Bioinformatics for Proteomics. Ann Loraine
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1 Bioinformatics for Proteomics Ann Loraine
2 What is bioinformatics? The science of collecting, processing, organizing, storing, analyzing, and mining biological information, especially data from high-throughput biology, such as genomic sequencing or proteomics. Combines aspects of computer science, statistics, biology. Different aspects more important at different times, depending on the biological question you want to answer.
3 Bioinformatics is... The computational wing of molecular biology. Just another tool in your research repertoire. Remember: computers (and computer software programs) are designed by humans for humans. Think about how the tool is designed - be aware of the interface and how it affects what you do.
4 Useful Texts David Mount Bioinformatics Philip Bourne & Helge Wessig Structural Bioinformatics Ian Korf, et al. BLAST Carl Branden & John Tooze Introduction to Protein Structure
5 Bioinformatic data...is information based on bioinformatic analysis of experimental results, such as large sequence databases....is based on many assumptions and judgement calls along the way. Should be used with care!
6 Questions you must answer... when dealing with bioinformatic data??? What is the origin of this information? experimental? computational? What evidence supports it? What are the uncertainties and underlying assumptions?
7 Scenario Using 2D gel electrophoresis and mass spectrometry, you identify a protein that is differentially expressed in an experimental sample (human tumor) versus a control (normal tissue). MASCOT tells you that the best match is SwissProt accession P Question: What is it? What is its biological role? Accession: an id (like a social security number) for an individual record in a sequence (or other type of) database. Each sequence (protein, mrna, DNA) in a database has a unique accession.
8 Questions What is it s biochemical function? Where is it localized in the cell? What other proteins or pathways does it interact with? Others?
9 Expert Protein Analysis System Search Protein sequence databases
10 SwissProt/trEMBL SwissProt, manually-curated protein sequence database; records come from the conceptual translations of full-length cdnas, usually submitted by individual labs. records (one per protein) contain core data (sequence & references) and annotations (bioinformatic analysis results) trembl: translated DNA sequence records from EMBL (European Molecular Biology Laboratory) and GenBank (US)
11 SwissProt record useful for finding similar proteins in model organisms, where function has been studied genetically, or other family members Gene name and accession are usually the best identifiers for cross-referencing to other databases.
12 References Links to PubMed records (redundant) literature references reporting protein and nucleotide sequences for sigma High-throughput sequencing projects, usually says little about single sequences.
13 Functional Annotations nucleotide protein Protein Data Bank 3-D structure known sequences excluding ESTs Gene information Esp. useful for finding other variant forms (such as due to alternative splicing) Mendelian Inheritance in Man, molecular and disease information
14 manually-curated, expert-approved
15 OMIM gene symbol Links to multiple PubMed records
16 Gene Ontology Annotations Click for evidence, tree view Gene Ontology - a structured, controlled vocabulary describing gene products. There are main branches: biological process, molecular function, and cellular component. The GOA project is annotating human proteins with GO terms.
17 Click for tree view. evidence!
18 GO tree view
19 A mis-annotation?? click for evidence
20 Evidence for extracellular stratifin? Question: Are you convinced?
21 Protein Sequence Analysis Tools Get fasta format sequence from SwissProt record >sp P S_HUMAN protein sigma (Stratifin) MERASLIQKAKLAEQAERYEDMAAFMKGAVEKGEELSCEERNLLSVAYKNVVGGQR VLSSIEQKSNEEGSEEKGPEVREYREKVETELQGVCDTVLGLLDSHLIKEAGDAES LKMKGDYYRYLAEVATGDDKKRIIDSARSAYQEAMDISKKEMPPTNPIRLGLALNF YEIANSPEEAISLAKTTFDEAMADLHTLSEDSYKDSTLIMQLLRDNLTLWTADNAG EAPQEPQS
22 InterPro links - more clues about function Database of Interacting Proteins
23 type is family - a group of proteins that share a common evolutionary history and usually a common function
24 InterPro
25 InterPro member databases
26 Profiles Built from multiple alignments involving proteins from many species (usually). Capture probability of observing specific amino acids at specific positions. Compare a sequence to a profile to get an idea of how well the sequence fits the profile. Is it a true member of the family? If yes, this gives you clues about the protein s function. This is a form of transitive annotation. Use with caution!
27 Alignment for Note - our original protein sequence
28 Profiles used in Structure Prediction
29 Genomic or gene view Especially useful for finding alternative forms due to alternative splicing. Entrez is an interface to all of NCBI s databases and search tools
30
31 No introns! Very unusual.
32 Search the genome Get fasta format sequence from SwissProt record
33 blat results query sequence strand (always + for protein) target sequence strand (genomic)
34 A potential variant form? Links detailed annotations, proteome browser A possible variant
35
36 other players p53, estrogen, etc.
37 Proteome Page structure prediction exon structure compare to other proteins/genes
38 Web pages versus bulk download Web pages (HTTP) usually for one at a time retrieval. Bulk download (FTP) usually for fetching entire databases, large files of data needed to answer genome-scale questions. NCBI: index.html SwissProt: ftp://us.expasy.org/
39 Build your own? Programming toolkits for bioinformatics python, perl (python is easier!) a topic for another lecture THANK YOU! most tools have command-line versions
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