were also able to detect proteins from the adenovirus used to immortalize these cells. We applied our method to a set of healthy human gut microbiome proteomic samples and showed a substantial increase in the number of identified peptides and proteins compared to previous metaproteomic analyses, while retaining a high degree of protein identification accuracy and allowing for a more in-depth characterization of the functional landscape of the samples. The combination of ComPIL with Blazmass allows proteomic searches to be performed with database sizes much larger than previously possible. These large database searches can be applied to complex meta-samples with unknown composition or proteomic samples where unexpected proteins may be identified. The protein database, proteomic search engine, and the proteomic data files for the 5 microbiome samples characterized and discussed herein are open source and available for use and additional analysis.

Mechanism-based Proteomic Screening Identifies Targets of Thioredoxin-like Proteins*

PubMed Central

Nakao, Lia S.; Everley, Robert A.; Marino, Stefano M.; Lo, Sze M.; de Souza, Luiz E.; Gygi, Steven P.; Gladyshev, Vadim N.

2015-01-01

Thioredoxin (Trx)-fold proteins are protagonists of numerous cellular pathways that are subject to thiol-based redox control. The best characterized regulator of thiols in proteins is Trx1 itself, which together with thioredoxin reductase 1 (TR1) and peroxiredoxins (Prxs) comprises a key redox regulatory system in mammalian cells. However, there are numerous other Trx-like proteins, whose functions and redox interactors are unknown. It is also unclear if the principles of Trx1-based redox control apply to these proteins. Here, we employed a proteomic strategy to four Trx-like proteins containing CXXC motifs, namely Trx1, Rdx12, Trx-like protein 1 (Txnl1) and nucleoredoxin 1 (Nrx1), whose cellular targets were trapped in vivo using mutant Trx-like proteins, under conditions of low endogenous expression of these proteins. Prxs were detected as key redox targets of Trx1, but this approach also supported the detection of TR1, which is the Trx1 reductant, as well as mitochondrial intermembrane proteins AIF and Mia40. In addition, glutathione peroxidase 4 was found to be a Rdx12 redox target. In contrast, no redox targets of Txnl1 and Nrx1 could be detected, suggesting that their CXXC motifs do not engage in mixed disulfides with cellular proteins. For some Trx-like proteins, the method allowed distinguishing redox and non-redox interactions. Parallel, comparative analyses of multiple thiol oxidoreductases revealed differences in the functions of their CXXC motifs, providing important insights into thiol-based redox control of cellular processes. PMID:25561728

Learning cellular sorting pathways using protein interactions and sequence motifs.

PubMed

Lin, Tien-Ho; Bar-Joseph, Ziv; Murphy, Robert F

2011-11-01

Proper subcellular localization is critical for proteins to perform their roles in cellular functions. Proteins are transported by different cellular sorting pathways, some of which take a protein through several intermediate locations until reaching its final destination. The pathway a protein is transported through is determined by carrier proteins that bind to specific sequence motifs. In this article, we present a new method that integrates protein interaction and sequence motif data to model how proteins are sorted through these sorting pathways. We use a hidden Markov model (HMM) to represent protein sorting pathways. The model is able to determine intermediate sorting states and to assign carrier proteins and motifs to the sorting pathways. In simulation studies, we show that the method can accurately recover an underlying sorting model. Using data for yeast, we show that our model leads to accurate prediction of subcellular localization. We also show that the pathways learned by our model recover many known sorting pathways and correctly assign proteins to the path they utilize. The learned model identified new pathways and their putative carriers and motifs and these may represent novel protein sorting mechanisms. Supplementary results and software implementation are available from http://murphylab.web.cmu.edu/software/2010_RECOMB_pathways/.

The Role of the Multifunctional BAG3 Protein in Cellular Protein Quality Control and in Disease.

PubMed

Stürner, Elisabeth; Behl, Christian

2017-01-01

In neurons, but also in all other cells the complex proteostasis network is monitored and tightly regulated by the cellular protein quality control (PQC) system. Beyond folding of newly synthesized polypeptides and their refolding upon misfolding the PQC also manages the disposal of aberrant proteins either by the ubiquitin-proteasome machinery or by the autophagic-lysosomal system. Aggregated proteins are primarily degraded by a process termed selective macroautophagy (or aggrephagy). One such recently discovered selective macroautophagy pathway is mediated by the multifunctional HSP70 co-chaperone BAG3 ( BCL-2-associated athanogene 3 ). Under acute stress and during cellular aging, BAG3 in concert with the molecular chaperones HSP70 and HSPB8 as well as the ubiquitin receptor p62/SQSTM1 specifically targets aggregation-prone proteins to autophagic degradation. Thereby, BAG3-mediated selective macroautophagy represents a pivotal adaptive safeguarding and emergency system of the PQC which is activated under pathophysiological conditions to ensure cellular proteostasis. Interestingly, BAG3-mediated selective macroautophagy is also involved in the clearance of aggregated proteins associated with age-related neurodegenerative disorders, like Alzheimer's disease (tau-protein), Huntington's disease (mutated huntingtin/polyQ proteins), and amyotrophic lateral sclerosis (mutated SOD1). In addition, based on its initial description BAG3 is an anti-apoptotic protein that plays a decisive role in other widespread diseases, including cancer and myopathies. Therefore, in the search for novel therapeutic intervention avenues in neurodegeneration, myopathies and cancer BAG3 is a promising candidate.

Equalizer technology followed by DIGE-based proteomics for detection of cellular proteins in artificial peritoneal dialysis effluents.

PubMed

Lichtenauer, Anton Michael; Herzog, Rebecca; Tarantino, Silvia; Aufricht, Christoph; Kratochwill, Klaus

2014-05-01

Peritoneal dialysis effluent (PDE) represents a rich pool of potential biomarkers for monitoring disease and therapy. Until now, proteomic studies have been hindered by the plasma-like composition of the PDE. Beads covered with a peptide library are a promising approach to remove high abundant proteins and concentrate the sample in one step. In this study, a novel approach for proteomic biomarker identification in PDEs consisting of a depletion and concentration step followed by 2D gel based protein quantification was established. To prove this experimental concept a model system of artificial PDEs was established by spiking unused peritoneal dialysis (PD) fluids with cellular proteins reflecting control conditions or cell stress. Using this procedure, we were able to reduce the amount of high abundant plasma proteins and concentrate low abundant proteins while preserving changes in abundance of proteins with cellular origin. The alterations in abundance of the investigated marker for cell stress, the heat shock proteins, showed similar abundance profiles in the artificial PDE as in pure cell culture samples. Our results demonstrate the efficacy of this system in detecting subtle changes in cellular protein expression triggered by unphysiological stress stimuli typical in PD, which could serve as biomarkers. Further studies using patients' PDE will be necessary to prove the concept in clinical PD and to assess whether this technique is also informative regarding enriching low abundant plasma derived protein biomarker in the PDE. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Combined RAF1 protein expression and p53 mutational status provides a strong predictor of cellular radiosensitivity

PubMed Central

Warenius, H M; Jones, M; Gorman, T; McLeish, R; Seabra, L; Barraclough, R; Rudland, P

2000-01-01

The tumour suppressor gene, p53, and genes coding for positive signal transduction factors can influence transit through cell-cycle checkpoints and modulate radiosensitivity. Here we examine the effects of RAF1 protein on the rate of exit from a G2/M block induced by γ-irradiation in relation to intrinsic cellular radiosensitivity in human cell lines expressing wild-type p53 (wtp53) protein as compared to mutant p53 (mutp53) protein. Cell lines which expressed mutp53 protein were all relatively radioresistant and exhibited no relationship between RAF1 protein and cellular radiosensitivity. Cell lines expressing wtp53 protein, however, showed a strong relationship between RAF1 protein levels and the radiosensitivity parameter SF2. In addition, when post-irradiation perturbation of G2/M transit was compared using the parameter T50 (time after the peak of G2/M delay at which 50% of the cells had exited from a block induced by 2 Gy of irradiation), RAF1 was related to T50 in wtp53, but not mutp53, cell lines. Cell lines which expressed wtp53 protein and high levels of RAF1 had shorter T50s and were also more radiosensitive. These results suggest a cooperative role for wtp53 and RAF1 protein in determining cellular radiosensitivity in human cells, which involves control of the G2/M checkpoint. © 2000 Cancer Research Campaign PMID:10993658

Inhibition of host protein synthesis by Sindbis virus: correlation with viral RNA replication and release of nuclear proteins to the cytoplasm.

PubMed

Sanz, Miguel A; García-Moreno, Manuel; Carrasco, Luis

2015-04-01

Infection of mammalian cells by Sindbis virus (SINV) profoundly blocks cellular mRNA translation. Experimental evidence points to viral non-structural proteins (nsPs), in particular nsP2, as the mediator of this inhibition. However, individual expression of nsP1, nsP2, nsP3 or nsP1-4 does not block cellular protein synthesis in BHK cells. Trans-complementation of a defective SINV replicon lacking most of the coding region for nsPs by the co-expression of nsP1-4 propitiates viral RNA replication at low levels, and inhibition of cellular translation is not observed. Exit of nuclear proteins including T-cell intracellular antigen and polypyrimidine tract-binding protein is clearly detected in SINV-infected cells, but not upon the expression of nsPs, even when the defective replicon was complemented. Analysis of a SINV variant with a point mutation in nsP2, exhibiting defects in the shut-off of host protein synthesis, indicates that both viral RNA replication and the release of nuclear proteins to the cytoplasm are greatly inhibited. Furthermore, nucleoside analogues that inhibit cellular and viral RNA synthesis impede the blockade of host mRNA translation, in addition to the release of nuclear proteins. Prevention of the shut-off of host mRNA translation by nucleoside analogues is not due to the inhibition of eIF2α phosphorylation, as this prevention is also observed in PKR(-/-) mouse embryonic fibroblasts that do not phosphorylate eIF2α after SINV infection. Collectively, our observations are consistent with the concept that for the inhibition of cellular protein synthesis to occur, viral RNA replication must take place at control levels, leading to the release of nuclear proteins to the cytoplasm. © 2014 John Wiley & Sons Ltd.

Cellular Chaperonin CCTγ Contributes to Rabies Virus Replication during Infection

PubMed Central

Zhang, Jinyang; Wu, Xiaopeng; Zan, Jie; Wu, Yongping; Ye, Chengjin; Ruan, Xizhen

2013-01-01

Rabies, as the oldest known infectious disease, remains a serious threat to public health worldwide. The eukaryotic cytosolic chaperonin TRiC/CCT complex facilitates the folding of proteins through ATP hydrolysis. Here, we investigated the expression, cellular localization, and function of neuronal CCTγ during neurotropic rabies virus (RABV) infection using mouse N2a cells as a model. Following RABV infection, 24 altered proteins were identified by using two-dimensional electrophoresis and mass spectrometry, including 20 upregulated proteins and 4 downregulated proteins. In mouse N2a cells infected with RABV or cotransfected with RABV genes encoding nucleoprotein (N) and phosphoprotein (P), confocal microscopy demonstrated that upregulated cellular CCTγ was colocalized with viral proteins N and P, which formed a hollow cricoid inclusion within the region around the nucleus. These inclusions, which correspond to Negri bodies (NBs), did not form in mouse N2a cells only expressing the viral protein N or P. Knockdown of CCTγ by lentivirus-mediated RNA interference led to significant inhibition of RABV replication. These results demonstrate that the complex consisting of viral proteins N and P recruits CCTγ to NBs and identify the chaperonin CCTγ as a host factor that facilitates intracellular RABV replication. This work illustrates how viruses can utilize cellular chaperonins and compartmentalization for their own benefit. PMID:23637400

Network-based function prediction and interactomics: the case for metabolic enzymes.

PubMed

Janga, S C; Díaz-Mejía, J Javier; Moreno-Hagelsieb, G

2011-01-01

As sequencing technologies increase in power, determining the functions of unknown proteins encoded by the DNA sequences so produced becomes a major challenge. Functional annotation is commonly done on the basis of amino-acid sequence similarity alone. Long after sequence similarity becomes undetectable by pair-wise comparison, profile-based identification of homologs can often succeed due to the conservation of position-specific patterns, important for a protein's three dimensional folding and function. Nevertheless, prediction of protein function from homology-driven approaches is not without problems. Homologous proteins might evolve different functions and the power of homology detection has already started to reach its maximum. Computational methods for inferring protein function, which exploit the context of a protein in cellular networks, have come to be built on top of homology-based approaches. These network-based functional inference techniques provide both a first hand hint into a proteins' functional role and offer complementary insights to traditional methods for understanding the function of uncharacterized proteins. Most recent network-based approaches aim to integrate diverse kinds of functional interactions to boost both coverage and confidence level. These techniques not only promise to solve the moonlighting aspect of proteins by annotating proteins with multiple functions, but also increase our understanding on the interplay between different functional classes in a cell. In this article we review the state of the art in network-based function prediction and describe some of the underlying difficulties and successes. Given the volume of high-throughput data that is being reported the time is ripe to employ these network-based approaches, which can be used to unravel the functions of the uncharacterized proteins accumulating in the genomic databases. © 2010 Elsevier Inc. All rights reserved.

Analysis of the Secretome and Identification of Novel Constituents from Culture Filtrate of Bacillus Calmette-Guérin Using High-resolution Mass Spectrometry*

PubMed Central

Zheng, Jianhua; Ren, Xianwen; Wei, Candong; Yang, Jian; Hu, Yongfeng; Liu, Liguo; Xu, Xingye; Wang, Jin; Jin, Qi

2013-01-01

Tuberculosis (TB) is an infectious bacterial disease that causes morbidity and mortality, especially in developing countries. Although its efficacy against TB has displayed a high degree of variability (0%–80%) in different trials, Mycobacterium bovis bacillus Calmette-Guérin (BCG) has been recognized as an important weapon for preventing TB worldwide for over 80 years. Because secreted proteins often play vital roles in the interaction between bacteria and host cells, the secretome of mycobacteria is considered to be an attractive reservoir of potential candidate antigens for the development of novel vaccines and diagnostic reagents. In this study, we performed a proteomic analysis of BCG culture filtrate proteins using SDS-PAGE and high-resolution Fourier transform mass spectrometry. In total, 239 proteins (1555 unique peptides) were identified, including 185 secreted proteins or lipoproteins. Furthermore, 17 novel protein products not annotated in the BCG database were detected and validated by means of RT-PCR at the transcriptional level. Additionally, the translational start sites of 52 proteins were confirmed, and 22 proteins were validated through extension of the translational start sites based on N-terminus-derived peptides. There are 103 secreted proteins that have not been reported in previous studies on the mycobacterial secretome and are unique to our study. The physicochemical characteristics of the secreted proteins were determined. Major components from the culture supernatant, including low-molecular-weight antigens, lipoproteins, Pro-Glu and Pro-Pro-Glu family proteins, and Mce family proteins, are discussed; some components represent potential predominant antigens in the humoral and cellular immune responses. PMID:23616670

Th1 stimulatory proteins of Leishmania donovani: comparative cellular and protective responses of rTriose phosphate isomerase, rProtein disulfide isomerase and rElongation factor-2 in combination with rHSP70 against visceral leishmaniasis.

PubMed

Jaiswal, Anil Kumar; Khare, Prashant; Joshi, Sumit; Kushawaha, Pramod Kumar; Sundar, Shyam; Dube, Anuradha

2014-01-01

In visceral leishmaniasis, the recovery from the disease is always associated with the generation of Th1-type of cellular responses. Based on this, we have previously identified several Th1-stimulatory proteins of Leishmania donovani -triose phosphate isomerase (TPI), protein disulfide isomerase (PDI) and elongation factor-2 (EL-2) etc. including heat shock protein 70 (HSP70) which induced Th1-type of cellular responses in both cured Leishmania patients/hamsters. Since, HSPs, being the logical targets for vaccines aimed at augmenting cellular immunity and can be early targets in the immune response against intracellular pathogens; they could be exploited as vaccine/adjuvant to induce long-term immunity more effectively. Therefore, in this study, we checked whether HSP70 can further enhance the immunogenicity and protective responses of the above said Th1-stimulatory proteins. Since, in most of the studies, immunogenicity of HSP70 of L. donovani was assessed in native condition, herein we generated recombinant HSP70 and tested its potential to stimulate immune responses in lymphocytes of cured Leishmania infected hamsters as well as in the peripheral blood mononuclear cells (PBMCs) of cured patients of VL either individually or in combination with above mentioned recombinant proteins. rLdHSP70 alone elicited strong cellular responses along with remarkable up-regulation of IFN-γ and IL-12 cytokines and extremely lower level of IL-4 and IL-10. Among the various combinations, rLdHSP70 + rLdPDI emerged as superior one augmenting improved cellular responses followed by rLdHSP70 + rLdEL-2. These combinations were further evaluated for its protective potential wherein rLdHSP70 + rLdPDI again conferred utmost protection (∼80%) followed by rLdHSP70 + rLdEL-2 (∼75%) and generated a strong cellular immune response with significant increase in the levels of iNOS transcript as well as IFN-γ and IL-12 cytokines which was further supported by the high level of IgG2 antibody in vaccinated animals. These observations indicated that vaccine(s) based on combination of HSP70 with Th1-stimulatory protein(s) may be a viable proposition against intracellular pathogens.

Th1 Stimulatory Proteins of Leishmania donovani: Comparative Cellular and Protective Responses of rTriose Phosphate Isomerase, rProtein Disulfide Isomerase and rElongation Factor-2 in Combination with rHSP70 against Visceral Leishmaniasis

PubMed Central

Jaiswal, Anil Kumar; Khare, Prashant; Joshi, Sumit; Kushawaha, Pramod Kumar; Sundar, Shyam; Dube, Anuradha

2014-01-01

In visceral leishmaniasis, the recovery from the disease is always associated with the generation of Th1-type of cellular responses. Based on this, we have previously identified several Th1-stimulatory proteins of Leishmania donovani -triose phosphate isomerase (TPI), protein disulfide isomerase (PDI) and elongation factor-2 (EL-2) etc. including heat shock protein 70 (HSP70) which induced Th1-type of cellular responses in both cured Leishmania patients/hamsters. Since, HSPs, being the logical targets for vaccines aimed at augmenting cellular immunity and can be early targets in the immune response against intracellular pathogens; they could be exploited as vaccine/adjuvant to induce long-term immunity more effectively. Therefore, in this study, we checked whether HSP70 can further enhance the immunogenicity and protective responses of the above said Th1-stimulatory proteins. Since, in most of the studies, immunogenicity of HSP70 of L. donovani was assessed in native condition, herein we generated recombinant HSP70 and tested its potential to stimulate immune responses in lymphocytes of cured Leishmania infected hamsters as well as in the peripheral blood mononuclear cells (PBMCs) of cured patients of VL either individually or in combination with above mentioned recombinant proteins. rLdHSP70 alone elicited strong cellular responses along with remarkable up-regulation of IFN-γ and IL-12 cytokines and extremely lower level of IL-4 and IL-10. Among the various combinations, rLdHSP70 + rLdPDI emerged as superior one augmenting improved cellular responses followed by rLdHSP70 + rLdEL-2. These combinations were further evaluated for its protective potential wherein rLdHSP70 + rLdPDI again conferred utmost protection (∼80%) followed by rLdHSP70 + rLdEL-2 (∼75%) and generated a strong cellular immune response with significant increase in the levels of iNOS transcript as well as IFN-γ and IL-12 cytokines which was further supported by the high level of IgG2 antibody in vaccinated animals. These observations indicated that vaccine(s) based on combination of HSP70 with Th1-stimulatory protein(s) may be a viable proposition against intracellular pathogens. PMID:25268700

Reduced native state stability in crowded cellular environment due to protein-protein interactions.

PubMed

Harada, Ryuhei; Tochio, Naoya; Kigawa, Takanori; Sugita, Yuji; Feig, Michael

2013-03-06

The effect of cellular crowding environments on protein structure and stability is a key issue in molecular and cellular biology. The classical view of crowding emphasizes the volume exclusion effect that generally favors compact, native states. Here, results from molecular dynamics simulations and NMR experiments show that protein crowders may destabilize native states via protein-protein interactions. In the model system considered here, mixtures of villin head piece and protein G at high concentrations, villin structures become increasingly destabilized upon increasing crowder concentrations. The denatured states observed in the simulation involve partial unfolding as well as more subtle conformational shifts. The unfolded states remain overall compact and only partially overlap with unfolded ensembles at high temperature and in the presence of urea. NMR measurements on the same systems confirm structural changes upon crowding based on changes of chemical shifts relative to dilute conditions. An analysis of protein-protein interactions and energetic aspects suggests the importance of enthalpic and solvation contributions to the crowding free energies that challenge an entropic-centered view of crowding effects.

Plant Abiotic Stress Proteomics: The Major Factors Determining Alterations in Cellular Proteome

PubMed Central

Kosová, Klára; Vítámvás, Pavel; Urban, Milan O.; Prášil, Ilja T.; Renaut, Jenny

2018-01-01

HIGHLIGHTS: Major environmental and genetic factors determining stress-related protein abundance are discussed.Major aspects of protein biological function including protein isoforms and PTMs, cellular localization and protein interactions are discussed.Functional diversity of protein isoforms and PTMs is discussed. Abiotic stresses reveal profound impacts on plant proteomes including alterations in protein relative abundance, cellular localization, post-transcriptional and post-translational modifications (PTMs), protein interactions with other protein partners, and, finally, protein biological functions. The main aim of the present review is to discuss the major factors determining stress-related protein accumulation and their final biological functions. A dynamics of stress response including stress acclimation to altered ambient conditions and recovery after the stress treatment is discussed. The results of proteomic studies aimed at a comparison of stress response in plant genotypes differing in stress adaptability reveal constitutively enhanced levels of several stress-related proteins (protective proteins, chaperones, ROS scavenging- and detoxification-related enzymes) in the tolerant genotypes with respect to the susceptible ones. Tolerant genotypes can efficiently adjust energy metabolism to enhanced needs during stress acclimation. Stress tolerance vs. stress susceptibility are relative terms which can reflect different stress-coping strategies depending on the given stress treatment. The role of differential protein isoforms and PTMs with respect to their biological functions in different physiological constraints (cellular compartments and interacting partners) is discussed. The importance of protein functional studies following high-throughput proteome analyses is presented in a broader context of plant biology. In summary, the manuscript tries to provide an overview of the major factors which have to be considered when interpreting data from proteomic studies on stress-treated plants. PMID:29472941

ISG15 in the tumorigenesis and treatment of cancer: An emerging role in malignancies of the digestive system

PubMed Central

Zuo, Chaohui; Sheng, Xinyi; Ma, Min; Xia, Man; Ouyang, Linda

2016-01-01

The interferon-stimulated gene 15 ubiquitin-like modifier (ISG15) encodes an IFN-inducible, ubiquitin-like protein. The ISG15 protein forms conjugates with numerous cellular proteins that are involved in a multitude of cellular functions, including interferon-induced immune responses and the regulation of cellular protein turnover. The expression of ISG15 and ISG15-mediated conjugation has been implicated in a wide range of human tumors and cancer cell lines, but the roles of ISG15 in tumorigenesis and responses to anticancer treatments remain largely unknown. In this review, we discuss the findings of recent studies with regard to the role of ISG15 pathways in cancers of the digestive system. PMID:27626310

ISG15 in the tumorigenesis and treatment of cancer: An emerging role in malignancies of the digestive system.

PubMed

Zuo, Chaohui; Sheng, Xinyi; Ma, Min; Xia, Man; Ouyang, Linda

2016-11-08

The interferon-stimulated gene 15 ubiquitin-like modifier (ISG15) encodes an IFN-inducible, ubiquitin-like protein. The ISG15 protein forms conjugates with numerous cellular proteins that are involved in a multitude of cellular functions, including interferon-induced immune responses and the regulation of cellular protein turnover. The expression of ISG15 and ISG15-mediated conjugation has been implicated in a wide range of human tumors and cancer cell lines, but the roles of ISG15 in tumorigenesis and responses to anticancer treatments remain largely unknown. In this review, we discuss the findings of recent studies with regard to the role of ISG15 pathways in cancers of the digestive system.

Fe-S Cluster Hsp70 Chaperones: The ATPase Cycle and Protein Interactions.

PubMed

Dutkiewicz, Rafal; Nowak, Malgorzata; Craig, Elizabeth A; Marszalek, Jaroslaw

2017-01-01

Hsp70 chaperones and their obligatory J-protein cochaperones function together in many cellular processes. Via cycles of binding to short stretches of exposed amino acids on substrate proteins, Hsp70/J-protein chaperones not only facilitate protein folding but also drive intracellular protein transport, biogenesis of cellular structures, and disassembly of protein complexes. The biogenesis of iron-sulfur (Fe-S) clusters is one of the critical cellular processes that require Hsp70/J-protein action. Fe-S clusters are ubiquitous cofactors critical for activity of proteins performing diverse functions in, for example, metabolism, RNA/DNA transactions, and environmental sensing. This biogenesis process can be divided into two sequential steps: first, the assembly of an Fe-S cluster on a conserved scaffold protein, and second, the transfer of the cluster from the scaffold to a recipient protein. The second step involves Hsp70/J-protein chaperones. Via binding to the scaffold, chaperones enable cluster transfer to recipient proteins. In eukaryotic cells mitochondria have a key role in Fe-S cluster biogenesis. In this review, we focus on methods that enabled us to dissect protein interactions critical for the function of Hsp70/J-protein chaperones in the mitochondrial process of Fe-S cluster biogenesis in the yeast Saccharomyces cerevisiae. © 2017 Elsevier Inc. All rights reserved.

Biomolecular interactions modulate macromolecular structure and dynamics in atomistic model of a bacterial cytoplasm

PubMed Central

Yu, Isseki; Mori, Takaharu; Ando, Tadashi; Harada, Ryuhei; Jung, Jaewoon; Sugita, Yuji; Feig, Michael

2016-01-01

Biological macromolecules function in highly crowded cellular environments. The structure and dynamics of proteins and nucleic acids are well characterized in vitro, but in vivo crowding effects remain unclear. Using molecular dynamics simulations of a comprehensive atomistic model cytoplasm we found that protein-protein interactions may destabilize native protein structures, whereas metabolite interactions may induce more compact states due to electrostatic screening. Protein-protein interactions also resulted in significant variations in reduced macromolecular diffusion under crowded conditions, while metabolites exhibited significant two-dimensional surface diffusion and altered protein-ligand binding that may reduce the effective concentration of metabolites and ligands in vivo. Metabolic enzymes showed weak non-specific association in cellular environments attributed to solvation and entropic effects. These effects are expected to have broad implications for the in vivo functioning of biomolecules. This work is a first step towards physically realistic in silico whole-cell models that connect molecular with cellular biology. DOI: http://dx.doi.org/10.7554/eLife.19274.001 PMID:27801646

Proteomic composition of Nipah virus-like particles.

PubMed

Vera-Velasco, Natalia Mara; García-Murria, Maria Jesús; Sánchez Del Pino, Manuel M; Mingarro, Ismael; Martinez-Gil, Luis

2018-02-10

Virions are often described as virus-only entities with no cellular components with the exception of the lipids in their membranes. However, advances in proteomics are revealing substantial amounts of host proteins in the viral particles. In the case of Nipah virus (NiV), the viral components in the virion have been known for some time. Nonetheless, no information has been obtained regarding the cellular proteins in the viral particles. To address this question, we produced Virus-Like Particles (VLPs) for NiV by expressing the F, G and M proteins in human-derived cells. Next, the proteomic content in these VLPs was analyzed by LC-MS/MS. We identified 67 human proteins including soluble and membrane-bound proteins involved in vesicle sorting and transport. Interestingly, many of them have been reported to interact with other viruses. Finally, thanks to the semi-quantitative nature of our data we were able to estimate the ratio among F, G and M proteins and also the ratio between cellular and viral proteins in the VLPs. We believe our data contribute to the better understanding of NiV life cycle and might facilitate future attempts for developing antiviral agents and the design of further experimental studies for this deadly infection. Traditionally viral particles have been described as pure entities carrying only viral-derived proteins. Advances in proteomics are changing this simplified view. Host proteins have been identified in many viruses (especially in enveloped viruses). These cell-derived proteins participate in multiple steps in the viral life cycle and might be as important for the survival of the virus as any other viral-encoded protein. In this work, we analyze utilizing LC-MS/MS the cellular proteins incorporated or bound to the virions of Nipah virus (NiV), an emerging, highly pathogenic, zoonotic virus from the Paramyxoviridiae family. Furthermore, we analyzed the ratio between cellular and viral proteins and among the viral F, G and M proteins in the viral particles. The characterization of the Nipah virus-human interactions occurring in the virion might facilitate the development of new therapeutic and prophylactic therapies for this viral illness. Copyright © 2017 Elsevier B.V. All rights reserved.

Purification of infectious human herpesvirus 6A virions and association of host cell proteins

PubMed Central

Hammarstedt, Maria; Ahlqvist, Jenny; Jacobson, Steven; Garoff, Henrik; Fogdell-Hahn, Anna

2007-01-01

Background Viruses that are incorporating host cell proteins might trigger autoimmune diseases. It is therefore of interest to identify possible host proteins associated with viruses, especially for enveloped viruses that have been suggested to play a role in autoimmune diseases, like human herpesvirus 6A (HHV-6A) in multiple sclerosis (MS). Results We have established a method for rapid and morphology preserving purification of HHV-6A virions, which in combination with parallel analyses with background control material released from mock-infected cells facilitates qualitative and quantitative investigations of the protein content of HHV-6A virions. In our iodixanol gradient purified preparation, we detected high levels of viral DNA by real-time PCR and viral proteins by metabolic labelling, silver staining and western blots. In contrast, the background level of cellular contamination was low in the purified samples as demonstrated by the silver staining and metabolic labelling analyses. Western blot analyses showed that the cellular complement protein CD46, the receptor for HHV-6A, is associated with the purified and infectious virions. Also, the cellular proteins clathrin, ezrin and Tsg101 are associated with intact HHV-6A virions. Conclusion Cellular proteins are associated with HHV-6A virions. The relevance of the association in disease and especially in autoimmunity will be further investigated. PMID:17949490

Signal sequence and keyword trap in silico for selection of full-length human cDNAs encoding secretion or membrane proteins from oligo-capped cDNA libraries.

PubMed

Otsuki, Tetsuji; Ota, Toshio; Nishikawa, Tetsuo; Hayashi, Koji; Suzuki, Yutaka; Yamamoto, Jun-ichi; Wakamatsu, Ai; Kimura, Kouichi; Sakamoto, Katsuhiko; Hatano, Naoto; Kawai, Yuri; Ishii, Shizuko; Saito, Kaoru; Kojima, Shin-ichi; Sugiyama, Tomoyasu; Ono, Tetsuyoshi; Okano, Kazunori; Yoshikawa, Yoko; Aotsuka, Satoshi; Sasaki, Naokazu; Hattori, Atsushi; Okumura, Koji; Nagai, Keiichi; Sugano, Sumio; Isogai, Takao

2005-01-01

We have developed an in silico method of selection of human full-length cDNAs encoding secretion or membrane proteins from oligo-capped cDNA libraries. Fullness rates were increased to about 80% by combination of the oligo-capping method and ATGpr, software for prediction of translation start point and the coding potential. Then, using 5'-end single-pass sequences, cDNAs having the signal sequence were selected by PSORT ('signal sequence trap'). We also applied 'secretion or membrane protein-related keyword trap' based on the result of BLAST search against the SWISS-PROT database for the cDNAs which could not be selected by PSORT. Using the above procedures, 789 cDNAs were primarily selected and subjected to full-length sequencing, and 334 of these cDNAs were finally selected as novel. Most of the cDNAs (295 cDNAs: 88.3%) were predicted to encode secretion or membrane proteins. In particular, 165(80.5%) of the 205 cDNAs selected by PSORT were predicted to have signal sequences, while 70 (54.2%) of the 129 cDNAs selected by 'keyword trap' preserved the secretion or membrane protein-related keywords. Many important cDNAs were obtained, including transporters, receptors, and ligands, involved in significant cellular functions. Thus, an efficient method of selecting secretion or membrane protein-encoding cDNAs was developed by combining the above four procedures.

LC-MS/MS based proteomic analysis and functional inference of hypothetical proteins in Desulfovibrio vulgaris

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhang, Weiwen; Culley, David E.; Gritsenko, Marina A.

2006-11-03

ABSTRACT In the previous study, the whole-genome gene expression profiles of D. vulgaris in response to oxidative stress and heat shock were determined. The results showed 24-28% of the responsive genes were hypothetical proteins that have not been experimentally characterized or whose function can not be deduced by simple sequence comparison. To further explore the protecting mechanisms employed in D. vulgaris against the oxidative stress and heat shock, attempt was made in this study to infer functions of these hypothetical proteins by phylogenomic profiling along with detailed sequence comparison against various publicly available databases. By this approach we were abletomore » assign possible functions to 25 responsive hypothetical proteins. The findings included that DVU0725, induced by oxidative stress, may be involved in lipopolysaccharide biosynthesis, implying that the alternation of lipopolysaccharide on cell surface might service as a mechanism against oxidative stress in D. vulgaris. In addition, two responsive proteins, DVU0024 encoding a putative transcriptional regulator and DVU1670 encoding predicted redox protein, were sharing co-evolution atterns with rubrerythrin in Archaeoglobus fulgidus and Clostridium perfringens, respectively, implying that they might be part of the stress response and protective systems in D. vulgaris. The study demonstrated that phylogenomic profiling is a useful tool in interpretation of experimental genomics data, and also provided further insight on cellular response to oxidative stress and heat shock in D. vulgaris.« less

hPDI: a database of experimental human protein-DNA interactions.

PubMed

Xie, Zhi; Hu, Shaohui; Blackshaw, Seth; Zhu, Heng; Qian, Jiang

2010-01-15

The human protein DNA Interactome (hPDI) database holds experimental protein-DNA interaction data for humans identified by protein microarray assays. The unique characteristics of hPDI are that it contains consensus DNA-binding sequences not only for nearly 500 human transcription factors but also for >500 unconventional DNA-binding proteins, which are completely uncharacterized previously. Users can browse, search and download a subset or the entire data via a web interface. This database is freely accessible for any academic purposes. http://bioinfo.wilmer.jhu.edu/PDI/.

MitoNuc: a database of nuclear genes coding for mitochondrial proteins. Update 2002.

PubMed

Attimonelli, Marcella; Catalano, Domenico; Gissi, Carmela; Grillo, Giorgio; Licciulli, Flavio; Liuni, Sabino; Santamaria, Monica; Pesole, Graziano; Saccone, Cecilia

2002-01-01

Mitochondria, besides their central role in energy metabolism, have recently been found to be involved in a number of basic processes of cell life and to contribute to the pathogenesis of many degenerative diseases. All functions of mitochondria depend on the interaction of nuclear and organelle genomes. Mitochondrial genomes have been extensively sequenced and analysed and data have been collected in several specialised databases. In order to collect information on nuclear coded mitochondrial proteins we developed MitoNuc, a database containing detailed information on sequenced nuclear genes coding for mitochondrial proteins in Metazoa. The MitoNuc database can be retrieved through SRS and is available via the web site http://bighost.area.ba.cnr.it/mitochondriome where other mitochondrial databases developed by our group, the complete list of the sequenced mitochondrial genomes, links to other mitochondrial sites and related information, are available. The MitoAln database, related to MitoNuc in the previous release, reporting the multiple alignments of the relevant homologous protein coding regions, is no longer supported in the present release. In order to keep the links among entries in MitoNuc from homologous proteins, a new field in the database has been defined: the cluster identifier, an alpha numeric code used to identify each cluster of homologous proteins. A comment field derived from the corresponding SWISS-PROT entry has been introduced; this reports clinical data related to dysfunction of the protein. The logic scheme of MitoNuc database has been implemented in the ORACLE DBMS. This will allow the end-users to retrieve data through a friendly interface that will be soon implemented.

ChlamyCyc: an integrative systems biology database and web-portal for Chlamydomonas reinhardtii.

PubMed

May, Patrick; Christian, Jan-Ole; Kempa, Stefan; Walther, Dirk

2009-05-04

The unicellular green alga Chlamydomonas reinhardtii is an important eukaryotic model organism for the study of photosynthesis and plant growth. In the era of modern high-throughput technologies there is an imperative need to integrate large-scale data sets from high-throughput experimental techniques using computational methods and database resources to provide comprehensive information about the molecular and cellular organization of a single organism. In the framework of the German Systems Biology initiative GoFORSYS, a pathway database and web-portal for Chlamydomonas (ChlamyCyc) was established, which currently features about 250 metabolic pathways with associated genes, enzymes, and compound information. ChlamyCyc was assembled using an integrative approach combining the recently published genome sequence, bioinformatics methods, and experimental data from metabolomics and proteomics experiments. We analyzed and integrated a combination of primary and secondary database resources, such as existing genome annotations from JGI, EST collections, orthology information, and MapMan classification. ChlamyCyc provides a curated and integrated systems biology repository that will enable and assist in systematic studies of fundamental cellular processes in Chlamydomonas. The ChlamyCyc database and web-portal is freely available under http://chlamycyc.mpimp-golm.mpg.de.

Using SQL Databases for Sequence Similarity Searching and Analysis.

PubMed

Pearson, William R; Mackey, Aaron J

2017-09-13

Relational databases can integrate diverse types of information and manage large sets of similarity search results, greatly simplifying genome-scale analyses. By focusing on taxonomic subsets of sequences, relational databases can reduce the size and redundancy of sequence libraries and improve the statistical significance of homologs. In addition, by loading similarity search results into a relational database, it becomes possible to explore and summarize the relationships between all of the proteins in an organism and those in other biological kingdoms. This unit describes how to use relational databases to improve the efficiency of sequence similarity searching and demonstrates various large-scale genomic analyses of homology-related data. It also describes the installation and use of a simple protein sequence database, seqdb_demo, which is used as a basis for the other protocols. The unit also introduces search_demo, a database that stores sequence similarity search results. The search_demo database is then used to explore the evolutionary relationships between E. coli proteins and proteins in other organisms in a large-scale comparative genomic analysis. © 2017 by John Wiley & Sons, Inc. Copyright © 2017 John Wiley & Sons, Inc.

SNAPPI-DB: a database and API of Structures, iNterfaces and Alignments for Protein–Protein Interactions

PubMed Central

Jefferson, Emily R.; Walsh, Thomas P.; Roberts, Timothy J.; Barton, Geoffrey J.

2007-01-01

SNAPPI-DB, a high performance database of Structures, iNterfaces and Alignments of Protein–Protein Interactions, and its associated Java Application Programming Interface (API) is described. SNAPPI-DB contains structural data, down to the level of atom co-ordinates, for each structure in the Protein Data Bank (PDB) together with associated data including SCOP, CATH, Pfam, SWISSPROT, InterPro, GO terms, Protein Quaternary Structures (PQS) and secondary structure information. Domain–domain interactions are stored for multiple domain definitions and are classified by their Superfamily/Family pair and interaction interface. Each set of classified domain–domain interactions has an associated multiple structure alignment for each partner. The API facilitates data access via PDB entries, domains and domain–domain interactions. Rapid development, fast database access and the ability to perform advanced queries without the requirement for complex SQL statements are provided via an object oriented database and the Java Data Objects (JDO) API. SNAPPI-DB contains many features which are not available in other databases of structural protein–protein interactions. It has been applied in three studies on the properties of protein–protein interactions and is currently being employed to train a protein–protein interaction predictor and a functional residue predictor. The database, API and manual are available for download at: . PMID:17202171

SALAD database: a motif-based database of protein annotations for plant comparative genomics

PubMed Central

Mihara, Motohiro; Itoh, Takeshi; Izawa, Takeshi

2010-01-01

Proteins often have several motifs with distinct evolutionary histories. Proteins with similar motifs have similar biochemical properties and thus related biological functions. We constructed a unique comparative genomics database termed the SALAD database (http://salad.dna.affrc.go.jp/salad/) from plant-genome-based proteome data sets. We extracted evolutionarily conserved motifs by MEME software from 209 529 protein-sequence annotation groups selected by BLASTP from the proteome data sets of 10 species: rice, sorghum, Arabidopsis thaliana, grape, a lycophyte, a moss, 3 algae, and yeast. Similarity clustering of each protein group was performed by pairwise scoring of the motif patterns of the sequences. The SALAD database provides a user-friendly graphical viewer that displays a motif pattern diagram linked to the resulting bootstrapped dendrogram for each protein group. Amino-acid-sequence-based and nucleotide-sequence-based phylogenetic trees for motif combination alignment, a logo comparison diagram for each clade in the tree, and a Pfam-domain pattern diagram are also available. We also developed a viewer named ‘SALAD on ARRAYs’ to view arbitrary microarray data sets of paralogous genes linked to the same dendrogram in a window. The SALAD database is a powerful tool for comparing protein sequences and can provide valuable hints for biological analysis. PMID:19854933

SALAD database: a motif-based database of protein annotations for plant comparative genomics.

PubMed

Mihara, Motohiro; Itoh, Takeshi; Izawa, Takeshi

2010-01-01

Proteins often have several motifs with distinct evolutionary histories. Proteins with similar motifs have similar biochemical properties and thus related biological functions. We constructed a unique comparative genomics database termed the SALAD database (http://salad.dna.affrc.go.jp/salad/) from plant-genome-based proteome data sets. We extracted evolutionarily conserved motifs by MEME software from 209,529 protein-sequence annotation groups selected by BLASTP from the proteome data sets of 10 species: rice, sorghum, Arabidopsis thaliana, grape, a lycophyte, a moss, 3 algae, and yeast. Similarity clustering of each protein group was performed by pairwise scoring of the motif patterns of the sequences. The SALAD database provides a user-friendly graphical viewer that displays a motif pattern diagram linked to the resulting bootstrapped dendrogram for each protein group. Amino-acid-sequence-based and nucleotide-sequence-based phylogenetic trees for motif combination alignment, a logo comparison diagram for each clade in the tree, and a Pfam-domain pattern diagram are also available. We also developed a viewer named 'SALAD on ARRAYs' to view arbitrary microarray data sets of paralogous genes linked to the same dendrogram in a window. The SALAD database is a powerful tool for comparing protein sequences and can provide valuable hints for biological analysis.

2'-5'-Oligoadenylate Synthetase-Like Protein Inhibits Respiratory Syncytial Virus Replication and Is Targeted by the Viral Nonstructural Protein 1.

PubMed

Dhar, Jayeeta; Cuevas, Rolando A; Goswami, Ramansu; Zhu, Jianzhong; Sarkar, Saumendra N; Barik, Sailen

2015-10-01

2'-5'-Oligoadenylate synthetase-like protein (OASL) is an interferon-inducible antiviral protein. Here we describe differential inhibitory activities of human OASL and the two mouse OASL homologs against respiratory syncytial virus (RSV) replication. Interestingly, nonstructural protein 1 (NS1) of RSV promoted proteasome-dependent degradation of specific OASL isoforms. We conclude that OASL acts as a cellular antiviral protein and that RSV NS1 suppresses this function to evade cellular innate immunity and allow virus growth. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

MEGADOCK-Web: an integrated database of high-throughput structure-based protein-protein interaction predictions.

PubMed

Hayashi, Takanori; Matsuzaki, Yuri; Yanagisawa, Keisuke; Ohue, Masahito; Akiyama, Yutaka

2018-05-08

Protein-protein interactions (PPIs) play several roles in living cells, and computational PPI prediction is a major focus of many researchers. The three-dimensional (3D) structure and binding surface are important for the design of PPI inhibitors. Therefore, rigid body protein-protein docking calculations for two protein structures are expected to allow elucidation of PPIs different from known complexes in terms of 3D structures because known PPI information is not explicitly required. We have developed rapid PPI prediction software based on protein-protein docking, called MEGADOCK. In order to fully utilize the benefits of computational PPI predictions, it is necessary to construct a comprehensive database to gather prediction results and their predicted 3D complex structures and to make them easily accessible. Although several databases exist that provide predicted PPIs, the previous databases do not contain a sufficient number of entries for the purpose of discovering novel PPIs. In this study, we constructed an integrated database of MEGADOCK PPI predictions, named MEGADOCK-Web. MEGADOCK-Web provides more than 10 times the number of PPI predictions than previous databases and enables users to conduct PPI predictions that cannot be found in conventional PPI prediction databases. In MEGADOCK-Web, there are 7528 protein chains and 28,331,628 predicted PPIs from all possible combinations of those proteins. Each protein structure is annotated with PDB ID, chain ID, UniProt AC, related KEGG pathway IDs, and known PPI pairs. Additionally, MEGADOCK-Web provides four powerful functions: 1) searching precalculated PPI predictions, 2) providing annotations for each predicted protein pair with an experimentally known PPI, 3) visualizing candidates that may interact with the query protein on biochemical pathways, and 4) visualizing predicted complex structures through a 3D molecular viewer. MEGADOCK-Web provides a huge amount of comprehensive PPI predictions based on docking calculations with biochemical pathways and enables users to easily and quickly assess PPI feasibilities by archiving PPI predictions. MEGADOCK-Web also promotes the discovery of new PPIs and protein functions and is freely available for use at http://www.bi.cs.titech.ac.jp/megadock-web/ .

ProteinWorldDB: querying radical pairwise alignments among protein sets from complete genomes.

PubMed

Otto, Thomas Dan; Catanho, Marcos; Tristão, Cristian; Bezerra, Márcia; Fernandes, Renan Mathias; Elias, Guilherme Steinberger; Scaglia, Alexandre Capeletto; Bovermann, Bill; Berstis, Viktors; Lifschitz, Sergio; de Miranda, Antonio Basílio; Degrave, Wim

2010-03-01

Many analyses in modern biological research are based on comparisons between biological sequences, resulting in functional, evolutionary and structural inferences. When large numbers of sequences are compared, heuristics are often used resulting in a certain lack of accuracy. In order to improve and validate results of such comparisons, we have performed radical all-against-all comparisons of 4 million protein sequences belonging to the RefSeq database, using an implementation of the Smith-Waterman algorithm. This extremely intensive computational approach was made possible with the help of World Community Grid, through the Genome Comparison Project. The resulting database, ProteinWorldDB, which contains coordinates of pairwise protein alignments and their respective scores, is now made available. Users can download, compare and analyze the results, filtered by genomes, protein functions or clusters. ProteinWorldDB is integrated with annotations derived from Swiss-Prot, Pfam, KEGG, NCBI Taxonomy database and gene ontology. The database is a unique and valuable asset, representing a major effort to create a reliable and consistent dataset of cross-comparisons of the whole protein content encoded in hundreds of completely sequenced genomes using a rigorous dynamic programming approach. The database can be accessed through http://proteinworlddb.org

LocSigDB: a database of protein localization signals

PubMed Central

Negi, Simarjeet; Pandey, Sanjit; Srinivasan, Satish M.; Mohammed, Akram; Guda, Chittibabu

2015-01-01

LocSigDB (http://genome.unmc.edu/LocSigDB/) is a manually curated database of experimental protein localization signals for eight distinct subcellular locations; primarily in a eukaryotic cell with brief coverage of bacterial proteins. Proteins must be localized at their appropriate subcellular compartment to perform their desired function. Mislocalization of proteins to unintended locations is a causative factor for many human diseases; therefore, collection of known sorting signals will help support many important areas of biomedical research. By performing an extensive literature study, we compiled a collection of 533 experimentally determined localization signals, along with the proteins that harbor such signals. Each signal in the LocSigDB is annotated with its localization, source, PubMed references and is linked to the proteins in UniProt database along with the organism information that contain the same amino acid pattern as the given signal. From LocSigDB webserver, users can download the whole database or browse/search for data using an intuitive query interface. To date, LocSigDB is the most comprehensive compendium of protein localization signals for eight distinct subcellular locations. Database URL: http://genome.unmc.edu/LocSigDB/ PMID:25725059

Clusterin in the protein corona plays a key role in the stealth effect of nanoparticles against phagocytes.

PubMed

Aoyama, Michihiko; Hata, Katsutomo; Higashisaka, Kazuma; Nagano, Kazuya; Yoshioka, Yasuo; Tsutsumi, Yasuo

2016-11-25

In biological fluids, nanoparticles interact with biological components such as proteins, and a layer called the "protein corona" forms around the nanoparticles. It is believed that the composition of the protein corona affects the cellular uptake and in vivo biodistribution of nanoparticles; however, the key proteins of the protein corona that control the biological fate of nanoparticles remain unclear. Recently, it was reported that clusterin binding to pegylated nanoparticles is important for the stealth effect of pegylated nanoparticles in phagocytes. However, the effect of clusterin on non-pegylated nanoparticles is unknown, although it is known that clusterin is present in the protein corona of non-pegylated nanoparticles. Here, we assessed the stealth effect of clusterin in the corona of non-pegylated silver nanoparticles and silica nanoparticles. We found that serum- and plasma-protein corona inhibited the cellular uptake of silver nanoparticles and silica nanoparticles in phagocytes and that the plasma-protein corona showed a greater stealth effect compared with the serum-protein corona. Clusterin was present in both the serum- and plasma-protein corona, but was present at a higher level in the plasma-protein corona than in the serum-protein corona. Clusterin binding to silver nanoparticles and silica nanoparticles suppressed the cellular uptake of nanoparticles in human macrophage-like cells (THP-1 cells). Although further studies are required to determine how clusterin suppresses non-specific cellular uptake in phagocytes, our data suggest that clusterin plays a key role in the stealth effect of not only pegylated nanoparticles but also non-pegylated nanoparticles. Copyright © 2016 Elsevier Inc. All rights reserved.

Thermosensitivity of growth is determined by chaperone-mediated proteome reallocation

PubMed Central

Chen, Ke; Gao, Ye; Mih, Nathan; O’Brien, Edward J.; Yang, Laurence; Palsson, Bernhard O.

2017-01-01

Maintenance of a properly folded proteome is critical for bacterial survival at notably different growth temperatures. Understanding the molecular basis of thermoadaptation has progressed in two main directions, the sequence and structural basis of protein thermostability and the mechanistic principles of protein quality control assisted by chaperones. Yet we do not fully understand how structural integrity of the entire proteome is maintained under stress and how it affects cellular fitness. To address this challenge, we reconstruct a genome-scale protein-folding network for Escherichia coli and formulate a computational model, FoldME, that provides statistical descriptions of multiscale cellular response consistent with many datasets. FoldME simulations show (i) that the chaperones act as a system when they respond to unfolding stress rather than achieving efficient folding of any single component of the proteome, (ii) how the proteome is globally balanced between chaperones for folding and the complex machinery synthesizing the proteins in response to perturbation, (iii) how this balancing determines growth rate dependence on temperature and is achieved through nonspecific regulation, and (iv) how thermal instability of the individual protein affects the overall functional state of the proteome. Overall, these results expand our view of cellular regulation, from targeted specific control mechanisms to global regulation through a web of nonspecific competing interactions that modulate the optimal reallocation of cellular resources. The methodology developed in this study enables genome-scale integration of environment-dependent protein properties and a proteome-wide study of cellular stress responses. PMID:29073085

Single-cell protein secretomic signatures as potential correlates to tumor cell lineage evolution and cell–cell interaction

PubMed Central

Kwak, Minsuk; Mu, Luye; Lu, Yao; Chen, Jonathan J.; Brower, Kara; Fan, Rong

2013-01-01

Secreted proteins including cytokines, chemokines, and growth factors represent important functional regulators mediating a range of cellular behavior and cell–cell paracrine/autocrine signaling, e.g., in the immunological system (Rothenberg, 2007), tumor microenvironment (Hanahan and Weinberg, 2011), or stem cell niche (Gnecchi etal., 2008). Detection of these proteins is of great value not only in basic cell biology but also for diagnosis and therapeutic monitoring of human diseases such as cancer. However, due to co-production of multiple effector proteins from a single cell, referred to as polyfunctionality, it is biologically informative to measure a panel of secreted proteins, or secretomic signature, at the level of single cells. Recent evidence further indicates that a genetically identical cell population can give rise to diverse phenotypic differences (Niepel etal., 2009). Non-genetic heterogeneity is also emerging as a potential barrier to accurate monitoring of cellular immunity and effective pharmacological therapies (Cohen etal., 2008; Gascoigne and Taylor, 2008), but can hardly assessed using conventional approaches that do not examine cellular phenotype at the functional level. It is known that cytokines, for example, in the immune system define the effector functions and lineage differentiation of immune cells. In this article, we hypothesize that protein secretion profile may represent a universal measure to identify the definitive correlate in the larger context of cellular functions to dissect cellular heterogeneity and evolutionary lineage relationship in human cancer. PMID:23390614

Therapeutic intervention at cellular quality control systems in Alzheimer's and Parkinson's diseases.

PubMed

Arduino, Daniela M; Esteves, A Raquel; Silva, Diana F F; Martins-Branco, Diogo; Santos, Daniel; Pimentel, Diana F Gomes; Cardoso, Sandra M

2011-01-01

Cellular homeostasis relies on quality control systems so that damaged biologic structures are either repaired or degraded and entirely replaced by newly formed proteins or even organelles. The clearance of dysfunctional cellular structures in long-lived postmitotic cells, like neurons, is essential to eliminate, per example, defective mitochondria, lipofuscin-loaded lysosomes and oxidized proteins. Short-lived proteins are degraded mainly by proteases and proteasomes whether most long-lived proteins and all organelles are digested by autophagy in the lysosomes. Recently, it an interplay was established between the ubiquitin-proteasome system and macroautophagy, so that both degradative mechanisms compensate for each other. In this article we describe each of these clearance systems and their contribution to neuronal quality control. We will highlight some of the findings that provide evidence for the dysfunction of these systems in Alzheimer's and Parkinson's diseases. Ultimately, we provide an outline on potential therapeutic interventions based on the modulation of cellular degradative systems.

Rice proteome analysis: a step toward functional analysis of the rice genome.

PubMed

Komatsu, Setsuko; Tanaka, Naoki

2005-03-01

The technique of proteome analysis using 2-DE has the power to monitor global changes that occur in the protein complement of tissues and subcellular compartments. In this review, we describe construction of the rice proteome database, the cataloging of rice proteins, and the functional characterization of some of the proteins identified. Initially, proteins extracted from various tissues and organelles were separated by 2-DE and an image analyzer was used to construct a display or reference map of the proteins. The rice proteome database currently contains 23 reference maps based on 2-DE of proteins from different rice tissues and subcellular compartments. These reference maps comprise 13 129 rice proteins, and the amino acid sequences of 5092 of these proteins are entered in the database. Major proteins involved in growth or stress responses have been identified by using a proteomics approach and some of these proteins have unique functions. Furthermore, initial work has also begun on analyzing the phosphoproteome and protein-protein interactions in rice. The information obtained from the rice proteome database will aid in the molecular cloning of rice genes and in predicting the function of unknown proteins.

Databases and Associated Tools for Glycomics and Glycoproteomics.

PubMed

Lisacek, Frederique; Mariethoz, Julien; Alocci, Davide; Rudd, Pauline M; Abrahams, Jodie L; Campbell, Matthew P; Packer, Nicolle H; Ståhle, Jonas; Widmalm, Göran; Mullen, Elaine; Adamczyk, Barbara; Rojas-Macias, Miguel A; Jin, Chunsheng; Karlsson, Niclas G

2017-01-01

The access to biodatabases for glycomics and glycoproteomics has proven to be essential for current glycobiological research. This chapter presents available databases that are devoted to different aspects of glycobioinformatics. This includes oligosaccharide sequence databases, experimental databases, 3D structure databases (of both glycans and glycorelated proteins) and association of glycans with tissue, disease, and proteins. Specific search protocols are also provided using tools associated with experimental databases for converting primary glycoanalytical data to glycan structural information. In particular, researchers using glycoanalysis methods by U/HPLC (GlycoBase), MS (GlycoWorkbench, UniCarb-DB, GlycoDigest), and NMR (CASPER) will benefit from this chapter. In addition we also include information on how to utilize glycan structural information to query databases that associate glycans with proteins (UniCarbKB) and with interactions with pathogens (SugarBind).

GALT protein database: querying structural and functional features of GALT enzyme.

PubMed

d'Acierno, Antonio; Facchiano, Angelo; Marabotti, Anna

2014-09-01

Knowledge of the impact of variations on protein structure can enhance the comprehension of the mechanisms of genetic diseases related to that protein. Here, we present a new version of GALT Protein Database, a Web-accessible data repository for the storage and interrogation of structural effects of variations of the enzyme galactose-1-phosphate uridylyltransferase (GALT), the impairment of which leads to classic Galactosemia, a rare genetic disease. This new version of this database now contains the models of 201 missense variants of GALT enzyme, including heterozygous variants, and it allows users not only to retrieve information about the missense variations affecting this protein, but also to investigate their impact on substrate binding, intersubunit interactions, stability, and other structural features. In addition, it allows the interactive visualization of the models of variants collected into the database. We have developed additional tools to improve the use of the database by nonspecialized users. This Web-accessible database (http://bioinformatica.isa.cnr.it/GALT/GALT2.0) represents a model of tools potentially suitable for application to other proteins that are involved in human pathologies and that are subjected to genetic variations. © 2014 WILEY PERIODICALS, INC.

Proteome of Caulobacter crescentus cell cycle publicly accessible on SWICZ server.

PubMed

Vohradsky, Jiri; Janda, Ivan; Grünenfelder, Björn; Berndt, Peter; Röder, Daniel; Langen, Hanno; Weiser, Jaroslav; Jenal, Urs

2003-10-01

Here we present the Swiss-Czech Proteomics Server (SWICZ), which hosts the proteomic database summarizing information about the cell cycle of the aquatic bacterium Caulobacter crescentus. The database provides a searchable tool for easy access of global protein synthesis and protein stability data as examined during the C. crescentus cell cycle. Protein synthesis data collected from five different cell cycle stages were determined for each protein spot as a relative value of the total amount of [(35)S]methionine incorporation. Protein stability of pulse-labeled extracts were measured during a chase period equivalent to one cell cycle unit. Quantitative information for individual proteins together with descriptive data such as protein identities, apparent molecular masses and isoelectric points, were combined with information on protein function, genomic context, and the cell cycle stage, and were then assembled in a relational database with a world wide web interface (http://proteom.biomed.cas.cz), which allows the database records to be searched and displays the recovered information. A total of 1250 protein spots were reproducibly detected on two-dimensional gel electropherograms, 295 of which were identified by mass spectroscopy. The database is accessible either through clickable two-dimensional gel electrophoretic maps or by means of a set of dedicated search engines. Basic characterization of the experimental procedures, data processing, and a comprehensive description of the web site are presented. In its current state, the SWICZ proteome database provides a platform for the incorporation of new data emerging from extended functional studies on the C. crescentus proteome.

The critical protein interactions and structures that elicit growth deregulation in cancer and viral replication

PubMed Central

Ou, Horng D.; May, Andrew P.

2010-01-01

One of the greatest challenges in biomedicine is to define the critical targets and network interactions that are subverted to elicit growth deregulation in human cells. Understanding and developing rational treatments for cancer requires a definition of the key molecular targets and how they interact to elicit the complex growth deregulation phenotype. Viral proteins provide discerning and powerful probes to understand both how cells work and how they can be manipulated using a minimal number of components. The small DNA viruses have evolved to target inherent weaknesses in cellular protein interaction networks to hijack the cellular DNA and protein replication machinery. In the battle to escape the inevitability of senescence and programmed cell death, cancers have converged on similar mechanisms, through the acquisition and selection of somatic mutations that drive unchecked cellular replication in tumors. Understanding the dynamic mechanisms through which a minimal number of viral proteins promote host cells to undergo unscheduled and pathological replication is a powerful strategy to identify critical targets that are also disrupted in cancer. Viruses can therefore be used as tools to probe the system-wide protein-protein interactions and structures that drive growth deregulation in human cells. Ultimately this can provide a path for developing system context-dependent therapeutics. This review will describe ongoing experimental approaches using viruses to study pathways deregulated in cancer, with a particular focus on viral cellular protein-protein interactions and structures. PMID:21061422

The adenovirus E4-ORF3 protein functions as a SUMO E3 ligase for TIF-1γ sumoylation and poly-SUMO chain elongation.

PubMed

Sohn, Sook-Young; Hearing, Patrick

2016-06-14

The adenovirus (Ad) early region 4 (E4)-ORF3 protein regulates diverse cellular processes to optimize the host environment for the establishment of Ad replication. E4-ORF3 self-assembles into multimers to form a nuclear scaffold in infected cells and creates distinct binding interfaces for different cellular target proteins. Previous studies have shown that the Ad5 E4-ORF3 protein induces sumoylation of multiple cellular proteins and subsequent proteasomal degradation of some of them, but the detailed mechanism of E4-ORF3 function remained unknown. Here, we investigate the role of E4-ORF3 in the sumoylation process by using transcription intermediary factor (TIF)-1γ as a substrate. Remarkably, we discovered that purified E4-ORF3 protein stimulates TIF-1γ sumoylation in vitro, demonstrating that E4-ORF3 acts as a small ubiquitin-like modifier (SUMO) E3 ligase. Furthermore, E4-ORF3 significantly increases poly-SUMO3 chain formation in vitro in the absence of substrate, showing that E4-ORF3 has SUMO E4 elongase activity. An E4-ORF3 mutant, which is defective in protein multimerization, exhibited severely decreased activity, demonstrating that E4-ORF3 self-assembly is required for these activities. Using a SUMO3 mutant, K11R, we found that E4-ORF3 facilitates the initial acceptor SUMO3 conjugation to TIF-1γ as well as poly-SUMO chain elongation. The E4-ORF3 protein displays no SUMO-targeted ubiquitin ligase activity in our assay system. These studies reveal the mechanism by which E4-ORF3 targets specific cellular proteins for sumoylation and proteasomal degradation and provide significant insight into how a small viral protein can play a role as a SUMO E3 ligase and E4-like SUMO elongase to impact a variety of cellular responses.

Proteomic analysis of protein interactions between Eimeria maxima sporozoites and chicken jejunal epithelial cells by shotgun LC-MS/MS.

PubMed

Huang, Jingwei; Liu, Tingqi; Li, Ke; Song, Xiaokai; Yan, Ruofeng; Xu, Lixin; Li, Xiangrui

2018-04-04

Eimeria maxima initiates infection by invading the jejunal epithelial cells of chicken. However, the proteins involved in invasion remain unknown. The research of the molecules that participate in the interactions between E. maxima sporozoites and host target cells will fill a gap in our understanding of the invasion system of this parasitic pathogen. In the present study, chicken jejunal epithelial cells were isolated and cultured in vitro. Western blot was employed to analyze the soluble proteins of E. maxima sporozoites that bound to chicken jejunal epithelial cells. Co-immunoprecipitation (co-IP) assay was used to separate the E. maxima proteins that bound to chicken jejunal epithelial cells. Shotgun LC-MS/MS technique was used for proteomics identification and Gene Ontology was employed for the bioinformatics analysis. The results of Western blot analysis showed that four proteins bands from jejunal epithelial cells co-cultured with soluble proteins of E. maxima sporozoites were recognized by the positive sera, with molecular weights of 70, 90, 95 and 130 kDa. The co-IP dilutions were analyzed by shotgun LC-MS/MS. A total of 204 proteins were identified in the E. maxima protein database using the MASCOT search engine. Thirty-five proteins including microneme protein 3 and 7 had more than two unique peptide counts and were annotated using Gene Ontology for molecular function, biological process and cellular localization. The results revealed that of the 35 annotated peptides, 22 (62.86%) were associated with binding activity and 15 (42.86%) were involved in catalytic activity. Our findings provide an insight into the interaction between E. maxima and the corresponding host cells and it is important for the understanding of molecular mechanisms underlying E. maxima invasion.

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

PubMed

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

2007-05-01

Currently, the understanding of the relationships between function, amino acid sequence, and protein structure continues to represent one of the major challenges of the modern protein science. As many as 50% of eukaryotic proteins are likely to contain functionally important long disordered regions. Many proteins are wholly disordered but still possess numerous biologically important functions. However, the number of experimentally confirmed disordered proteins with known biological functions is substantially smaller than their actual number in nature. Therefore, there is a crucial need for novel bionformatics approaches that allow projection of the current knowledge from a few experimentally verified examples to much larger groups of known and potential proteins. The elaboration of a bioinformatics tool for the analysis of functional diversity of intrinsically disordered proteins and application of this data mining tool to >200 000 proteins from the Swiss-Prot database, each annotated with at least one of the 875 functional keywords, was described in the first paper of this series (Xie, H.; Vucetic, S.; Iakoucheva, L. M.; Oldfield, C. J.; Dunker, A. K.; Obradovic, Z.; Uversky, V.N. Functional anthology of intrinsic disorder. 1. Biological processes and functions of proteins with long disordered regions. J. Proteome Res. 2007, 5, 1882-1898). Using this tool, we have found that out of the 710 Swiss-Prot functional keywords associated with at least 20 proteins, 262 were strongly positively correlated with long intrinsically disordered regions, and 302 were strongly negatively correlated. Illustrative examples of functional disorder or order were found for the vast majority of keywords showing strongest positive or negative correlation with intrinsic disorder, respectively. Some 80 Swiss-Prot keywords associated with disorder- and order-driven biological processes and protein functions were described in the first paper (see above). The second paper of the series was devoted to the presentation of 87 Swiss-Prot keywords attributed to the cellular components, domains, technical terms, developmental processes, and coding sequence diversities possessing strong positive and negative correlation with long disordered regions (Vucetic, S.; Xie, H.; Iakoucheva, L. M.; Oldfield, C. J.; Dunker, A. K.; Obradovic, Z.; Uversky, V. N. Functional anthology of intrinsic disorder. 2. Cellular components, domains, technical terms, developmental processes, and coding sequence diversities correlated with long disordered regions. J. Proteome Res. 2007, 5, 1899-1916). Protein structure and functionality can be modulated by various post-translational modifications or/and as a result of binding of specific ligands. Numerous human diseases are associated with protein misfolding/misassembly/misfunctioning. This work concludes the series of papers dedicated to the functional anthology of intrinsic disorder and describes approximately 80 Swiss-Prot functional keywords that are related to ligands, post-translational modifications, and diseases possessing strong positive or negative correlation with the predicted long disordered regions in proteins.

The coming of age of chaperone-mediated autophagy.

PubMed

Kaushik, Susmita; Cuervo, Ana Maria

2018-06-01

Chaperone-mediated autophagy (CMA) was the first studied process that indicated that degradation of intracellular components by the lysosome can be selective - a concept that is now well accepted for other forms of autophagy. Lysosomes can degrade cellular cytosol in a nonspecific manner but can also discriminate what to target for degradation with the involvement of a degradation tag, a chaperone and a sophisticated mechanism to make the selected proteins cross the lysosomal membrane through a dedicated translocation complex. Recent studies modulating CMA activity in vivo using transgenic mouse models have demonstrated that selectivity confers on CMA the ability to participate in the regulation of multiple cellular functions. Timely degradation of specific cellular proteins by CMA modulates, for example, glucose and lipid metabolism, DNA repair, cellular reprograming and the cellular response to stress. These findings expand the physiological relevance of CMA beyond its originally identified role in protein quality control and reveal that CMA failure with age may aggravate diseases, such as ageing-associated neurodegeneration and cancer.

Cellular Restriction Factors of Feline Immunodeficiency Virus

PubMed Central

Zielonka, Jörg; Münk, Carsten

2011-01-01

Lentiviruses are known for their narrow cell- and species-tropisms, which are determined by cellular proteins whose absence or presence either support viral replication (dependency factors, cofactors) or inhibit viral replication (restriction factors). Similar to Human immunodeficiency virus type 1 (HIV-1), the cat lentivirus Feline immunodeficiency virus (FIV) is sensitive to recently discovered cellular restriction factors from non-host species that are able to stop viruses from replicating. Of particular importance are the cellular proteins APOBEC3, TRIM5α and tetherin/BST-2. In general, lentiviruses counteract or escape their species’ own variant of the restriction factor, but are targeted by the orthologous proteins of distantly related species. Most of the knowledge regarding lentiviral restriction factors has been obtained in the HIV-1 system; however, much less is known about their effects on other lentiviruses. We describe here the molecular mechanisms that explain how FIV maintains its replication in feline cells, but is largely prevented from cross-species infections by cellular restriction factors. PMID:22069525

Discrimination of Self and Non-Self Ribonucleic Acids

PubMed Central

Gebhardt, Anna; Laudenbach, Beatrice T.

2017-01-01

Most virus infections are controlled through the innate and adaptive immune system. A surprisingly limited number of so-called pattern recognition receptors (PRRs) have the ability to sense a large variety of virus infections. The reason for the broad activity of PRRs lies in the ability to recognize viral nucleic acids. These nucleic acids lack signatures that are present in cytoplasmic cellular nucleic acids and thereby marking them as pathogen-derived. Accumulating evidence suggests that these signatures, which are predominantly sensed by a class of PRRs called retinoic acid-inducible gene I (RIG-I)-like receptors and other proteins, are not unique to viruses but rather resemble immature forms of cellular ribonucleic acids generated by cellular polymerases. RIG-I-like receptors, and other cellular antiviral proteins, may therefore have mainly evolved to sense nonprocessed nucleic acids typically generated by primitive organisms and pathogens. This capability has not only implications on induction of antiviral immunity but also on the function of cellular proteins to handle self-derived RNA with stimulatory potential. PMID:28475460

KnotProt: a database of proteins with knots and slipknots

PubMed Central

Jamroz, Michal; Niemyska, Wanda; Rawdon, Eric J.; Stasiak, Andrzej; Millett, Kenneth C.; Sułkowski, Piotr; Sulkowska, Joanna I.

2015-01-01

The protein topology database KnotProt, http://knotprot.cent.uw.edu.pl/, collects information about protein structures with open polypeptide chains forming knots or slipknots. The knotting complexity of the cataloged proteins is presented in the form of a matrix diagram that shows users the knot type of the entire polypeptide chain and of each of its subchains. The pattern visible in the matrix gives the knotting fingerprint of a given protein and permits users to determine, for example, the minimal length of the knotted regions (knot's core size) or the depth of a knot, i.e. how many amino acids can be removed from either end of the cataloged protein structure before converting it from a knot to a different type of knot. In addition, the database presents extensive information about the biological functions, families and fold types of proteins with non-trivial knotting. As an additional feature, the KnotProt database enables users to submit protein or polymer chains and generate their knotting fingerprints. PMID:25361973

Gene Discovery through Genomic Sequencing of Brucella abortus

PubMed Central

Sánchez, Daniel O.; Zandomeni, Ruben O.; Cravero, Silvio; Verdún, Ramiro E.; Pierrou, Ester; Faccio, Paula; Diaz, Gabriela; Lanzavecchia, Silvia; Agüero, Fernán; Frasch, Alberto C. C.; Andersson, Siv G. E.; Rossetti, Osvaldo L.; Grau, Oscar; Ugalde, Rodolfo A.

2001-01-01

Brucella abortus is the etiological agent of brucellosis, a disease that affects bovines and human. We generated DNA random sequences from the genome of B. abortus strain 2308 in order to characterize molecular targets that might be useful for developing immunological or chemotherapeutic strategies against this pathogen. The partial sequencing of 1,899 clones allowed the identification of 1,199 genomic sequence surveys (GSSs) with high homology (BLAST expect value < 10−5) to sequences deposited in the GenBank databases. Among them, 925 represent putative novel genes for the Brucella genus. Out of 925 nonredundant GSSs, 470 were classified in 15 categories based on cellular function. Seven hundred GSSs showed no significant database matches and remain available for further studies in order to identify their function. A high number of GSSs with homology to Agrobacterium tumefaciens and Rhizobium meliloti proteins were observed, thus confirming their close phylogenetic relationship. Among them, several GSSs showed high similarity with genes related to nodule nitrogen fixation, synthesis of nod factors, nodulation protein symbiotic plasmid, and nodule bacteroid differentiation. We have also identified several B. abortus homologs of virulence and pathogenesis genes from other pathogens, including a homolog to both the Shda gene from Salmonella enterica serovar Typhimurium and the AidA-1 gene from Escherichia coli. Other GSSs displayed significant homologies to genes encoding components of the type III and type IV secretion machineries, suggesting that Brucella might also have an active type III secretion machinery. PMID:11159979

Integrated Controlling System and Unified Database for High Throughput Protein Crystallography Experiments

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gaponov, Yu.A.; Igarashi, N.; Hiraki, M.

2004-05-12

An integrated controlling system and a unified database for high throughput protein crystallography experiments have been developed. Main features of protein crystallography experiments (purification, crystallization, crystal harvesting, data collection, data processing) were integrated into the software under development. All information necessary to perform protein crystallography experiments is stored (except raw X-ray data that are stored in a central data server) in a MySQL relational database. The database contains four mutually linked hierarchical trees describing protein crystals, data collection of protein crystal and experimental data processing. A database editor was designed and developed. The editor supports basic database functions to view,more » create, modify and delete user records in the database. Two search engines were realized: direct search of necessary information in the database and object oriented search. The system is based on TCP/IP secure UNIX sockets with four predefined sending and receiving behaviors, which support communications between all connected servers and clients with remote control functions (creating and modifying data for experimental conditions, data acquisition, viewing experimental data, and performing data processing). Two secure login schemes were designed and developed: a direct method (using the developed Linux clients with secure connection) and an indirect method (using the secure SSL connection using secure X11 support from any operating system with X-terminal and SSH support). A part of the system has been implemented on a new MAD beam line, NW12, at the Photon Factory Advanced Ring for general user experiments.« less

Introducing the CPL/MUW proteome database: interpretation of human liver and liver cancer proteome profiles by referring to isolated primary cells.

PubMed

Wimmer, Helge; Gundacker, Nina C; Griss, Johannes; Haudek, Verena J; Stättner, Stefan; Mohr, Thomas; Zwickl, Hannes; Paulitschke, Verena; Baron, David M; Trittner, Wolfgang; Kubicek, Markus; Bayer, Editha; Slany, Astrid; Gerner, Christopher

2009-06-01

Interpretation of proteome data with a focus on biomarker discovery largely relies on comparative proteome analyses. Here, we introduce a database-assisted interpretation strategy based on proteome profiles of primary cells. Both 2-D-PAGE and shotgun proteomics are applied. We obtain high data concordance with these two different techniques. When applying mass analysis of tryptic spot digests from 2-D gels of cytoplasmic fractions, we typically identify several hundred proteins. Using the same protein fractions, we usually identify more than thousand proteins by shotgun proteomics. The data consistency obtained when comparing these independent data sets exceeds 99% of the proteins identified in the 2-D gels. Many characteristic differences in protein expression of different cells can thus be independently confirmed. Our self-designed SQL database (CPL/MUW - database of the Clinical Proteomics Laboratories at the Medical University of Vienna accessible via www.meduniwien.ac.at/proteomics/database) facilitates (i) quality management of protein identification data, which are based on MS, (ii) the detection of cell type-specific proteins and (iii) of molecular signatures of specific functional cell states. Here, we demonstrate, how the interpretation of proteome profiles obtained from human liver tissue and hepatocellular carcinoma tissue is assisted by the Clinical Proteomics Laboratories at the Medical University of Vienna-database. Therefore, we suggest that the use of reference experiments supported by a tailored database may substantially facilitate data interpretation of proteome profiling experiments.

Comparative Analysis of Cellular Immune Responses in Treated Leishmania Patients and Hamsters against Recombinant Th1 Stimulatory Proteins of Leishmania donovani

PubMed Central

Joshi, Sumit; Yadav, Narendra K.; Rawat, Keerti; Tripathi, Chandra Dev P.; Jaiswal, Anil K.; Khare, Prashant; Tandon, Rati; Baharia, Rajendra K.; Das, Sanchita; Gupta, Reema; Kushawaha, Pramod K.; Sundar, Shyam; Sahasrabuddhe, Amogh A.; Dube, Anuradha

2016-01-01

Our prior studies demonstrated that cellular response of T helper 1 (Th1) type was generated by a soluble antigenic fraction (ranging from 89.9 to 97.1 kDa) of Leishmania donovani promastigote, in treated Leishmania patients as well as hamsters and showed significant prophylactic potential against experimental visceral leishmaniasis (VL). Eighteen Th1 stimulatory proteins were identified through proteomic analysis of this subfraction, out of which 15 were developed as recombinant proteins. In the present work, we have evaluated these 15 recombinant proteins simultaneously for their comparative cellular responses in treated Leishmania patients and hamsters. Six proteins viz. elongation factor-2, enolase, aldolase, triose phosphate isomerase, protein disulfide isomerase, and p45 emerged as most immunogenic as they produced a significant lymphoproliferative response, nitric oxide generation and Th1 cytokine response in PBMCs and lymphocytes of treated Leishmania patients and hamsters respectively. The results suggested that these proteins may be exploited for developing a successful poly-protein and/or poly-epitope vaccine against VL. PMID:27047452

Responses of Plant Proteins to Heavy Metal Stress—A Review

PubMed Central

Hasan, Md. Kamrul; Cheng, Yuan; Kanwar, Mukesh K.; Chu, Xian-Yao; Ahammed, Golam J.; Qi, Zhen-Yu

2017-01-01

Plants respond to environmental pollutants such as heavy metal(s) by triggering the expression of genes that encode proteins involved in stress response. Toxic metal ions profoundly affect the cellular protein homeostasis by interfering with the folding process and aggregation of nascent or non-native proteins leading to decreased cell viability. However, plants possess a range of ubiquitous cellular surveillance systems that enable them to efficiently detoxify heavy metals toward enhanced tolerance to metal stress. As proteins constitute the major workhorses of living cells, the chelation of metal ions in cytosol with phytochelatins and metallothioneins followed by compartmentalization of metals in the vacuoles as well as the repair of stress-damaged proteins or removal and degradation of proteins that fail to achieve their native conformations are critical for plant tolerance to heavy metal stress. In this review, we provide a broad overview of recent advances in cellular protein research with regards to heavy metal tolerance in plants. We also discuss how plants maintain functional and healthy proteomes for survival under such capricious surroundings. PMID:28928754

Protein Corona in Response to Flow: Effect on Protein Concentration and Structure.

PubMed

Jayaram, Dhanya T; Pustulka, Samantha M; Mannino, Robert G; Lam, Wilbur A; Payne, Christine K

2018-04-09

Nanoparticles used in cellular applications encounter free serum proteins that adsorb onto the surface of the nanoparticle, forming a protein corona. This protein layer controls the interaction of nanoparticles with cells. For nanomedicine applications, it is important to consider how intravenous injection and the subsequent shear flow will affect the protein corona. Our goal was to determine if shear flow changed the composition of the protein corona and if these changes affected cellular binding. Colorimetric assays of protein concentration and gel electrophoresis demonstrate that polystyrene nanoparticles subjected to flow have a greater concentration of serum proteins adsorbed on the surface, especially plasminogen. Plasminogen, in the absence of nanoparticles, undergoes changes in structure in response to flow, characterized by fluorescence and circular dichroism spectroscopy. The protein-nanoparticle complexes formed from fetal bovine serum after flow had decreased cellular binding, as measured with flow cytometry. In addition to the relevance for nanomedicine, these results also highlight the technical challenges of protein corona studies. The composition of the protein corona was highly dependent on the initial mixing step: rocking, vortexing, or flow. Overall, these results reaffirm the importance of the protein corona in nanoparticle-cell interactions and point toward the challenges of predicting corona composition based on nanoparticle properties. Copyright © 2018 Biophysical Society. Published by Elsevier Inc. All rights reserved.

Transcriptome of interstitial cells of Cajal reveals unique and selective gene signatures

PubMed Central

Park, Paul J.; Fuchs, Robert; Wei, Lai; Jorgensen, Brian G.; Redelman, Doug; Ward, Sean M.; Sanders, Kenton M.

2017-01-01

Transcriptome-scale data can reveal essential clues into understanding the underlying molecular mechanisms behind specific cellular functions and biological processes. Transcriptomics is a continually growing field of research utilized in biomarker discovery. The transcriptomic profile of interstitial cells of Cajal (ICC), which serve as slow-wave electrical pacemakers for gastrointestinal (GI) smooth muscle, has yet to be uncovered. Using copGFP-labeled ICC mice and flow cytometry, we isolated ICC populations from the murine small intestine and colon and obtained their transcriptomes. In analyzing the transcriptome, we identified a unique set of ICC-restricted markers including transcription factors, epigenetic enzymes/regulators, growth factors, receptors, protein kinases/phosphatases, and ion channels/transporters. This analysis provides new and unique insights into the cellular and biological functions of ICC in GI physiology. Additionally, we constructed an interactive ICC genome browser (http://med.unr.edu/physio/transcriptome) based on the UCSC genome database. To our knowledge, this is the first online resource that provides a comprehensive library of all known genetic transcripts expressed in primary ICC. Our genome browser offers a new perspective into the alternative expression of genes in ICC and provides a valuable reference for future functional studies. PMID:28426719

Extraction, integration and analysis of alternative splicing and protein structure distributed information

PubMed Central

D'Antonio, Matteo; Masseroli, Marco

2009-01-01

Background Alternative splicing has been demonstrated to affect most of human genes; different isoforms from the same gene encode for proteins which differ for a limited number of residues, thus yielding similar structures. This suggests possible correlations between alternative splicing and protein structure. In order to support the investigation of such relationships, we have developed the Alternative Splicing and Protein Structure Scrutinizer (PASS), a Web application to automatically extract, integrate and analyze human alternative splicing and protein structure data sparsely available in the Alternative Splicing Database, Ensembl databank and Protein Data Bank. Primary data from these databases have been integrated and analyzed using the Protein Identifier Cross-Reference, BLAST, CLUSTALW and FeatureMap3D software tools. Results A database has been developed to store the considered primary data and the results from their analysis; a system of Perl scripts has been implemented to automatically create and update the database and analyze the integrated data; a Web interface has been implemented to make the analyses easily accessible; a database has been created to manage user accesses to the PASS Web application and store user's data and searches. Conclusion PASS automatically integrates data from the Alternative Splicing Database with protein structure data from the Protein Data Bank. Additionally, it comprehensively analyzes the integrated data with publicly available well-known bioinformatics tools in order to generate structural information of isoform pairs. Further analysis of such valuable information might reveal interesting relationships between alternative splicing and protein structure differences, which may be significantly associated with different functions. PMID:19828075

Structural classification of proteins using texture descriptors extracted from the cellular automata image.

PubMed

Kavianpour, Hamidreza; Vasighi, Mahdi

2017-02-01

Nowadays, having knowledge about cellular attributes of proteins has an important role in pharmacy, medical science and molecular biology. These attributes are closely correlated with the function and three-dimensional structure of proteins. Knowledge of protein structural class is used by various methods for better understanding the protein functionality and folding patterns. Computational methods and intelligence systems can have an important role in performing structural classification of proteins. Most of protein sequences are saved in databanks as characters and strings and a numerical representation is essential for applying machine learning methods. In this work, a binary representation of protein sequences is introduced based on reduced amino acids alphabets according to surrounding hydrophobicity index. Many important features which are hidden in these long binary sequences can be clearly displayed through their cellular automata images. The extracted features from these images are used to build a classification model by support vector machine. Comparing to previous studies on the several benchmark datasets, the promising classification rates obtained by tenfold cross-validation imply that the current approach can help in revealing some inherent features deeply hidden in protein sequences and improve the quality of predicting protein structural class.

RNA-Binding Protein FXR1 Regulates p21 and TERC RNA to Bypass p53-Mediated Cellular Senescence in OSCC

PubMed Central

Majumder, Mrinmoyee; House, Reniqua; Palanisamy, Nallasivam; Qie, Shuo; Day, Terrence A.; Neskey, David; Diehl, J. Alan

2016-01-01

RNA-binding proteins (RBP) regulate numerous aspects of co- and post-transcriptional gene expression in cancer cells. Here, we demonstrate that RBP, fragile X-related protein 1 (FXR1), plays an essential role in cellular senescence by utilizing mRNA turnover pathway. We report that overexpressed FXR1 in head and neck squamous cell carcinoma targets (G-quadruplex (G4) RNA structure within) both mRNA encoding p21 (Cyclin-Dependent Kinase Inhibitor 1A (CDKN1A, Cip1) and the non-coding RNA Telomerase RNA Component (TERC), and regulates their turnover to avoid senescence. Silencing of FXR1 in cancer cells triggers the activation of Cyclin-Dependent Kinase Inhibitors, p53, increases DNA damage, and ultimately, cellular senescence. Overexpressed FXR1 binds and destabilizes p21 mRNA, subsequently reduces p21 protein expression in oral cancer cells. In addition, FXR1 also binds and stabilizes TERC RNA and suppresses the cellular senescence possibly through telomerase activity. Finally, we report that FXR1-regulated senescence is irreversible and FXR1-depleted cells fail to form colonies to re-enter cellular proliferation. Collectively, FXR1 displays a novel mechanism of controlling the expression of p21 through p53-dependent manner to bypass cellular senescence in oral cancer cells. PMID:27606879

GRBase, a new gene regulation data base available by anonymous ftp.

PubMed Central

Collier, B; Danielsen, M

1994-01-01

The Gene Regulation Database (GRBase) is a compendium of information on the structure and function of proteins involved in the control of gene expression in eukaryotes. These proteins include transcription factors, proteins involved in signal transduction, and receptors. The database can be obtained by FTP in Filemaker Pro, text, and postscript formats. The database will be expanded in the coming year to include reviews on families of proteins involved in gene regulation and to allow online searching. PMID:7937071

Exosomes and Homeostatic Synaptic Plasticity Are Linked to Each other and to Huntington's, Parkinson's, and Other Neurodegenerative Diseases by Database-Enabled Analyses of Comprehensively Curated Datasets

PubMed Central

Wang, James K. T.; Langfelder, Peter; Horvath, Steve; Palazzolo, Michael J.

2017-01-01

Huntington's disease (HD) is a progressive and autosomal dominant neurodegeneration caused by CAG expansion in the huntingtin gene (HTT), but the pathophysiological mechanism of mutant HTT (mHTT) remains unclear. To study HD using systems biological methodologies on all published data, we undertook the first comprehensive curation of two key PubMed HD datasets: perturbation genes that impact mHTT-driven endpoints and therefore are putatively linked causally to pathogenic mechanisms, and the protein interactome of HTT that reflects its biology. We perused PubMed articles containing co-citation of gene IDs and MeSH terms of interest to generate mechanistic gene sets for iterative enrichment analyses and rank ordering. The HD Perturbation database of 1,218 genes highly overlaps the HTT Interactome of 1,619 genes, suggesting links between normal HTT biology and mHTT pathology. These two HD datasets are enriched for protein networks of key genes underlying two mechanisms not previously implicated in HD nor in each other: exosome synaptic functions and homeostatic synaptic plasticity. Moreover, proteins, possibly including HTT, and miRNA detected in exosomes from a wide variety of sources also highly overlap the HD datasets, suggesting both mechanistic and biomarker links. Finally, the HTT Interactome highly intersects protein networks of pathogenic genes underlying Parkinson's, Alzheimer's and eight non-HD polyglutamine diseases, ALS, and spinal muscular atrophy. These protein networks in turn highly overlap the exosome and homeostatic synaptic plasticity gene sets. Thus, we hypothesize that HTT and other neurodegeneration pathogenic genes form a large interlocking protein network involved in exosome and homeostatic synaptic functions, particularly where the two mechanisms intersect. Mutant pathogenic proteins cause dysfunctions at distinct points in this network, each altering the two mechanisms in specific fashion that contributes to distinct disease pathologies, depending on the gene mutation and the cellular and biological context. This protein network is rich with drug targets, and exosomes may provide disease biomarkers, thus enabling drug discovery. All the curated datasets are made available for other investigators. Elucidating the roles of pathogenic neurodegeneration genes in exosome and homeostatic synaptic functions may provide a unifying framework for the age-dependent, progressive and tissue selective nature of multiple neurodegenerative diseases. PMID:28611571

Exosomes and Homeostatic Synaptic Plasticity Are Linked to Each other and to Huntington's, Parkinson's, and Other Neurodegenerative Diseases by Database-Enabled Analyses of Comprehensively Curated Datasets.

PubMed

Wang, James K T; Langfelder, Peter; Horvath, Steve; Palazzolo, Michael J

2017-01-01

Huntington's disease (HD) is a progressive and autosomal dominant neurodegeneration caused by CAG expansion in the huntingtin gene ( HTT ), but the pathophysiological mechanism of mutant HTT (mHTT) remains unclear. To study HD using systems biological methodologies on all published data, we undertook the first comprehensive curation of two key PubMed HD datasets: perturbation genes that impact mHTT-driven endpoints and therefore are putatively linked causally to pathogenic mechanisms, and the protein interactome of HTT that reflects its biology. We perused PubMed articles containing co-citation of gene IDs and MeSH terms of interest to generate mechanistic gene sets for iterative enrichment analyses and rank ordering. The HD Perturbation database of 1,218 genes highly overlaps the HTT Interactome of 1,619 genes, suggesting links between normal HTT biology and mHTT pathology. These two HD datasets are enriched for protein networks of key genes underlying two mechanisms not previously implicated in HD nor in each other: exosome synaptic functions and homeostatic synaptic plasticity. Moreover, proteins, possibly including HTT, and miRNA detected in exosomes from a wide variety of sources also highly overlap the HD datasets, suggesting both mechanistic and biomarker links. Finally, the HTT Interactome highly intersects protein networks of pathogenic genes underlying Parkinson's, Alzheimer's and eight non-HD polyglutamine diseases, ALS, and spinal muscular atrophy. These protein networks in turn highly overlap the exosome and homeostatic synaptic plasticity gene sets. Thus, we hypothesize that HTT and other neurodegeneration pathogenic genes form a large interlocking protein network involved in exosome and homeostatic synaptic functions, particularly where the two mechanisms intersect. Mutant pathogenic proteins cause dysfunctions at distinct points in this network, each altering the two mechanisms in specific fashion that contributes to distinct disease pathologies, depending on the gene mutation and the cellular and biological context. This protein network is rich with drug targets, and exosomes may provide disease biomarkers, thus enabling drug discovery. All the curated datasets are made available for other investigators. Elucidating the roles of pathogenic neurodegeneration genes in exosome and homeostatic synaptic functions may provide a unifying framework for the age-dependent, progressive and tissue selective nature of multiple neurodegenerative diseases.

Steap4 Plays a Critical Role in Osteoclastogenesis in Vitro by Regulating Cellular Iron/Reactive Oxygen Species (ROS) Levels and cAMP Response Element-binding Protein (CREB) Activation*

PubMed Central

Zhou, Jian; Ye, Shiqiao; Fujiwara, Toshifumi; Manolagas, Stavros C.; Zhao, Haibo

2013-01-01

Iron is essential for osteoclast differentiation, and iron overload in a variety of hematologic diseases is associated with excessive bone resorption. Iron uptake by osteoclast precursors via the transferrin cycle increases mitochondrial biogenesis, reactive oxygen species production, and activation of cAMP response element-binding protein, a critical transcription factor downstream of receptor activator of NF-κB-ligand-induced calcium signaling. These changes are required for the differentiation of osteoclast precursors to mature bone-resorbing osteoclasts. However, the molecular mechanisms regulating cellular iron metabolism in osteoclasts remain largely unknown. In this report, we provide evidence that Steap4, a member of the six-transmembrane epithelial antigen of prostate (Steap) family proteins, is an endosomal ferrireductase with a critical role in cellular iron utilization in osteoclasts. Specifically, we show that Steap4 is the only Steap family protein that is up-regulated during osteoclast differentiation. Knocking down Steap4 expression in vitro by lentivirus-mediated short hairpin RNAs inhibits osteoclast formation and decreases cellular ferrous iron, reactive oxygen species, and the activation of cAMP response element-binding protein. These results demonstrate that Steap4 is a critical enzyme for cellular iron uptake and utilization in osteoclasts and, thus, indispensable for osteoclast development and function. PMID:23990467

Bioinformatics: Cheap and robust method to explore biomaterial from Indonesia biodiversity

NASA Astrophysics Data System (ADS)

Widodo

2015-02-01

Indonesia has a huge amount of biodiversity, which may contain many biomaterials for pharmaceutical application. These resources potency should be explored to discover new drugs for human wealth. However, the bioactive screening using conventional methods is very expensive and time-consuming. Therefore, we developed a methodology for screening the potential of natural resources based on bioinformatics. The method is developed based on the fact that organisms in the same taxon will have similar genes, metabolism and secondary metabolites product. Then we employ bioinformatics to explore the potency of biomaterial from Indonesia biodiversity by comparing species with the well-known taxon containing the active compound through published paper or chemical database. Then we analyze drug-likeness, bioactivity and the target proteins of the active compound based on their molecular structure. The target protein was examined their interaction with other proteins in the cell to determine action mechanism of the active compounds in the cellular level, as well as to predict its side effects and toxicity. By using this method, we succeeded to screen anti-cancer, immunomodulators and anti-inflammation from Indonesia biodiversity. For example, we found anticancer from marine invertebrate by employing the method. The anti-cancer was explore based on the isolated compounds of marine invertebrate from published article and database, and then identified the protein target, followed by molecular pathway analysis. The data suggested that the active compound of the invertebrate able to kill cancer cell. Further, we collect and extract the active compound from the invertebrate, and then examined the activity on cancer cell (MCF7). The MTT result showed that the methanol extract of marine invertebrate was highly potent in killing MCF7 cells. Therefore, we concluded that bioinformatics is cheap and robust way to explore bioactive from Indonesia biodiversity for source of drug and another pharmaceutical material.

Integrated cellular network of transcription regulations and protein-protein interactions

PubMed Central

2010-01-01

Background With the accumulation of increasing omics data, a key goal of systems biology is to construct networks at different cellular levels to investigate cellular machinery of the cell. However, there is currently no satisfactory method to construct an integrated cellular network that combines the gene regulatory network and the signaling regulatory pathway. Results In this study, we integrated different kinds of omics data and developed a systematic method to construct the integrated cellular network based on coupling dynamic models and statistical assessments. The proposed method was applied to S. cerevisiae stress responses, elucidating the stress response mechanism of the yeast. From the resulting integrated cellular network under hyperosmotic stress, the highly connected hubs which are functionally relevant to the stress response were identified. Beyond hyperosmotic stress, the integrated network under heat shock and oxidative stress were also constructed and the crosstalks of these networks were analyzed, specifying the significance of some transcription factors to serve as the decision-making devices at the center of the bow-tie structure and the crucial role for rapid adaptation scheme to respond to stress. In addition, the predictive power of the proposed method was also demonstrated. Conclusions We successfully construct the integrated cellular network which is validated by literature evidences. The integration of transcription regulations and protein-protein interactions gives more insight into the actual biological network and is more predictive than those without integration. The method is shown to be powerful and flexible and can be used under different conditions and for different species. The coupling dynamic models of the whole integrated cellular network are very useful for theoretical analyses and for further experiments in the fields of network biology and synthetic biology. PMID:20211003

Integrated cellular network of transcription regulations and protein-protein interactions.

PubMed

Wang, Yu-Chao; Chen, Bor-Sen

2010-03-08

With the accumulation of increasing omics data, a key goal of systems biology is to construct networks at different cellular levels to investigate cellular machinery of the cell. However, there is currently no satisfactory method to construct an integrated cellular network that combines the gene regulatory network and the signaling regulatory pathway. In this study, we integrated different kinds of omics data and developed a systematic method to construct the integrated cellular network based on coupling dynamic models and statistical assessments. The proposed method was applied to S. cerevisiae stress responses, elucidating the stress response mechanism of the yeast. From the resulting integrated cellular network under hyperosmotic stress, the highly connected hubs which are functionally relevant to the stress response were identified. Beyond hyperosmotic stress, the integrated network under heat shock and oxidative stress were also constructed and the crosstalks of these networks were analyzed, specifying the significance of some transcription factors to serve as the decision-making devices at the center of the bow-tie structure and the crucial role for rapid adaptation scheme to respond to stress. In addition, the predictive power of the proposed method was also demonstrated. We successfully construct the integrated cellular network which is validated by literature evidences. The integration of transcription regulations and protein-protein interactions gives more insight into the actual biological network and is more predictive than those without integration. The method is shown to be powerful and flexible and can be used under different conditions and for different species. The coupling dynamic models of the whole integrated cellular network are very useful for theoretical analyses and for further experiments in the fields of network biology and synthetic biology.

The Role of the Multifunctional BAG3 Protein in Cellular Protein Quality Control and in Disease

PubMed Central

Stürner, Elisabeth; Behl, Christian

2017-01-01

In neurons, but also in all other cells the complex proteostasis network is monitored and tightly regulated by the cellular protein quality control (PQC) system. Beyond folding of newly synthesized polypeptides and their refolding upon misfolding the PQC also manages the disposal of aberrant proteins either by the ubiquitin-proteasome machinery or by the autophagic-lysosomal system. Aggregated proteins are primarily degraded by a process termed selective macroautophagy (or aggrephagy). One such recently discovered selective macroautophagy pathway is mediated by the multifunctional HSP70 co-chaperone BAG3 (BCL-2-associated athanogene 3). Under acute stress and during cellular aging, BAG3 in concert with the molecular chaperones HSP70 and HSPB8 as well as the ubiquitin receptor p62/SQSTM1 specifically targets aggregation-prone proteins to autophagic degradation. Thereby, BAG3-mediated selective macroautophagy represents a pivotal adaptive safeguarding and emergency system of the PQC which is activated under pathophysiological conditions to ensure cellular proteostasis. Interestingly, BAG3-mediated selective macroautophagy is also involved in the clearance of aggregated proteins associated with age-related neurodegenerative disorders, like Alzheimer’s disease (tau-protein), Huntington’s disease (mutated huntingtin/polyQ proteins), and amyotrophic lateral sclerosis (mutated SOD1). In addition, based on its initial description BAG3 is an anti-apoptotic protein that plays a decisive role in other widespread diseases, including cancer and myopathies. Therefore, in the search for novel therapeutic intervention avenues in neurodegeneration, myopathies and cancer BAG3 is a promising candidate. PMID:28680391

Columba: an integrated database of proteins, structures, and annotations.

PubMed

Trissl, Silke; Rother, Kristian; Müller, Heiko; Steinke, Thomas; Koch, Ina; Preissner, Robert; Frömmel, Cornelius; Leser, Ulf

2005-03-31

Structural and functional research often requires the computation of sets of protein structures based on certain properties of the proteins, such as sequence features, fold classification, or functional annotation. Compiling such sets using current web resources is tedious because the necessary data are spread over many different databases. To facilitate this task, we have created COLUMBA, an integrated database of annotations of protein structures. COLUMBA currently integrates twelve different databases, including PDB, KEGG, Swiss-Prot, CATH, SCOP, the Gene Ontology, and ENZYME. The database can be searched using either keyword search or data source-specific web forms. Users can thus quickly select and download PDB entries that, for instance, participate in a particular pathway, are classified as containing a certain CATH architecture, are annotated as having a certain molecular function in the Gene Ontology, and whose structures have a resolution under a defined threshold. The results of queries are provided in both machine-readable extensible markup language and human-readable format. The structures themselves can be viewed interactively on the web. The COLUMBA database facilitates the creation of protein structure data sets for many structure-based studies. It allows to combine queries on a number of structure-related databases not covered by other projects at present. Thus, information on both many and few protein structures can be used efficiently. The web interface for COLUMBA is available at http://www.columba-db.de.

Pericentrin in cellular function and disease

PubMed Central

Delaval, Benedicte

2010-01-01

Pericentrin is an integral component of the centrosome that serves as a multifunctional scaffold for anchoring numerous proteins and protein complexes. Through these interactions, pericentrin contributes to a diversity of fundamental cellular processes. Recent studies link pericentrin to a growing list of human disorders. Studies on pericentrin at the cellular, molecular, and, more recently, organismal level, provide a platform for generating models to elucidate the etiology of these disorders. Although the complexity of phenotypes associated with pericentrin-mediated disorders is somewhat daunting, insights into the cellular basis of disease are beginning to come into focus. In this review, we focus on human conditions associated with loss or elevation of pericentrin and propose cellular and molecular models that might explain them. PMID:19951897

Coordinating Center: Molecular and Cellular Findings of Screen-Detected Lesions | Division of Cancer Prevention

Cancer.gov

The Molecular and Cellular Characterization of Screen‐Detected Lesions ‐ Coordinating Center and Data Management Group will provide support for the participating studies responding to RFA CA14‐10. The coordinating center supports three main domains: network coordination, statistical support and computational analysis and protocol development and database support. Support for

Learning Cellular Sorting Pathways Using Protein Interactions and Sequence Motifs

PubMed Central

Lin, Tien-Ho; Bar-Joseph, Ziv

2011-01-01

Abstract Proper subcellular localization is critical for proteins to perform their roles in cellular functions. Proteins are transported by different cellular sorting pathways, some of which take a protein through several intermediate locations until reaching its final destination. The pathway a protein is transported through is determined by carrier proteins that bind to specific sequence motifs. In this article, we present a new method that integrates protein interaction and sequence motif data to model how proteins are sorted through these sorting pathways. We use a hidden Markov model (HMM) to represent protein sorting pathways. The model is able to determine intermediate sorting states and to assign carrier proteins and motifs to the sorting pathways. In simulation studies, we show that the method can accurately recover an underlying sorting model. Using data for yeast, we show that our model leads to accurate prediction of subcellular localization. We also show that the pathways learned by our model recover many known sorting pathways and correctly assign proteins to the path they utilize. The learned model identified new pathways and their putative carriers and motifs and these may represent novel protein sorting mechanisms. Supplementary results and software implementation are available from http://murphylab.web.cmu.edu/software/2010_RECOMB_pathways/. PMID:21999284

Deciphering the Acute Cellular Phosphoproteome Response to Irradiation with X-rays, Protons and Carbon Ions*

PubMed Central

Winter, Martin; Dokic, Ivana; Schlegel, Julian; Warnken, Uwe; Debus, Jürgen; Abdollahi, Amir; Schnölzer, Martina

2017-01-01

Radiotherapy is a cornerstone of cancer therapy. The recently established particle therapy with raster-scanning protons and carbon ions landmarks a new era in the field of high-precision cancer medicine. However, molecular mechanisms governing radiation induced intracellular signaling remain elusive. Here, we present the first comprehensive proteomic and phosphoproteomic study applying stable isotope labeling by amino acids in cell culture (SILAC) in combination with high-resolution mass spectrometry to decipher cellular response to irradiation with X-rays, protons and carbon ions. At protein expression level limited alterations were observed 2 h post irradiation of human lung adenocarcinoma cells. In contrast, 181 phosphorylation sites were found to be differentially regulated out of which 151 sites were not hitherto attributed to radiation response as revealed by crosscheck with the PhosphoSitePlus database. Radiation-induced phosphorylation of the p(S/T)Q motif was the prevailing regulation pattern affecting proteins involved in DNA damage response signaling. Because radiation doses were selected to produce same level of cell kill and DNA double-strand breakage for each radiation quality, DNA damage responsive phosphorylation sites were regulated to same extent. However, differential phosphorylation between radiation qualities was observed for 55 phosphorylation sites indicating the existence of distinct signaling circuitries induced by X-ray versus particle (proton/carbon) irradiation beyond the canonical DNA damage response. This unexpected finding was confirmed in targeted spike-in experiments using synthetic isotope labeled phosphopeptides. Herewith, we successfully validated uniform DNA damage response signaling coexisting with altered signaling involved in apoptosis and metabolic processes induced by X-ray and particle based treatments. In summary, the comprehensive insight into the radiation-induced phosphoproteome landscape is instructive for the design of functional studies aiming to decipher cellular signaling processes in response to radiotherapy, space radiation or ionizing radiation per se. Further, our data will have a significant impact on the ongoing debate about patient treatment modalities. PMID:28302921

Deciphering the Acute Cellular Phosphoproteome Response to Irradiation with X-rays, Protons and Carbon Ions.

PubMed

Winter, Martin; Dokic, Ivana; Schlegel, Julian; Warnken, Uwe; Debus, Jürgen; Abdollahi, Amir; Schnölzer, Martina

2017-05-01

Radiotherapy is a cornerstone of cancer therapy. The recently established particle therapy with raster-scanning protons and carbon ions landmarks a new era in the field of high-precision cancer medicine. However, molecular mechanisms governing radiation induced intracellular signaling remain elusive. Here, we present the first comprehensive proteomic and phosphoproteomic study applying stable isotope labeling by amino acids in cell culture (SILAC) in combination with high-resolution mass spectrometry to decipher cellular response to irradiation with X-rays, protons and carbon ions. At protein expression level limited alterations were observed 2 h post irradiation of human lung adenocarcinoma cells. In contrast, 181 phosphorylation sites were found to be differentially regulated out of which 151 sites were not hitherto attributed to radiation response as revealed by crosscheck with the PhosphoSitePlus database.Radiation-induced phosphorylation of the p(S/T)Q motif was the prevailing regulation pattern affecting proteins involved in DNA damage response signaling. Because radiation doses were selected to produce same level of cell kill and DNA double-strand breakage for each radiation quality, DNA damage responsive phosphorylation sites were regulated to same extent. However, differential phosphorylation between radiation qualities was observed for 55 phosphorylation sites indicating the existence of distinct signaling circuitries induced by X-ray versus particle (proton/carbon) irradiation beyond the canonical DNA damage response. This unexpected finding was confirmed in targeted spike-in experiments using synthetic isotope labeled phosphopeptides. Herewith, we successfully validated uniform DNA damage response signaling coexisting with altered signaling involved in apoptosis and metabolic processes induced by X-ray and particle based treatments.In summary, the comprehensive insight into the radiation-induced phosphoproteome landscape is instructive for the design of functional studies aiming to decipher cellular signaling processes in response to radiotherapy, space radiation or ionizing radiation per se Further, our data will have a significant impact on the ongoing debate about patient treatment modalities. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

The Molecular Chaperone TRiC/CCT Binds to the Trp-Asp 40 (WD40) Repeat Protein WDR68 and Promotes Its Folding, Protein Kinase DYRK1A Binding, and Nuclear Accumulation*

PubMed Central

Miyata, Yoshihiko; Shibata, Takeshi; Aoshima, Masato; Tsubata, Takuichi; Nishida, Eisuke

2014-01-01

Trp-Asp (WD) repeat protein 68 (WDR68) is an evolutionarily conserved WD40 repeat protein that binds to several proteins, including dual specificity tyrosine phosphorylation-regulated protein kinase (DYRK1A), MAPK/ERK kinase kinase 1 (MEKK1), and Cullin4-damage-specific DNA-binding protein 1 (CUL4-DDB1). WDR68 affects multiple and diverse physiological functions, such as controlling anthocyanin synthesis in plants, tissue growth in insects, and craniofacial development in vertebrates. However, the biochemical basis and the regulatory mechanism of WDR68 activity remain largely unknown. To better understand the cellular function of WDR68, here we have isolated and identified cellular WDR68 binding partners using a phosphoproteomic approach. More than 200 cellular proteins with wide varieties of biochemical functions were identified as WDR68-binding protein candidates. Eight T-complex protein 1 (TCP1) subunits comprising the molecular chaperone TCP1 ring complex/chaperonin-containing TCP1 (TRiC/CCT) were identified as major WDR68-binding proteins, and phosphorylation sites in both WDR68 and TRiC/CCT were identified. Co-immunoprecipitation experiments confirmed the binding between TRiC/CCT and WDR68. Computer-aided structural analysis suggested that WDR68 forms a seven-bladed β-propeller ring. Experiments with a series of deletion mutants in combination with the structural modeling showed that three of the seven β-propeller blades of WDR68 are essential and sufficient for TRiC/CCT binding. Knockdown of cellular TRiC/CCT by siRNA caused an abnormal WDR68 structure and led to reduction of its DYRK1A-binding activity. Concomitantly, nuclear accumulation of WDR68 was suppressed by the knockdown of TRiC/CCT, and WDR68 formed cellular aggregates when overexpressed in the TRiC/CCT-deficient cells. Altogether, our results demonstrate that the molecular chaperone TRiC/CCT is essential for correct protein folding, DYRK1A binding, and nuclear accumulation of WDR68. PMID:25342745

Cellular repressor of E1A-stimulated genes is a bona fide lysosomal protein which undergoes proteolytic maturation during its biosynthesis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Schaehs, Philipp; Weidinger, Petra; Probst, Olivia C.

2008-10-01

Cellular repressor of E1A-stimulated genes (CREG) has been reported to be a secretory glycoprotein implicated in cellular growth and differentiation. We now show that CREG is predominantly localized within intracellular compartments. Intracellular CREG was found to lack an N-terminal peptide present in the secreted form of the protein. In contrast to normal cells, CREG is largely secreted by fibroblasts missing both mannose 6-phosphate receptors. This is not observed in cells lacking only one of them. Mass spectrometric analysis of recombinant CREG revealed that the protein contains phosphorylated oligosaccharides at either of its two N-glycosylation sites. Cellular CREG was found tomore » cosediment with lysosomal markers upon subcellular fractionation by density-gradient centrifugation. In fibroblasts expressing a CREG-GFP fusion construct, the heterologous protein was detected in compartments containing lysosomal proteins. Immunolocalization of endogenous CREG confirmed that intracellular CREG is localized in lysosomes. Proteolytic processing of intracellular CREG involves the action of lysosomal cysteine proteinases. These results establish that CREG is a lysosomal protein that undergoes proteolytic maturation in the course of its biosynthesis, carries the mannose 6-phosphate recognition marker and depends on the interaction with mannose 6-phosphate receptors for efficient delivery to lysosomes.« less

Elucidation of the Ebola virus VP24 cellular interactome and disruption of virus biology through targeted inhibition of host-cell protein function.

PubMed

García-Dorival, Isabel; Wu, Weining; Dowall, Stuart; Armstrong, Stuart; Touzelet, Olivier; Wastling, Jonathan; Barr, John N; Matthews, David; Carroll, Miles; Hewson, Roger; Hiscox, Julian A

2014-11-07

Viral pathogenesis in the infected cell is a balance between antiviral responses and subversion of host-cell processes. Many viral proteins specifically interact with host-cell proteins to promote virus biology. Understanding these interactions can lead to knowledge gains about infection and provide potential targets for antiviral therapy. One such virus is Ebola, which has profound consequences for human health and causes viral hemorrhagic fever where case fatality rates can approach 90%. The Ebola virus VP24 protein plays a critical role in the evasion of the host immune response and is likely to interact with multiple cellular proteins. To map these interactions and better understand the potential functions of VP24, label-free quantitative proteomics was used to identify cellular proteins that had a high probability of forming the VP24 cellular interactome. Several known interactions were confirmed, thus placing confidence in the technique, but new interactions were also discovered including one with ATP1A1, which is involved in osmoregulation and cell signaling. Disrupting the activity of ATP1A1 in Ebola-virus-infected cells with a small molecule inhibitor resulted in a decrease in progeny virus, thus illustrating how quantitative proteomics can be used to identify potential therapeutic targets.

Skeletal muscle plasticity: cellular and molecular responses to altered physical activity paradigms

NASA Technical Reports Server (NTRS)

Baldwin, Kenneth M.; Haddad, Fadia

2002-01-01

The goal of this article is to examine our current understanding of the chain of events known to be involved in the adaptive process whereby specific genes and their protein products undergo altered expression; specifically, skeletal muscle adaptation in response to altered loading states will be discussed, with a special focus on the regulation of the contractile protein, myosin heavy chain gene expression. This protein, which is both an important structural and regulatory protein comprising the contractile apparatus, can be expressed as different isoforms, thereby having an impact on the functional diversity of the muscle. Because the regulation of the myosin gene family is under the control of a complex set of processes including, but not limited to, activity, hormonal, and metabolic factors, this protein will serve as a cellular "marker" for studies of muscle plasticity in response to various mechanical perturbations in which the quantity and type of myosin isoform, along with other important cellular proteins, are altered in expression.

Hacking the Cell: Network Intrusion and Exploitation by Adenovirus E1A.

PubMed

King, Cason R; Zhang, Ali; Tessier, Tanner M; Gameiro, Steven F; Mymryk, Joe S

2018-05-01

As obligate intracellular parasites, viruses are dependent on their infected hosts for survival. Consequently, viruses are under enormous selective pressure to utilize available cellular components and processes to their own advantage. As most, if not all, cellular activities are regulated at some level via protein interactions, host protein interaction networks are particularly vulnerable to viral exploitation. Indeed, viral proteins frequently target highly connected "hub" proteins to "hack" the cellular network, defining the molecular basis for viral control over the host. This widespread and successful strategy of network intrusion and exploitation has evolved convergently among numerous genetically distinct viruses as a result of the endless evolutionary arms race between pathogens and hosts. Here we examine the means by which a particularly well-connected viral hub protein, human adenovirus E1A, compromises and exploits the vulnerabilities of eukaryotic protein interaction networks. Importantly, these interactions identify critical regulatory hubs in the human proteome and help define the molecular basis of their function. Copyright © 2018 King et al.

Hacking the Cell: Network Intrusion and Exploitation by Adenovirus E1A

PubMed Central

King, Cason R.; Zhang, Ali; Tessier, Tanner M.; Gameiro, Steven F.

2018-01-01

ABSTRACT As obligate intracellular parasites, viruses are dependent on their infected hosts for survival. Consequently, viruses are under enormous selective pressure to utilize available cellular components and processes to their own advantage. As most, if not all, cellular activities are regulated at some level via protein interactions, host protein interaction networks are particularly vulnerable to viral exploitation. Indeed, viral proteins frequently target highly connected “hub” proteins to “hack” the cellular network, defining the molecular basis for viral control over the host. This widespread and successful strategy of network intrusion and exploitation has evolved convergently among numerous genetically distinct viruses as a result of the endless evolutionary arms race between pathogens and hosts. Here we examine the means by which a particularly well-connected viral hub protein, human adenovirus E1A, compromises and exploits the vulnerabilities of eukaryotic protein interaction networks. Importantly, these interactions identify critical regulatory hubs in the human proteome and help define the molecular basis of their function. PMID:29717008

Altered Protein Expression of Streptococcus oralis Cultured at Low pH Revealed by Two-Dimensional Gel Electrophoresis

PubMed Central

Wilkins, Joanna C.; Homer, Karen A.; Beighton, David

2001-01-01

Streptococcus oralis is the predominant aciduric nonmutans streptococcus isolated from the human dentition, but the role of this organism in the initiation and progression of dental caries has yet to be established. To identify proteins that are differentially expressed by S. oralis growing under conditions of low pH, soluble cellular proteins extracted from bacteria grown in batch culture at pH 5.2 or 7.0 were analyzed by two-dimensional (2-D) gel electrophoresis. Thirty-nine proteins had altered expression at low pH; these were excised, digested with trypsin using an in-gel protocol, and further analyzed by peptide mass fingerprinting using matrix-assisted laser desorption ionization mass spectrometry. The resulting fingerprints were compared with the genomic database for Streptococcus pneumoniae, an organism that is phylogenetically closely related to S. oralis, and putative functions for the majority of these proteins were determined on the basis of functional homology. Twenty-eight proteins were up-regulated following growth at pH 5.2; these included enzymes of the glycolytic pathway (glyceraldehyde-3-phosphate dehydrogenase and lactate dehydrogenase), the polypeptide chains comprising ATP synthase, and proteins that are considered to play a role in the general stress response of bacteria, including the 60-kDa chaperone, Hsp33, and superoxide dismutase, and three distinct ABC transporters. These data identify, for the first time, gene products that may be important in the survival and proliferation of nonmutans aciduric S. oralis under conditions of low pH that are likely to be encountered by this organism in vivo. PMID:11472910

sc-PDB: a 3D-database of ligandable binding sites—10 years on

PubMed Central

Desaphy, Jérémy; Bret, Guillaume; Rognan, Didier; Kellenberger, Esther

2015-01-01

The sc-PDB database (available at http://bioinfo-pharma.u-strasbg.fr/scPDB/) is a comprehensive and up-to-date selection of ligandable binding sites of the Protein Data Bank. Sites are defined from complexes between a protein and a pharmacological ligand. The database provides the all-atom description of the protein, its ligand, their binding site and their binding mode. Currently, the sc-PDB archive registers 9283 binding sites from 3678 unique proteins and 5608 unique ligands. The sc-PDB database was publicly launched in 2004 with the aim of providing structure files suitable for computational approaches to drug design, such as docking. During the last 10 years we have improved and standardized the processes for (i) identifying binding sites, (ii) correcting structures, (iii) annotating protein function and ligand properties and (iv) characterizing their binding mode. This paper presents the latest enhancements in the database, specifically pertaining to the representation of molecular interaction and to the similarity between ligand/protein binding patterns. The new website puts emphasis in pictorial analysis of data. PMID:25300483

MiCroKit 3.0: an integrated database of midbody, centrosome and kinetochore.

PubMed

Ren, Jian; Liu, Zexian; Gao, Xinjiao; Jin, Changjiang; Ye, Mingliang; Zou, Hanfa; Wen, Longping; Zhang, Zhaolei; Xue, Yu; Yao, Xuebiao

2010-01-01

During cell division/mitosis, a specific subset of proteins is spatially and temporally assembled into protein super complexes in three distinct regions, i.e. centrosome/spindle pole, kinetochore/centromere and midbody/cleavage furrow/phragmoplast/bud neck, and modulates cell division process faithfully. Although many experimental efforts have been carried out to investigate the characteristics of these proteins, no integrated database was available. Here, we present the MiCroKit database (http://microkit.biocuckoo.org) of proteins that localize in midbody, centrosome and/or kinetochore. We collected into the MiCroKit database experimentally verified microkit proteins from the scientific literature that have unambiguous supportive evidence for subcellular localization under fluorescent microscope. The current version of MiCroKit 3.0 provides detailed information for 1489 microkit proteins from seven model organisms, including Saccharomyces cerevisiae, Schizasaccharomyces pombe, Caenorhabditis elegans, Drosophila melanogaster, Xenopus laevis, Mus musculus and Homo sapiens. Moreover, the orthologous information was provided for these microkit proteins, and could be a useful resource for further experimental identification. The online service of MiCroKit database was implemented in PHP + MySQL + JavaScript, while the local packages were developed in JAVA 1.5 (J2SE 5.0).

MiCroKit 3.0: an integrated database of midbody, centrosome and kinetochore

PubMed Central

Liu, Zexian; Gao, Xinjiao; Jin, Changjiang; Ye, Mingliang; Zou, Hanfa; Wen, Longping; Zhang, Zhaolei; Xue, Yu; Yao, Xuebiao

2010-01-01

During cell division/mitosis, a specific subset of proteins is spatially and temporally assembled into protein super complexes in three distinct regions, i.e. centrosome/spindle pole, kinetochore/centromere and midbody/cleavage furrow/phragmoplast/bud neck, and modulates cell division process faithfully. Although many experimental efforts have been carried out to investigate the characteristics of these proteins, no integrated database was available. Here, we present the MiCroKit database (http://microkit.biocuckoo.org) of proteins that localize in midbody, centrosome and/or kinetochore. We collected into the MiCroKit database experimentally verified microkit proteins from the scientific literature that have unambiguous supportive evidence for subcellular localization under fluorescent microscope. The current version of MiCroKit 3.0 provides detailed information for 1489 microkit proteins from seven model organisms, including Saccharomyces cerevisiae, Schizasaccharomyces pombe, Caenorhabditis elegans, Drosophila melanogaster, Xenopus laevis, Mus musculus and Homo sapiens. Moreover, the orthologous information was provided for these microkit proteins, and could be a useful resource for further experimental identification. The online service of MiCroKit database was implemented in PHP + MySQL + JavaScript, while the local packages were developed in JAVA 1.5 (J2SE 5.0). PMID:19783819

SASD: the Synthetic Alternative Splicing Database for identifying novel isoform from proteomics

PubMed Central

2013-01-01

Background Alternative splicing is an important and widespread mechanism for generating protein diversity and regulating protein expression. High-throughput identification and analysis of alternative splicing in the protein level has more advantages than in the mRNA level. The combination of alternative splicing database and tandem mass spectrometry provides a powerful technique for identification, analysis and characterization of potential novel alternative splicing protein isoforms from proteomics. Therefore, based on the peptidomic database of human protein isoforms for proteomics experiments, our objective is to design a new alternative splicing database to 1) provide more coverage of genes, transcripts and alternative splicing, 2) exclusively focus on the alternative splicing, and 3) perform context-specific alternative splicing analysis. Results We used a three-step pipeline to create a synthetic alternative splicing database (SASD) to identify novel alternative splicing isoforms and interpret them at the context of pathway, disease, drug and organ specificity or custom gene set with maximum coverage and exclusive focus on alternative splicing. First, we extracted information on gene structures of all genes in the Ensembl Genes 71 database and incorporated the Integrated Pathway Analysis Database. Then, we compiled artificial splicing transcripts. Lastly, we translated the artificial transcripts into alternative splicing peptides. The SASD is a comprehensive database containing 56,630 genes (Ensembl gene IDs), 95,260 transcripts (Ensembl transcript IDs), and 11,919,779 Alternative Splicing peptides, and also covering about 1,956 pathways, 6,704 diseases, 5,615 drugs, and 52 organs. The database has a web-based user interface that allows users to search, display and download a single gene/transcript/protein, custom gene set, pathway, disease, drug, organ related alternative splicing. Moreover, the quality of the database was validated with comparison to other known databases and two case studies: 1) in liver cancer and 2) in breast cancer. Conclusions The SASD provides the scientific community with an efficient means to identify, analyze, and characterize novel Exon Skipping and Intron Retention protein isoforms from mass spectrometry and interpret them at the context of pathway, disease, drug and organ specificity or custom gene set with maximum coverage and exclusive focus on alternative splicing. PMID:24267658

Prediction of protein-protein interactions from amino acid sequences with ensemble extreme learning machines and principal component analysis.

PubMed

You, Zhu-Hong; Lei, Ying-Ke; Zhu, Lin; Xia, Junfeng; Wang, Bing

2013-01-01

Protein-protein interactions (PPIs) play crucial roles in the execution of various cellular processes and form the basis of biological mechanisms. Although large amount of PPIs data for different species has been generated by high-throughput experimental techniques, current PPI pairs obtained with experimental methods cover only a fraction of the complete PPI networks, and further, the experimental methods for identifying PPIs are both time-consuming and expensive. Hence, it is urgent and challenging to develop automated computational methods to efficiently and accurately predict PPIs. We present here a novel hierarchical PCA-EELM (principal component analysis-ensemble extreme learning machine) model to predict protein-protein interactions only using the information of protein sequences. In the proposed method, 11188 protein pairs retrieved from the DIP database were encoded into feature vectors by using four kinds of protein sequences information. Focusing on dimension reduction, an effective feature extraction method PCA was then employed to construct the most discriminative new feature set. Finally, multiple extreme learning machines were trained and then aggregated into a consensus classifier by majority voting. The ensembling of extreme learning machine removes the dependence of results on initial random weights and improves the prediction performance. When performed on the PPI data of Saccharomyces cerevisiae, the proposed method achieved 87.00% prediction accuracy with 86.15% sensitivity at the precision of 87.59%. Extensive experiments are performed to compare our method with state-of-the-art techniques Support Vector Machine (SVM). Experimental results demonstrate that proposed PCA-EELM outperforms the SVM method by 5-fold cross-validation. Besides, PCA-EELM performs faster than PCA-SVM based method. Consequently, the proposed approach can be considered as a new promising and powerful tools for predicting PPI with excellent performance and less time.

Characterization of the African Swine Fever Virus Decapping Enzyme during Infection

PubMed Central

Quintas, Ana; Pérez-Núñez, Daniel; Sánchez, Elena G.; Nogal, Maria L.; Hentze, Matthias W.; Castelló, Alfredo

2017-01-01

ABSTRACT African swine fever virus (ASFV) infection is characterized by a progressive decrease in cellular protein synthesis with a concomitant increase in viral protein synthesis, though the mechanism by which the virus achieves this is still unknown. Decrease of cellular mRNA is observed during ASFV infection, suggesting that inhibition of cellular proteins is due to an active mRNA degradation process. ASFV carries a gene (Ba71V D250R/Malawi g5R) that encodes a decapping protein (ASFV-DP) that has a Nudix hydrolase motif and decapping activity in vitro. Here, we show that ASFV-DP was expressed from early times and accumulated throughout the infection with a subcellular localization typical of the endoplasmic reticulum, colocalizing with the cap structure and interacting with the ribosomal protein L23a. ASFV-DP was capable of interaction with poly(A) RNA in cultured cells, primarily mediated by the N-terminal region of the protein. ASFV-DP also interacted with viral and cellular RNAs in the context of infection, and its overexpression in infected cells resulted in decreased levels of both types of transcripts. This study points to ASFV-DP as a viral decapping enzyme involved in both the degradation of cellular mRNA and the regulation of viral transcripts. IMPORTANCE Virulent ASFV strains cause a highly infectious and lethal disease in domestic pigs for which there is no vaccine. Since 2007, an outbreak in the Caucasus region has spread to Russia, jeopardizing the European pig population and making it essential to deepen knowledge about the virus. Here, we demonstrate that ASFV-DP is a novel RNA-binding protein implicated in the regulation of mRNA metabolism during infection, making it a good target for vaccine development. PMID:29021398

Characterization of the African Swine Fever Virus Decapping Enzyme during Infection.

PubMed

Quintas, Ana; Pérez-Núñez, Daniel; Sánchez, Elena G; Nogal, Maria L; Hentze, Matthias W; Castelló, Alfredo; Revilla, Yolanda

2017-12-15

African swine fever virus (ASFV) infection is characterized by a progressive decrease in cellular protein synthesis with a concomitant increase in viral protein synthesis, though the mechanism by which the virus achieves this is still unknown. Decrease of cellular mRNA is observed during ASFV infection, suggesting that inhibition of cellular proteins is due to an active mRNA degradation process. ASFV carries a gene (Ba71V D250R/Malawi g5R) that encodes a decapping protein (ASFV-DP) that has a Nudix hydrolase motif and decapping activity in vitro Here, we show that ASFV-DP was expressed from early times and accumulated throughout the infection with a subcellular localization typical of the endoplasmic reticulum, colocalizing with the cap structure and interacting with the ribosomal protein L23a. ASFV-DP was capable of interaction with poly(A) RNA in cultured cells, primarily mediated by the N-terminal region of the protein. ASFV-DP also interacted with viral and cellular RNAs in the context of infection, and its overexpression in infected cells resulted in decreased levels of both types of transcripts. This study points to ASFV-DP as a viral decapping enzyme involved in both the degradation of cellular mRNA and the regulation of viral transcripts. IMPORTANCE Virulent ASFV strains cause a highly infectious and lethal disease in domestic pigs for which there is no vaccine. Since 2007, an outbreak in the Caucasus region has spread to Russia, jeopardizing the European pig population and making it essential to deepen knowledge about the virus. Here, we demonstrate that ASFV-DP is a novel RNA-binding protein implicated in the regulation of mRNA metabolism during infection, making it a good target for vaccine development. Copyright © 2017 Quintas et al.

A novel angiotensin II type 1 receptor-associated protein induces cellular hypertrophy in rat vascular smooth muscle and renal proximal tubular cells.

PubMed

Guo, Deng-Fu; Tardif, Valerie; Ghelima, Karin; Chan, John S D; Ingelfinger, Julie R; Chen, XiangMei; Chenier, Isabelle

2004-05-14

Angiotensin II stimulates cellular hypertrophy in cultured vascular smooth muscle and renal proximal tubular cells. This effect is believed to be one of earliest morphological changes of heart and renal failure. However, the precise molecular mechanism involved in angiotensin II-induced hypertrophy is poorly understood. In the present study we report the isolation of a novel angiotensin II type 1 receptor-associated protein. It encodes a 531-amino acid protein. Its mRNA is detected in all human tissues examined but highly expressed in the human kidney, pancreas, heart, and human embryonic kidney cells as well as rat vascular smooth muscle and renal proximal tubular cells. Protein synthesis and relative cell size analyzed by flow cytometry studies indicate that overexpression of the novel angiotensin II type 1 receptor-associated protein induces cellular hypertrophy in cultured rat vascular smooth muscle and renal proximal tubular cells. In contrast, the hypertrophic effects was reversed in renal proximal tubular cell lines expressing the novel gene in the antisense orientation and its dominant negative mutant, which lacks the last 101 amino acids in its carboxyl-terminal tail. The hypertrophic effects are at least in part mediated via protein kinase B activation or cyclin-dependent kinase inhibitor, p27(kip1) protein expression level in vascular smooth muscle, and renal proximal tubular cells. Moreover, angiotensin II could not stimulate cellular hypertrophy in renal proximal tubular cells expressing the novel gene in the antisense orientation and its mutant. These findings may provide new molecular mechanisms to understand hypertrophic agents such as angiotensin II-induced cellular hypertrophy.

Coordinated regulation by two VPS9 domain-containing guanine nucleotide exchange factors in small GTPase Rab5 signaling pathways in fission yeast

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tsukamoto, Yuta; Kagiwada, Satoshi; Shimazu, Sayuri

The small GTPase Rab5 is reported to regulate various cellular functions, such as vesicular transport and endocytosis. VPS9 domain-containing proteins are thought to activate Rab5(s) by their guanine-nucleotide exchange activities. Numerous VPS9 proteins have been identified and are structurally conserved from yeast to mammalian cells. However, the functional relationships among VPS9 proteins in cells remain unclear. Only one Rab5 and two VPS9 proteins were identified in the Schizosaccharomyces pombe genome. Here, we examined the cellular function of two VPS9 proteins and the relationship between these proteins in cellular functions. Vps901-GFP and Vps902-GFP exhibited dotted signals in vegetative and differentiated cells.more » vps901 deletion mutant (Δvps901) cells exhibited a phenotype deficient in the mating process and responses to high concentrations of ions, such as calcium and metals, and Δvps901Δvps902 double mutant cells exhibited round cell shapes similar to ypt5-909 (Rab5 mutant allele) cells. Deletion of both vps901 and vps902 genes completely abolished the mating process and responses to various stresses. A lack of vacuole formation and aberrant inner cell membrane structures were also observed in Δvps901Δvps902 cells by electron microscopy. These data strongly suggest that Vps901 and Vps902 are cooperatively involved in the regulation of cellular functions, such as cell morphology, sexual development, response to ion stresses, and vacuole formation, via Rab5 signaling pathways in fission yeast cells. - Highlights: • Roles of Rab5 activator VPS9 proteins in cellular functions. • Cooperation between VPS9 proteins in Rab5 signaling pathway. • Roles of each VPS9 protein in Rab5 signaling pathway are discussed.« less

Fasting and refeeding induces changes in the mouse hepatic lipid droplet proteome.

PubMed

Kramer, David A; Quiroga, Ariel D; Lian, Jihong; Fahlman, Richard P; Lehner, Richard

2018-06-15

During fasting, the liver increases lipid storage as a mean to reserve and provide energy for vital cellular functions. After re-feeding, hepatocytes rapidly decrease the amount of triacylglycerol that is stored in lipid droplets (LDs), visible as the size of hepatic LDs significantly decreases after re-feeding. Little is known about the changes in the liver LD proteome that occur during the fasting/re-feeding transition. This study aimed to investigate the hepatic LD proteome in fasted and re-fed conditions in the mouse. Using label-free LC-MS/MS analysis the relative abundance of 817 proteins was determined in highly purified LDs. Comparative analysis for differential protein abundance with respect to feeding states revealed 130 with higher abundance in LDs from fasted mice and 31 in LDs from re-fed mice. Among proteins observed to have higher abundance on LDs in the fasted state we found perilipin-5, and several mitochondrial and peroxisomal marker proteins, supporting the role of LDs in the provision of substrates for fatty acid oxidation. Proteins of higher abundance upon re-feeding included several peroxisomal and mitochondrial marker proteins and expand our understanding of the dynamic nature of the hepatic LD proteome according to the energetic requirements of the cell. Proteomic investigations have been revealing the complexities and dynamics of cellular LDs from a variety of cell types. As these sub-cellular structures are truly dynamic in nature, our investigations reveal that simply the feeding state of an animal leads to significant changes to the protein composition of LDs and suggest a variety of dynamic interactions with other cellular organelles, such as the mitochondria and peroxisomes. As such, the experimental design for investigations of this cellular structure must consider this dynamic baseline. Lastly our analysis on global protein abundance has revealed the unforeseen high abundance of murine major urinary proteins associated with hepatic lipid droplets, which warrants further investigations. Crown Copyright © 2018. Published by Elsevier B.V. All rights reserved.

Why are they missing? : Bioinformatics characterization of missing human proteins.

PubMed

Elguoshy, Amr; Magdeldin, Sameh; Xu, Bo; Hirao, Yoshitoshi; Zhang, Ying; Kinoshita, Naohiko; Takisawa, Yusuke; Nameta, Masaaki; Yamamoto, Keiko; El-Refy, Ali; El-Fiky, Fawzy; Yamamoto, Tadashi

2016-10-21

NeXtProt is a web-based protein knowledge platform that supports research on human proteins. NeXtProt (release 2015-04-28) lists 20,060 proteins, among them, 3373 canonical proteins (16.8%) lack credible experimental evidence at protein level (PE2:PE5). Therefore, they are considered as "missing proteins". A comprehensive bioinformatic workflow has been proposed to analyze these "missing" proteins. The aims of current study were to analyze physicochemical properties, existence and distribution of the tryptic cleavage sites, and to pinpoint the signature peptides of the missing proteins. Our findings showed that 23.7% of missing proteins were hydrophobic proteins possessing transmembrane domains (TMD). Also, forty missing entries generate tryptic peptides were either out of mass detection range (>30aa) or mapped to different proteins (<9aa). Additionally, 21% of missing entries didn't generate any unique tryptic peptides. In silico endopeptidase combination strategy increased the possibility of missing proteins identification. Coherently, using both mature protein database and signal peptidome database could be a promising option to identify some missing proteins by targeting their unique N-terminal tryptic peptide from mature protein database and or C-terminus tryptic peptide from signal peptidome database. In conclusion, Identification of missing protein requires additional consideration during sample preparation, extraction, digestion and data analysis to increase its incidence of identification. Copyright © 2016. Published by Elsevier B.V.

Harnessing Drug Resistance: Using ABC Transporter Proteins To Target Cancer Cells

PubMed Central

Leitner, Heather M.; Kachadourian, Remy; Day, Brian J.

2007-01-01

The ATP-binding cassette (ABC) class of proteins is one of the most functionally diverse transporter families found in biological systems. Although the abundance of ABC proteins varies between species, they are highly conserved in sequence and often demonstrate similar functions across prokaryotic and eukaryotic organisms. Beginning with a brief summary of the events leading to our present day knowledge of ABC transporters, the purpose of this review is to discuss the potential for utilizing ABC transporters as a means for cellular glutathione (GSH) modulation. GSH is one of the most abundant thiol antioxidants in cells. It is involved in cellular division, protein and DNA synthesis, maintenance of cellular redox status and xenobiotic metabolism. Cellular GSH levels are often altered in many disease states including cancer. Over the past two decades there has been considerable emphasis on methods to sensitize cancer cells to chemotherapeutics and ionization radiation therapy by GSH depletion. We contend that ABC transporters, particularly multi-drug resistant proteins (MRPs), may be used as therapeutic targets for applications aimed at modulation of GSH levels. This review will emphasize MRP-mediated modulation of intracellular GSH levels as a potential alternative and adjunctive approach for cancer therapy. PMID:17585883

Engineering microbial phenotypes through rewiring of genetic networks

PubMed Central

Rodrigues, Rui T.L.; Lee, Sangjin; Haines, Matthew

2017-01-01

Abstract The ability to program cellular behaviour is a major goal of synthetic biology, with applications in health, agriculture and chemicals production. Despite efforts to build ‘orthogonal’ systems, interactions between engineered genetic circuits and the endogenous regulatory network of a host cell can have a significant impact on desired functionality. We have developed a strategy to rewire the endogenous cellular regulatory network of yeast to enhance compatibility with synthetic protein and metabolite production. We found that introducing novel connections in the cellular regulatory network enabled us to increase the production of heterologous proteins and metabolites. This strategy is demonstrated in yeast strains that show significantly enhanced heterologous protein expression and higher titers of terpenoid production. Specifically, we found that the addition of transcriptional regulation between free radical induced signalling and nitrogen regulation provided robust improvement of protein production. Assessment of rewired networks revealed the importance of key topological features such as high betweenness centrality. The generation of rewired transcriptional networks, selection for specific phenotypes, and analysis of resulting library members is a powerful tool for engineering cellular behavior and may enable improved integration of heterologous protein and metabolite pathways. PMID:28369627

Systematic analyses of the ultraviolet radiation resistance-associated gene product (UVRAG) protein interactome by tandem affinity purification.

PubMed

Son, Ji-Hye; Hwang, Eurim C; Kim, Joungmok

2016-03-01

Ultraviolet radiation resistance-associated gene product (UVRAG) was originally identified as a protein involved in cellular responses to UV irradiation. Subsequent studies have demonstrated that UVRAG plays as an important role in autophagy, a lysosome-dependent catabolic program, as a part of a pro-autophagy PIK3C3/VPS34 lipid kinase complex. Several recent studies have shown that UVRAG is also involved in autophagy-independent cellular functions, such as DNA repair/stability and vesicular trafficking/fusion. Here, we examined the UVRAG protein interactome to obtain information about its functional network. To this end, we screened UVRAG-interacting proteins using a tandem affinity purification method coupled with MALDI-TOF/MS analysis. Our results demonstrate that UVRAG interacts with various proteins involved in a wide spectrum of cellular functions, including genome stability, protein translational elongation, protein localization (trafficking), vacuole organization, transmembrane transport as well as autophagy. Notably, the interactome list of high-confidence UVRAG-interacting proteins is enriched for proteins involved in the regulation of genome stability. Our systematic UVRAG interactome analysis should provide important clues for understanding a variety of UVRAG functions.

Genome-Wide Identification and Expression Profiling Analysis of the Xyloglucan Endotransglucosylase/Hydrolase Gene Family in Tobacco (Nicotiana tabacum L.).

PubMed

Wang, Meng; Xu, Zongchang; Ding, Anming; Kong, Yingzhen

2018-05-24

Xyloglucan endotransglucosylase/hydrolase genes ( XTHs ) encode enzymes required for the reconstruction and modification of xyloglucan backbones, which will result in changes of cell wall extensibility during growth. A total of 56 NtXTH genes were identified from common tobacco, and 50 cDNA fragments were verified by PCR amplification. The 56 NtXTH genes could be classified into two subfamilies: Group I/II and Group III according to their phylogenetic relationships. The gene structure, chromosomal localization, conserved protein domains prediction, sub-cellular localization of NtXTH proteins and evolutionary relationships among Nicotiana tabacum , Nicotiana sylvestrisis , Nicotiana tomentosiformis , Arabidopsis , and rice were also analyzed. The NtXTHs expression profiles analyzed by the TobEA database and qRT-PCR revealed that NtXTHs display different expression patterns in different tissues. Notably, the expression patterns of 12 NtXTHs responding to environment stresses, including salinity, alkali, heat, chilling, and plant hormones, including IAA and brassinolide, were characterized. All the results would be useful for the function study of NtXTHs during different growth cycles and stresses.

ProteinWorldDB: querying radical pairwise alignments among protein sets from complete genomes

PubMed Central

Otto, Thomas Dan; Catanho, Marcos; Tristão, Cristian; Bezerra, Márcia; Fernandes, Renan Mathias; Elias, Guilherme Steinberger; Scaglia, Alexandre Capeletto; Bovermann, Bill; Berstis, Viktors; Lifschitz, Sergio; de Miranda, Antonio Basílio; Degrave, Wim

2010-01-01

Motivation: Many analyses in modern biological research are based on comparisons between biological sequences, resulting in functional, evolutionary and structural inferences. When large numbers of sequences are compared, heuristics are often used resulting in a certain lack of accuracy. In order to improve and validate results of such comparisons, we have performed radical all-against-all comparisons of 4 million protein sequences belonging to the RefSeq database, using an implementation of the Smith–Waterman algorithm. This extremely intensive computational approach was made possible with the help of World Community Grid™, through the Genome Comparison Project. The resulting database, ProteinWorldDB, which contains coordinates of pairwise protein alignments and their respective scores, is now made available. Users can download, compare and analyze the results, filtered by genomes, protein functions or clusters. ProteinWorldDB is integrated with annotations derived from Swiss-Prot, Pfam, KEGG, NCBI Taxonomy database and gene ontology. The database is a unique and valuable asset, representing a major effort to create a reliable and consistent dataset of cross-comparisons of the whole protein content encoded in hundreds of completely sequenced genomes using a rigorous dynamic programming approach. Availability: The database can be accessed through http://proteinworlddb.org Contact: otto@fiocruz.br PMID:20089515

Cellular Factors Required for Lassa Virus Budding

PubMed Central

Urata, Shuzo; Noda, Takeshi; Kawaoka, Yoshihiro; Yokosawa, Hideyoshi; Yasuda, Jiro

2006-01-01

It is known that Lassa virus Z protein is sufficient for the release of virus-like particles (VLPs) and that it has two L domains, PTAP and PPPY, in its C terminus. However, little is known about the cellular factor for Lassa virus budding. We examined which cellular factors are used in Lassa virus Z budding. We demonstrated that Lassa Z protein efficiently produces VLPs and uses cellular factors, Vps4A, Vps4B, and Tsg101, in budding, suggesting that Lassa virus budding uses the multivesicular body pathway functionally. Our data may provide a clue to develop an effective antiviral strategy for Lassa virus. PMID:16571837

p53-Mediated Cellular Response to DNA Damage in Cells with Replicative Hepatitis B Virus

NASA Astrophysics Data System (ADS)

Puisieux, Alain; Ji, Jingwei; Guillot, Celine; Legros, Yann; Soussi, Thierry; Isselbacher, Kurt; Ozturk, Mehmet

1995-02-01

Wild-type p53 acts as a tumor suppressor gene by protecting cells from deleterious effects of genotoxic agents through the induction of a G_1/S arrest or apoptosis as a response to DNA damage. Transforming proteins of several oncogenic DNA viruses inactivate tumor suppressor activity of p53 by blocking this cellular response. To test whether hepatitis B virus displays a similar effect, we studied the p53-mediated cellular response to DNA damage in 2215 hepatoma cells with replicative hepatitis B virus. We demonstrate that hepatitis B virus replication does not interfere with known cellular functions of p53 protein.

A Reference Viral Database (RVDB) To Enhance Bioinformatics Analysis of High-Throughput Sequencing for Novel Virus Detection

PubMed Central

Goodacre, Norman; Aljanahi, Aisha; Nandakumar, Subhiksha; Mikailov, Mike

2018-01-01

ABSTRACT Detection of distantly related viruses by high-throughput sequencing (HTS) is bioinformatically challenging because of the lack of a public database containing all viral sequences, without abundant nonviral sequences, which can extend runtime and obscure viral hits. Our reference viral database (RVDB) includes all viral, virus-related, and virus-like nucleotide sequences (excluding bacterial viruses), regardless of length, and with overall reduced cellular sequences. Semantic selection criteria (SEM-I) were used to select viral sequences from GenBank, resulting in a first-generation viral database (VDB). This database was manually and computationally reviewed, resulting in refined, semantic selection criteria (SEM-R), which were applied to a new download of updated GenBank sequences to create a second-generation VDB. Viral entries in the latter were clustered at 98% by CD-HIT-EST to reduce redundancy while retaining high viral sequence diversity. The viral identity of the clustered representative sequences (creps) was confirmed by BLAST searches in NCBI databases and HMMER searches in PFAM and DFAM databases. The resulting RVDB contained a broad representation of viral families, sequence diversity, and a reduced cellular content; it includes full-length and partial sequences and endogenous nonretroviral elements, endogenous retroviruses, and retrotransposons. Testing of RVDBv10.2, with an in-house HTS transcriptomic data set indicated a significantly faster run for virus detection than interrogating the entirety of the NCBI nonredundant nucleotide database, which contains all viral sequences but also nonviral sequences. RVDB is publically available for facilitating HTS analysis, particularly for novel virus detection. It is meant to be updated on a regular basis to include new viral sequences added to GenBank. IMPORTANCE To facilitate bioinformatics analysis of high-throughput sequencing (HTS) data for the detection of both known and novel viruses, we have developed a new reference viral database (RVDB) that provides a broad representation of different virus species from eukaryotes by including all viral, virus-like, and virus-related sequences (excluding bacteriophages), regardless of their size. In particular, RVDB contains endogenous nonretroviral elements, endogenous retroviruses, and retrotransposons. Sequences were clustered to reduce redundancy while retaining high viral sequence diversity. A particularly useful feature of RVDB is the reduction of cellular sequences, which can enhance the run efficiency of large transcriptomic and genomic data analysis and increase the specificity of virus detection. PMID:29564396

A Reference Viral Database (RVDB) To Enhance Bioinformatics Analysis of High-Throughput Sequencing for Novel Virus Detection.

PubMed

Goodacre, Norman; Aljanahi, Aisha; Nandakumar, Subhiksha; Mikailov, Mike; Khan, Arifa S

2018-01-01

Detection of distantly related viruses by high-throughput sequencing (HTS) is bioinformatically challenging because of the lack of a public database containing all viral sequences, without abundant nonviral sequences, which can extend runtime and obscure viral hits. Our reference viral database (RVDB) includes all viral, virus-related, and virus-like nucleotide sequences (excluding bacterial viruses), regardless of length, and with overall reduced cellular sequences. Semantic selection criteria (SEM-I) were used to select viral sequences from GenBank, resulting in a first-generation viral database (VDB). This database was manually and computationally reviewed, resulting in refined, semantic selection criteria (SEM-R), which were applied to a new download of updated GenBank sequences to create a second-generation VDB. Viral entries in the latter were clustered at 98% by CD-HIT-EST to reduce redundancy while retaining high viral sequence diversity. The viral identity of the clustered representative sequences (creps) was confirmed by BLAST searches in NCBI databases and HMMER searches in PFAM and DFAM databases. The resulting RVDB contained a broad representation of viral families, sequence diversity, and a reduced cellular content; it includes full-length and partial sequences and endogenous nonretroviral elements, endogenous retroviruses, and retrotransposons. Testing of RVDBv10.2, with an in-house HTS transcriptomic data set indicated a significantly faster run for virus detection than interrogating the entirety of the NCBI nonredundant nucleotide database, which contains all viral sequences but also nonviral sequences. RVDB is publically available for facilitating HTS analysis, particularly for novel virus detection. It is meant to be updated on a regular basis to include new viral sequences added to GenBank. IMPORTANCE To facilitate bioinformatics analysis of high-throughput sequencing (HTS) data for the detection of both known and novel viruses, we have developed a new reference viral database (RVDB) that provides a broad representation of different virus species from eukaryotes by including all viral, virus-like, and virus-related sequences (excluding bacteriophages), regardless of their size. In particular, RVDB contains endogenous nonretroviral elements, endogenous retroviruses, and retrotransposons. Sequences were clustered to reduce redundancy while retaining high viral sequence diversity. A particularly useful feature of RVDB is the reduction of cellular sequences, which can enhance the run efficiency of large transcriptomic and genomic data analysis and increase the specificity of virus detection.

Algorithms for database-dependent search of MS/MS data.

PubMed

Matthiesen, Rune

2013-01-01

The frequent used bottom-up strategy for identification of proteins and their associated modifications generate nowadays typically thousands of MS/MS spectra that normally are matched automatically against a protein sequence database. Search engines that take as input MS/MS spectra and a protein sequence database are referred as database-dependent search engines. Many programs both commercial and freely available exist for database-dependent search of MS/MS spectra and most of the programs have excellent user documentation. The aim here is therefore to outline the algorithm strategy behind different search engines rather than providing software user manuals. The process of database-dependent search can be divided into search strategy, peptide scoring, protein scoring, and finally protein inference. Most efforts in the literature have been put in to comparing results from different software rather than discussing the underlining algorithms. Such practical comparisons can be cluttered by suboptimal implementation and the observed differences are frequently caused by software parameters settings which have not been set proper to allow even comparison. In other words an algorithmic idea can still be worth considering even if the software implementation has been demonstrated to be suboptimal. The aim in this chapter is therefore to split the algorithms for database-dependent searching of MS/MS data into the above steps so that the different algorithmic ideas become more transparent and comparable. Most search engines provide good implementations of the first three data analysis steps mentioned above, whereas the final step of protein inference are much less developed for most search engines and is in many cases performed by an external software. The final part of this chapter illustrates how protein inference is built into the VEMS search engine and discusses a stand-alone program SIR for protein inference that can import a Mascot search result.

The structure and dipole moment of globular proteins in solution and crystalline states: use of NMR and X-ray databases for the numerical calculation of dipole moment.

PubMed

Takashima, S

2001-04-05

The large dipole moment of globular proteins has been well known because of the detailed studies using dielectric relaxation and electro-optical methods. The search for the origin of these dipolemoments, however, must be based on the detailed knowledge on protein structure with atomic resolutions. At present, we have two sources of information on the structure of protein molecules: (1) x-ray databases obtained in crystalline state; (2) NMR databases obtained in solution state. While x-ray databases consist of only one model, NMR databases, because of the fluctuation of the protein folding in solution, consist of a number of models, thus enabling the computation of dipole moment repeated for all these models. The aim of this work, using these databases, is the detailed investigation on the interdependence between the structure and dipole moment of protein molecules. The dipole moment of protein molecules has roughly two components: one dipole moment is due to surface charges and the other, core dipole moment, is due to polar groups such as N--H and C==O bonds. The computation of surface charge dipole moment consists of two steps: (A) calculation of the pK shifts of charged groups for electrostatic interactions and (B) calculation of the dipole moment using the pK corrected for electrostatic shifts. The dipole moments of several proteins were computed using both NMR and x-ray databases. The dipole moments of these two sets of calculations are, with a few exceptions, in good agreement with one another and also with measured dipole moments.

Inconsistencies in the red blood cell membrane proteome analysis: generation of a database for research and diagnostic applications

PubMed Central

Hegedűs, Tamás; Chaubey, Pururawa Mayank; Várady, György; Szabó, Edit; Sarankó, Hajnalka; Hofstetter, Lia; Roschitzki, Bernd; Sarkadi, Balázs

2015-01-01

Based on recent results, the determination of the easily accessible red blood cell (RBC) membrane proteins may provide new diagnostic possibilities for assessing mutations, polymorphisms or regulatory alterations in diseases. However, the analysis of the current mass spectrometry-based proteomics datasets and other major databases indicates inconsistencies—the results show large scattering and only a limited overlap for the identified RBC membrane proteins. Here, we applied membrane-specific proteomics studies in human RBC, compared these results with the data in the literature, and generated a comprehensive and expandable database using all available data sources. The integrated web database now refers to proteomic, genetic and medical databases as well, and contains an unexpected large number of validated membrane proteins previously thought to be specific for other tissues and/or related to major human diseases. Since the determination of protein expression in RBC provides a method to indicate pathological alterations, our database should facilitate the development of RBC membrane biomarker platforms and provide a unique resource to aid related further research and diagnostics. Database URL: http://rbcc.hegelab.org PMID:26078478

[Non-ciliary functions of cilia proteins].

PubMed

Taulet, Nicolas; Delaval, Bénédicte

2014-11-01

Cilia proteins have long been characterized for their role in cilia formation and function, and their implications in ciliopathies. However, several cellular defects induced by cilia proteins deregulation suggest that they could have non-ciliary roles. Indeed, several non-ciliary functions have been recently characterized for cilia proteins including roles in intra-cellular and in vesicular transport, in spindle orientation or in the maintenance of genomic stability. These observations thus raise the crucial question of the contribution of non-ciliary functions of cilia proteins to the pathological manifestations associated with ciliopathies such as polycystic kidney disease. © 2014 médecine/sciences – Inserm.

Origins of the protein synthesis cycle

NASA Technical Reports Server (NTRS)

Fox, S. W.

1981-01-01

Largely derived from experiments in molecular evolution, a theory of protein synthesis cycles has been constructed. The sequence begins with ordered thermal proteins resulting from the self-sequencing of mixed amino acids. Ordered thermal proteins then aggregate to cell-like structures. When they contained proteinoids sufficiently rich in lysine, the structures were able to synthesize offspring peptides. Since lysine-rich proteinoid (LRP) also catalyzes the polymerization of nucleoside triphosphate to polynucleotides, the same microspheres containing LRP could have synthesized both original cellular proteins and cellular nucleic acids. The LRP within protocells would have provided proximity advantageous for the origin and evolution of the genetic code.