Bioinformatics Resources and Tools on the Web: A Primer

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Bioinformatics Resources and Tools on the Web A
Primer

• Joel H. Graber
• Center for Advanced Biotechnology
• Boston University

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Outline

• Introduction What is bioinformatics?
• The basics
• The five sites that all biologists should know
• Some examples
• Using the tools in a somewhat less-than-naïve
  manner
• Questions/comments are welcome at all points
• Much of this material comes from the Boston
  University course BF527 Bioinformatic
  Applications (http//matrix.bu.edu/BF527/)

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What is bioinformatics?
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Examples of Bioinformatics

• Database interfaces
• Genbank/EMBL/DDBJ, Medline, SwissProt, PDB,
• Sequence alignment
• BLAST, FASTA
• Multiple sequence alignment
• Clustal, MultAlin, DiAlign
• Gene finding
• Genscan, GenomeScan, GeneMark, GRAIL
• Protein Domain analysis and identification
• pfam, BLOCKS, ProDom,
• Pattern Identification/Characterization
• Gibbs Sampler, AlignACE, MEME
• Protein Folding prediction
• PredictProtein, SwissModeler

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Things to know and remember about using web
server-based tools

• You are using someone elses computer
• You are (probably) getting a reduced set of
  options or capacity
• Servers are great for sporadic or
  proof-of-principle work, but for intensive work,
  the software should be obtained and run locally

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Five websites that all biologists should know

• NCBI (The National Center for Biotechnology
  Information
• http//www.ncbi.nlm.nih.gov/
• EBI (The European Bioinformatics Institute)
• http//www.ebi.ac.uk/
• The Canadian Bioinformatics Resource
• http//www.cbr.nrc.ca/
• SwissProt/ExPASy (Swiss Bioinformatics Resource)
• http//expasy.cbr.nrc.ca/sprot/
• PDB (The Protein Databank)
• http//www.rcsb.org/PDB/

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NCBI (http//www.ncbi.nlm.nih.gov/)

• Entrez interface to databases
• Medline/OMIM
• Genbank/Genpept/Structures
• BLAST server(s)
• Five-plus flavors of blast
• Draft Human Genome
• Much, much more

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EBI (http//www.ebi.ac.uk/)

• SRS database interface
• EMBL, SwissProt, and many more
• Many server-based tools
• ClustalW, DALI,

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SwissProt (http//expasy.cbr.nrc.ca/sprot/)

• Curation!!!
• Error rate in the information is greatly reduced
  in comparison to most other databases.
• Extensive cross-linking to other data sources
• SwissProt is the gold-standard by which other
  databases can be measured, and is the best place
  to start if you have a specific protein to
  investigate

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A few more resources to be aware of

• Human Genome Working Draft
• http//genome.ucsc.edu/
• TIGR (The Institute for Genomics Research)
• http//www.tigr.org/
• Celera
• http//www.celera.com/
• (Model) Organism specific information
• Yeast http//genome-www.stanford.edu/Saccharomyce
  s/
• Arabidopis http//www.tair.org/
• Mouse http//www.jax.org/
• Fruitfly http//www.fruitfly.org/
• Nematode http//www.wormbase.org/
• Nucleic Acids Research Database Issue
• http//nar.oupjournals.org/ (First issue every
  year)

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Example 1 Searching a new genome for a specific
protein

• Specific problem We want to find the closest
  match in C. elegans of D. melanogaster protein
  NTF1, a transcription factor
• First- understanding the different forms of blast

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The different versions of BLAST
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1st Step Search the proteins

• blastp is used to search for C. elegans proteins
  that are similar to NTF1
• Two reasonable hits are found, but the hits have
  suspicious characteristics
• besides the fact that they werent included in
  the complete genome!

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2nd Step Search the nucleotides

• tblastn is used to search for translations of C.
  elegans nucleotide that are similar to NTF1
• Now we have only one hit
• How are they related?

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Conclusion Incorrect gene prediction/annotation

• The two predicted proteins have essentially
  identical annotation
• The protein-protein alignments are disjoint and
  consecutive on the protein
• The protein-nucleotide alignment includes both
  protein-protein alignments in the proper order
• Why/how does this happen?

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Final(?) Check Gene prediction

• Genscan is the best available ab initio gene
  predictor
• http//genes.mit.edu/GENSCAN.html
• Genscans prediction spans both protein-protein
  alignments, reinforcing our conclusion of a bad
  prediction

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Ab initio vs. similarity vs. hybrid models for
gene finding

• Ab initio The gene looks like the average of
  many genes
• Genscan, GeneMark, GRAIL
• Similarity The gene looks like a specific known
  gene
• Procrustes,
• Hybrid A combination of both
• Genomescan (http//genes.mit.edu/genomescan/)

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A similar example Fruitfly homolog of mRNA
localization protein VERA

• Similar procedure as just described
• Tblastn search with BLOSUM45 produces an
  unexpected exon
• Conclusion Incomplete (as opposed to incorrect)
  annotation
• We have verified the existence of the rare
  isoform through RT-PCR

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Another example Find all genes with pdz domains

• Multiple methods are possible
• The best method will depend on many things
• How much do you know about the domain?
• Do you know the exact extent of the domain?
• How many examples do you expect to find?

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Some possible methods if the domain is a known
domain

• SwissProt
• text search capabilities
• good annotation of known domains
• crosslinks to other databases (domains)
• Databases of known domains
• BLOCKS (http//blocks.fhcrc.org/)
• Pfam (http//pfam.wustl.edu/)
• Others (ProDom, ProSite, DOMO,)

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Determination of the nature of conservation in a
domain

• For new domains, multiple alignment is your best
  option
• Global clustalw
• Local DiAlign
• Hidden Markov Model HMMER
• For known domains, this work has largely been
  done for you
• BLOCKS
• Pfam

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If you have a protein, and want to search it to
known domains

• Search/Analysis tools
• Pfam
• BLOCKS
• PredictProtein (http//cubic.bioc.columbia.edu/pre
  dictprotein/predictprotein.html)

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Different representations of conserved domains

• BLOCKS
• Gapless regions
• Often multiple blocks for one domain
• PFAM
• Statistical model, based on HMM
• Since gaps are allowed, most domains have only
  one pfam model

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Conclusions

• We have only touched small parts of the elephant
• Trial and error (intelligently) is often your
  best tool
• Keep up with the main five sites, and youll have
  a pretty good idea of what is happening and
  available