PsiBLAST, Prosite, UCSC Genome Browser Lecture 3

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Psi-BLAST,Prosite, UCSC Genome
BrowserLecture 3
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Searching for remote homologs

• Sometimes BLAST isnt enough
• Large protein family, and BLAST only finds close
  members. We want more distant members

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PSI-BLAST

• Position Specific Iterative BLAST

Regular blast
Construct profile from blast results
Blast profile search
Final results
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Consensus, Pattern, PSSM
Consensus the most frequent character in the
column is chosen
Pattern represents the alignment as a regular
expression
Profile PSSM Position Specific Score Matrix
Pos
Nuc
A A C T T G
A-TA-C-T-T-GC
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Pos
Nuc
S(AACCAA)10.6711.25.33 S(GACCAA)0 Sequences
with higher scores -gt higher chance of being
related to the PSSM
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PSI-BLAST

• Position Specific Iterative BLAST

Regular blast
Construct profile from blast results
Blast profile search
Final results
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BLAST PSI-Blast
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PSI-Blast - results
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PSI-BLAST

• Advantage PSI-BLAST looks for seqs that are
  close to the query, and learns from them to
  extend the circle of friends
• Disadvantage if we obtained a WRONG hit, we will
  get to unrelated sequences (contamination). This
  gets worse and worse each iteration

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PSI-BLAST

• Which of the following is/are correct?
• PSI-BLAST is expected to give more hits than
  BLAST
• PSI-BLAST is an iterative search method
• PSI-BLAST is faster than BLAST
• Each iteration of PSI-BLAST can only improve the
  results of the previous iteration

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Turning information into knowledge

• The outcome of a sequencing project are masses of
  raw data
• The challenge is to turn these raw data into
  biological knowledge
• A valuable tool for this challenge is an
  automated diagnostic pipe through which newly
  determined sequences can be streamlined

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From sequence to function

• Nature tends to innovate rather than invent
• Proteins are composed of functional elements
  domains and motifs
• Domains are structural
• units that carry out a
• certain function. They are
• shared between different
• proteins
• Motifs are shorter
• and are usually critical
• for the biological activity

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http//www.expasy.ch/prosite
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Prosite

• From analyzing conserved regions in protein
  sequences it is possible to derive signatures of
  motifs and domains
• Prosite consists of annotated sites/motifs/signatu
  res/fingerprints
• Given an uncharacterized translated protein
  sequence, prosite tries to predict which motifs
  and domains make up the protein and thus identify
  the family to which it belongs

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Prosite

• Prosite represents entries with patterns or
  profiles

profile
pattern
A-TA-C-T-T-GC

• Profiles are used in prosite when the motif is
  relatively divergent, and is difficult to
  represent as a pattern
• Profiles also characterize domains over their
  entire length, not just the motif

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Prosite sequence query
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Patterns with a high probability of occurrence

• Entries describing commonly found
  post-translational modifications or
  compositionally biased regions
• Found in the majority of known protein sequences
• High probability of occurrence
• Prosite filters them by default

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Scanning Prosite
Query sequence
Query pattern
Result all patterns found in the sequence
Result all sequences which adhere to this pattern
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Prosite pattern query
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UCSC Genome Browser
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UCSC Genome Browser - Gateway
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UCSC Genome Browser - Gateway
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Results
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Annotation tracks
Base position
UTR
UCSC Genes
RefSeq Genes
mRNAs (GenBank)
Intron
Coding
GeneDirection
SNPs
Repeats
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UCSC Gene
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UCSC Genome Browser - movement
Zoom x3 Center
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Controllingannotationtracks
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BLAT

• BLAT Blast-Like Alignment Tool
• BLAT is designed to find similarity of gt95 on
  DNA, gt80 for protein
• Rapid search by indexing entire genome
• Good for
• Finding genomic coordinates of cDNA
• Determining exons/introns
• Finding human (or chimp, dog, cow) homologs of
  another vertebrate sequence

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BLAT on UCSC Genome Browser
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BLAT search
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BLAT Results
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BLAT Results
query
Match
Non-Match(mismatch/indel)
hit
Indel boundaries