1. Construction of Substitution Matrices
BLOSUM: BLOcks SUbstitution Matrix
PAM: Point Accepted Mutations

2. Substitution Matrices
Contain 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

3. Substitution Matrices
Contain 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
Are constructed from a large and diverse sample of sequence alignments

4. Substitution Matrices
Contain 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
Are constructed from a large and diverse sample of sequence alignments
Multiple alignment of well studied gene sequences from different species

5. Substitution Matrices
Contain 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
Are constructed from a large and diverse sample of sequence alignments
Multiple alignment of well studied gene sequences from different species
Use orthologs - functionally similar

6. Substitution Matrices
Contain 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
Are constructed from a large and diverse sample of sequence alignments
Multiple alignment of well studied gene sequences from different species
Use orthologs - functionally similar
Observed substitutions tend to preserve functions

7. Substitution Matrices
Contain 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
Are constructed from a large and diverse sample of sequence alignments
Multiple alignment of well studied gene sequences from different species
Use orthologs - functionally similar
Observed substitutions tend to preserve functions
Minimal gaps

8. How to Construct Substitution Matrices
Tabulate substitutions
A to A: 9867 times
A to R: 2 times
A to N: 9 times
etc….

9. How to Construct Substitution Matrices

10. How to Construct Substitution Matrices (BLOSUM)

11. How to Construct Substitution Matrices (BLOSUM)

12. How to Construct Substitution Matrices

13. Finding the Random Mutation Rate
Compute overall occurrence of an amino acid in a protein database

14. Finding the Random Mutation Rate
Compute overall occurrence of an amino acid in a protein database

15. Finding the Random Mutation Rate
Compute overall occurrence of an amino acid in a protein database

16. Finding the Random Mutation Rate
Example:
Expected random mutation rate is 1 in and observed mutation rate of W to R is 1 in 10
Score = log (0.1/0.0001) = log (1000) = +3

21. PAM Matrices [1 point mutation per 100 amino acids]
does not take into account different evolutionary rates between conserved and non-conserved regions
PAM1 is 1% average change in amino acids
PAM 250:??

22. PAM Matrices

23. PAM vs. BLOSUM

24. Basic Local Alignment Search Tool (BLAST)
Heuristic method

25. BLAST Algorithm

26. BLAST Algorithm

27. BLAST Algorithm

30. What can we search and compare?
DNA vs DNA
Protein vs Protein
DNA vs Protein
Protein vs DNA

31. Reading Frames

32. The best BLAST program