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Construction of Substitution matrices BLOSUM BLO CKS SU BSTITUTION M ATRIX PAM

Construction of Substitution matrices BLOSUM BLO CKS SU BSTITUTION M ATRIX PAM P OINT A CCEPTED M UTATIONS. Substitution matrices

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Construction of Substitution matrices BLOSUM BLO CKS SU BSTITUTION M ATRIX PAM

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  1. Construction of Substitution matrices • BLOSUM • BLOCKS SUBSTITUTION MATRIX • PAM • POINT ACCEPTED MUTATIONS

  2. Substitution matrices • Substitution matrix contains values proportional to the probability that amino acid A mutates into amino acid B for all pairs of amino acids through a period of evolution • Substitution matrices are constructed from a large and diverse sample of sequence alignments

  3. How to construct substitution matrices • Multiple alignment of well studies gene sequences from different species • use orthologs: functionally similar • observed substitutions tend to preserve functions • minimal gaps

  4. How to construct substitution matrices ? • Tabulate substitutions • A to A: 9867 times • A to R: 2 times • A to N: 9 times • etc….

  5. How to construct substitution matrices ?

  6. Construction of Substitution matrices • BLOSUM

  7. Construction of Substitution matrices • BLOSUM

  8. How to construct substitution matrices ? Substitution matrix score = Log Observed mutation rate in alignment Expected random mutation rate

  9. How do we find the random mutation rate?

  10. The random mutation rate • compute the overall occurrence of an amino acid in a protein database

  11. The random mutation rate • compute the overall occurrence of an amino acid in a protein database http://www.ebi.ac.uk/swissprot/sptr_stats/index.html

  12. The random mutation rate • Example: • Expected random mutation rate is 1 in 10000 and observed mutation rate of W to R is 1 in 10 • Score = log (0.1/0.0001) = log (1000) = +3

  13. Calculating BLOSUM62 scores

  14. Calculating BLOSUM62 scores

  15. Calculating BLOSUM62 scores

  16. Calculating BLOSUM62 scores

  17. Calculating BLOSUM62 scores

  18. Calculating BLOSUM62 scores

  19. Calculating BLOSUM62 scores

  20. Calculating BLOSUM62 scores

  21. Calculating BLOSUM62 scores

  22. Calculating BLOSUM62 scores

  23. PAM matrices • Point Accepted Mutations • [1 point mutation per 100 amino acids] • does not take into account different evolutionary rates between conserved and non-conserved regions

  24. PAM1 is 1% average change in amino acids • PAM 250:??

  25. Why use substitution matrices?????

  26. Why use substitution matrices? • Database searches

  27. Database searching

  28. Database searching

  29. Database searching • Query Sequence; Database sequences

  30. Database searching: Filtering • Dynamic programming is computationally expensive • Apply DP to sequence pairs that are likely to be similar • find short words: query-database • DNA 7-28bases (BLAST?) • PROTEIN 3 amino acids (BLAST?)

  31. BLAST • Basic Local Alignment Search Tool • Heuristic method?

  32. Blast output parameter E value

  33. E value • number of alignments one can expect see by chance. • Number of alignments having the same or greater score. • Dependent on size of database and length of query seq.

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