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Hsueh-Fen Juan ( 阮雪芬 ) NTUT Oct 9, 2003 Bioinformatics Data Analysis in Proteomics
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Yuki Juan’s Systems Biology Lab Outline Introduction to proteomics Protein Functions Protein-protein interactions Pathways Protein Structures and Drug Discovery Bioinformatics on the Web
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Yuki Juan’s Systems Biology Lab Proteomics Network Identify Proteins Drug Discovery Structures Protein-Protein Interactions Pathways Protein Functions
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Yuki Juan’s Systems Biology Lab Outline Introduction to proteomics Protein Functions Protein-protein interactions Pathways Protein Structures and Drug Discovery Bioinformatics on the Web
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Yuki Juan’s Systems Biology Lab What Is Proteomes ?
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Yuki Juan’s Systems Biology Lab Proteomes Gene + Chromosome Genome Protein +Genome Proteome Proteomes are dynamics Proteome changes as a function of: time development extracellular condition intracellular condition
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Yuki Juan’s Systems Biology Lab Definitions of Proteomics First coined in 1995 by Wilkins Be defined as the large-scale characterization of the entire protein complement of a cell line, tissue, or organism. The study of proteomes Goal: -To obtain a more global and integrated view of biology by studying all the proteins of a cell rather than each one individually.
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Yuki Juan’s Systems Biology Lab Proteomics Origins In 1975, the introduction of the 2D gel by O’Farrell who began mapping proteins from E. coli. The first major technology to emerge for the identification of proteins was the sequencing of proteins by Edman degradation picomole MS technology has replaced Edman degradation to identify proteins femtomole
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Yuki Juan’s Systems Biology Lab How Proteomics Can Help Drug Development http://www.sciam.com.tw/read/readshow.asp?FDocNo=63&CL=18
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Yuki Juan’s Systems Biology Lab Types of Proteomics and Their Applications to Biology
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Yuki Juan’s Systems Biology Lab Mechanisms by Which a Single Gene Can Give Rise to Multiple Gene Products In bacteria, 1 or 2 proteins/gene In yeast, 3 proteins/gene In human, 3 or more proteins/gene Glycosylation
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Yuki Juan’s Systems Biology Lab Two-dimensional Gel Approach Nature 2000, 405, 837-846
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Yuki Juan’s Systems Biology Lab 3.510 42 60 70 150 kDa pH Increase of 50% Decrease of 50% Unmatched spots Matched spots Image Matching
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Yuki Juan’s Systems Biology Lab www.expasy.ch/ch2d
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Yuki Juan’s Systems Biology Lab
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http://www.expasy.ch/melanie/
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Yuki Juan’s Systems Biology Lab
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Standard Proteome Analysis by 2DE-MS Current Opinion in Chemical Biology 2000, 4:489–494 Mass Fingerprint Searching in http://www.expas y.ch/tools/peptide nt.html
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Yuki Juan’s Systems Biology Lab ControlTreated sample Example: Gel
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Yuki Juan’s Systems Biology Lab + - gel membrane Transfer proteins from gel to PVDF membrane Sequencing (Very time-Consuming)
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Yuki Juan’s Systems Biology Lab Mass spectrometry
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Yuki Juan’s Systems Biology Lab For peptide mass fingerprint data
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Yuki Juan’s Systems Biology Lab Species pI and MW
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Yuki Juan’s Systems Biology Lab Peptide Mass Fingerprinting
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Yuki Juan’s Systems Biology Lab World Wide Web Tools for Searching Databases Site nameURLInformation available MOWSE http://srs.hgmp.mrc.ac.uk/cgi-bin/mowsePeptide mass mapping and sequencing ProFoundhttp://prowl.rockefeller.edu/cgi- bin/ProFound Peptide mass mapping and sequencing PeptIdenthttp://www.expasy.ch/tools/peptident.Peptide mass mapping and sequencing PepSeahttp://195.41.108.38/PepSeaIntro.htmlPeptide mass mapping and sequencing MASCOThttp://www.matrixscience.com/Peptide mass mapping and sequencing PepFraghttp://www.proteometrics.com/Peptide mass mapping and sequencing Protein Prospectorhttp://prospector.ucsf.edu/Peptide mass mapping and sequencing FindModhttp://www.expasy.ch/tools/findmod/Posttranslational modification SEAQUESThttp://fields.scripps.edu/sequest/Uninterpreted MS/MS searching FASTA Search Programs http://fasta.bioch.virginia.edu/Protein and nucleotide database searching Cleaved Radioactivity of Phosphopeptides http://fasta.bioch.virginia.edu/crpProtein phosphorylation site mapping
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Yuki Juan’s Systems Biology Lab The Information Stored in Genes Is Expressed by a Multistage Process
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Yuki Juan’s Systems Biology Lab DNA 和蛋白質合成的地方 DNA Proteins Sugar Chain
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Yuki Juan’s Systems Biology Lab Post-translational Modification Genomics provides comprehensive data-bases of sequence information, DNA and mRNA provide no information concerning the activities and post- translational modifications of proteins. The number of documented protein co- and post- translational modifications now exceeds 400 (http: / / abrf.org / index-.cfm/dm.home). The elucidation of protein post-translational modifications is perhaps the most important justification for proteomics as a scientific endeavor.
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Yuki Juan’s Systems Biology Lab Chemical modification Covalent linkage of chemical group to amino acid in protein Several types: Acetylation (common on first amino acid) Phosphorylation (often involved in regulation) Lipidation (attachment to membrane) Glycosylation (common outside cell)
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Yuki Juan’s Systems Biology Lab Processing Removal of part of protein by either: Cleavage (protease cuts protein into smaller pieces) Splicing (self-removal and rejoining of ends)
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Yuki Juan’s Systems Biology Lab
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Post-translational Modification
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Yuki Juan’s Systems Biology Lab Post-translational Modification Glycosylation Mass spectrometric “sequencing” of oligosaccharides Increase the complexity of the proteome
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Yuki Juan’s Systems Biology Lab Outline Introduction to proteomics Protein Functions Protein-protein interactions Pathways Protein Structures and Drug Discovery Bioinformatics on the Web
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Yuki Juan’s Systems Biology Lab Protein Functions Gene Ontology http://www.geneontology.org/ GoMiner http://discover.nci.nih.gov/gominer
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Yuki Juan’s Systems Biology Lab Gene Ontology
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Yuki Juan’s Systems Biology Lab Gene Ontology
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Yuki Juan’s Systems Biology Lab GoMiner
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Yuki Juan’s Systems Biology Lab GoMiner Genome Biology 2003, 4:R28
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Yuki Juan’s Systems Biology Lab Outline Introduction to proteomics Protein Functions Protein-protein interactions Pathways Protein Structures and Drug Discovery Bioinformatics on the Web
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Yuki Juan’s Systems Biology Lab Protein-protein Interactions Introduction Mass Spectrometry Yeast Two-hybrid Assay
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Yuki Juan’s Systems Biology Lab Introduction Kanehisa (2000)said: Interaction NetworkFunction Post-genome informatics (2000)
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Yuki Juan’s Systems Biology Lab Introduction Protein-protein interactions are intrinsic to every cellular process. Form the basis of phenomena DNA replication and transcription Metabolism Signal transduction Cell cycle control Secretion
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Yuki Juan’s Systems Biology Lab Introduction Knowledge of interacting proteins Provide insight into the function of important genes Elucidates relevant pathways Facilitates the identification of potential drug targets Use in developing novel therapeutics
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Yuki Juan’s Systems Biology Lab Interaction Discovery Methods TiBS(2003), 27(12), 633-638.
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Yuki Juan’s Systems Biology Lab Structural Biology of Interactions and Complexes X-ray crystallography Nuclear Magnetic Resonance (NMR) Electron microscopy (EI)
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Yuki Juan’s Systems Biology Lab Examples of Three-dimensional Protein Interactions and Complexes TiBS(2003), 27(12), 633-638.
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Yuki Juan’s Systems Biology Lab The Study of Protein-protein Interaction by Mass Spectrometry bait S14 ? ? ? ? ** ** SDS- PAGE MASS
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Yuki Juan’s Systems Biology Lab Peptide Mass Fingerprinting
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Yuki Juan’s Systems Biology Lab Detecting Protein-protein Interactions by Protein Microarray Science, 2000, 289, 1761-1763
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Yuki Juan’s Systems Biology Lab Yeast Two-hybrid System Useful in the study of various interactions The technology was originally developed during the late 1980's in the laboratory Dr. Stanley Fields (see Fields and Song, 1989, Nature).
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Yuki Juan’s Systems Biology Lab Yeast Two-hybrid Assay GAL4 DNA- binding domain GAL4 DNA- activation domain Nature, 2000
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Yuki Juan’s Systems Biology Lab Yeast Two-hybrid Assay Library-based yeast two-hybrid screening method Nature, 2000
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Yuki Juan’s Systems Biology Lab Information Extraction (IE) A vast amount of data on protein- protein interactions residues in the published literature, which never been entered into databases. IE have been applied to gaining information on protein-protein interactions.
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Yuki Juan’s Systems Biology Lab Mining Literature for Protein-protein Interactions
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Yuki Juan’s Systems Biology Lab Mining Literature for Protein-protein Interactions
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Yuki Juan’s Systems Biology Lab Protein-protein Interactions on the Web Yeast http://depts.washington.edu/sfields/yplm/data/index.html http://portal.curagen.com http://mips.gsf.de/proj/yeast/CYGD/interaction/ http://www.pnas.org/cgi/content/full/97/3/1143/DC1 http://dip.doe-mbi.ucla.edu/ http://genome.c.kanazawa-u.ac.jp/Y2H C. Elegans http://cancerbiology.dfci.harvard.edu/cancerbiology/ResLabs/Vid al/ H. Pylori http://pim/hybrigenics.com Drosophila http://gifts.univ-mrs.fr/FlyNets/Flynets_home_page.html
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Yuki Juan’s Systems Biology Lab Yeast Protein Linkage Map Data New protein-protein interactions in yeast Stanley Fields Lab http://depts.washington.edu/sfields/yplm/data List of interactions with links to YPD
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Yuki Juan’s Systems Biology Lab Yeast Protein Linkage Map Data
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Yuki Juan’s Systems Biology Lab GeneScape PathwayCalling: Protein interaction and pathway Analysis http://portal.curagen.com PATHCALLING YEAST DATABASE
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Yuki Juan’s Systems Biology Lab GeneScape
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Yuki Juan’s Systems Biology Lab Outline Introduction to proteomics Protein Functions Protein-protein interactions Pathways Protein Structures and Drug Discovery Bioinformatics on the Web
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Yuki Juan’s Systems Biology Lab Biological Pathways
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Yuki Juan’s Systems Biology Lab Biological Pathways Metabolic and biochemical Transcription, regulation and protein synthesis Signal transduction
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Yuki Juan’s Systems Biology Lab Metabolic and Biochemical Pathway The synthesis of fatty acids
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Yuki Juan’s Systems Biology Lab Transcription, Regulation and Protein Synthesis Transcription Initiation Elongation Termination
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Yuki Juan’s Systems Biology Lab Initiation of Transcription
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Yuki Juan’s Systems Biology Lab The Elongation of Transcrption
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Yuki Juan’s Systems Biology Lab The Termination of Transcription
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Yuki Juan’s Systems Biology Lab Transcription, Regulation and Protein Synthesis Translation Initiation Elongation Termination
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Yuki Juan’s Systems Biology Lab 30S Subunit and Shine-Dalgarno Sequence IF: Initiating protein factor E: Exit site P: Peptide site A: Aminoacyl site
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Yuki Juan’s Systems Biology Lab Initiation of Translation in E. coli
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Yuki Juan’s Systems Biology Lab The Elongation Process in Protein Synthesis EF-Tu: Elongation Factor Peptide transferase
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Yuki Juan’s Systems Biology Lab Termination of Protein Synthesis Stop codon: UAG, UAA, UGA Cytoplasmic protein release factor
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Yuki Juan’s Systems Biology Lab Signal Transduction EDF signaling pathway Protein kinase
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Yuki Juan’s Systems Biology Lab Pathway Conservation
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Yuki Juan’s Systems Biology Lab Databases of Pathways---KEGG http://www.genome.ad.jp/kegg/
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Yuki Juan’s Systems Biology Lab Glycolysis
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Yuki Juan’s Systems Biology Lab Pathway Software BIOCARTA http://biocarta.com/ Browse all pathway
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Yuki Juan’s Systems Biology Lab Pathway Software BIOCARTA
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Yuki Juan’s Systems Biology Lab Pathway Result 1: Enolase Glycolysis Pyruvate Acetyl-CoA ethanol lactate Cancer cells BIOCARTA
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Yuki Juan’s Systems Biology Lab Pathway Result 2 : Retinoic Acid Receptor RXR-alpha BIOCARTA
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Yuki Juan’s Systems Biology Lab Development of the Network- based Pathway Paradigm Ti BS (2003), 28(5), 250-258
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Yuki Juan’s Systems Biology Lab Development of the Network- based Pathway Paradigm Ti BS (2003), 28(5), 250-258
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Yuki Juan’s Systems Biology Lab BIND http://bind.mshri.on.ca/
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Yuki Juan’s Systems Biology Lab Outline Introduction to proteomics Protein Functions Protein-protein interactions Pathways Protein Structures and Drug Discovery Bioinformatics on the Web
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Yuki Juan’s Systems Biology Lab
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Protein Structure Data Bank- PDB http://www.rcsb.org/pdb/
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Yuki Juan’s Systems Biology Lab Therapeutic Target Classes
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Yuki Juan’s Systems Biology Lab Examples of Structure-based Drug Design HIV protease with the inhibitor amprenavir (Agenerase) bound Close-up of zanamivir (Relenza) bound to influenza neuraminidase
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Yuki Juan’s Systems Biology Lab Outline Introduction to proteomics Protein Functions Protein-protein interactions Pathways Protein Structures and Drug Discovery Bioinformatics on the Web
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Yuki Juan’s Systems Biology Lab Useful Bio Websites PROSITE http://tw.expasy.org/prosite/ Amos' WWW links page http://tw.expasy.org/alinks.html Phosphoprotein Database (PPDB) http://www- lmmb.ncifcrf.gov/phosphoDB/
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Yuki Juan’s Systems Biology Lab PROSITE is a database of protein families and domains. http://tw.expasy.org/prosite/
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Yuki Juan’s Systems Biology Lab
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http://tw.expasy.org/alinks.html
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Yuki Juan’s Systems Biology Lab http://tw.expasy.org/alinks.html#PTM
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Yuki Juan’s Systems Biology Lab http://www.cbs.dtu.dk/databases/PhosphoBase/
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Yuki Juan’s Systems Biology Lab
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Proteomics Network Identify Proteins Drug Discovery Structures Protein-Protein Interactions Pathways Protein Functions
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Yuki Juan’s Systems Biology Lab Thank You!
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