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Sequence comparisons
April 9, 2002
Review homework
Learning objectives-Review amino acids.
Understand difference between identity, similarity
and homology. Understand difference between
global alignment and local alignment.
Workshop-Perform sliding window to compare
two sequences
Homework #3 due on Thurs.
Amino acid characteristics
Review of amino acid
characteristics
http://info.bio.cmu.edu/Courses/BiochemMols/AAViewer/AAVFrameset.htm
http://info.bio.cmu.edu/Courses/BiochemMols/BCMolecules.html
Purpose of finding differences
and similarities of amino acids.
Infer structural information
Infer functional information
Infer evolutionary relationships
Evolutionary Basis of Sequence
Alignment
1. Similarity: Quantity that relates how much
two amino acid sequences are alike.
2. Identity: Quantity that describes how much
two sequences are alike in the strictest terms.
3. Homology: a conclusion drawn from data
suggesting that two genes share a common
evolutionary history.
Evolutionary Basis of Sequence
Alignment (Cont. 1)
1. Example: Shown on the next page is a pairwise alignment of
two proteins. One is mouse trypsin and the other is crayfish
trypsin. They are homologous proteins. The sequences
share 41% identity.
2. Underlined residues are identical. Asterisks and diamond
represent those residues that participate in catalysis. Five
gaps are placed to optimize the alignment.
Evolutionary Basis of Sequence
Alignment (Cont. 2)
Why are there regions of identity?
1) Conserved function-residues participate in reaction.
2) Structural (For example, conserved cysteine residues that
form a disulfide linkage)
3) Historical-Residues that are conserved solely due to a
common ancestor gene.
Evolutionary Basis of Sequence
Alignment (Cont. 3)
Note: it is possible that two proteins share a high degree of
similarity but have two different functions. For example,
human gamma-crystallin is a lens protein that has no known
enzymatic activity. It shares a high percentage of identity with
E. coli quinone oxidoreductase. These proteins likely had a
common ancestor but their functions diverged.
Analogous to railroad car and diner function.
Modular nature of proteins
The previous alignment was global. However,
many proteins do not display global patterns of
similarity. Instead, they possess local regions of
similarity.
Proteins can be thought of as assemblies of
modular domains. It is thought that this may, in
some cases, be due to a process known as exon
shuffling.
Modular nature of proteins (cont. 1)
Gene A
Exon 1a
Exon 2a
Duplication of Exon 2a
Gene A
Exon 1a
Exon 2a
Exon 2a
Exchange with Gene B
Gene B
Exon 1b
Exon 2b
Exon 2b
Gene A
Exon 1a
Exon 2a
Exon 3 (Exon 2b from Gene B)
Gene B
Exon 1b
Exon 2b
Exon 3 (Exon 2a from Gene A)
Dot Plots
A
A
T
G
C
C
T
A
G
T
G
C C
*
T
*
*
A
G
*
*
*
* *
* *
*
*
*
*
*
*
Window = 1
Note that 25% of
the table will be
filled due to random
chance. 1 in 4 chance
at each position
Dot Plots with window = 2
A
A
{
T
{
G
{
C
{
C
{
T
{A
{G
T
G
C C
T
A
*
*
*
G
Window = 2
The larger the window
the more noise can
be filtered
What is the
percent chance that
you will receive a
match randomly?
1/16 * 100 = 6.25%
*
*
*
*
Similarity
It is easy to score if an amino acid is identical to another (the
score is 1 if identical and 0 if not). However, it is not easy to
give a score for amino acids that are somewhat similar.
+NH
3
CO2-
+NH
3
CO2-
Isoleucine
Leucine
Should they get a 0 (non-identical) or a 1 (identical) or
Something in between?
Identity Matrix
A
C
I
L
1
0
0
0
A
1
0 1
0 0
C I
1
L
Simplest type of scoring matrix
The Point-Accepted-Mutation (PAM) model
of evolution and the PAM scoring matrix
It implies that each amino acid (AA) mutates independently of
each other with a probability which depends only on the AA.
Since there are 20 AA, the transition probabilities are
described by a 20X20-mutation matrix, denoted by M.
A standard M, which defines a 1-PAM change.
Point Accepted Mutation (PAM) Distance: A 1-PAM unit changes 1%
of the amino acids on average:
where fi is the frequency of AA i. One PAM is a unit of evolutionary
divergence in which 1% of the amino acids have been changed.
The Point-Accepted-Mutation (PAM) model
of evolution and the PAM scoring matrix
(cont. 1)
A 2-PAM unit is equivalent to two 1-PAM unit evolution
(or M2).
A k-PAM unit is equivalent to k 1-PAM unit evolution
(or Mk). Example 1:
CNGTTDQVDKIVKILNEGQIASTDVVEVVVSPPYVFLPVVKSQLRPEIQV
|||||||||||||| |||||||||||||||||||||||||||||||||||
CNGTTDQVDKIVKIRNEGQIASTDVVEVVVSPPYVFLPVVKSQLRPEIQV
lengths = 50
1 Mismatch
PAM distance = 2
Two proteins that are similar in
certain regions
Tissue plasminogen activator (PLAT)
Coagulation factor 12 (F12).
The Dotter Program
• Program consists of three components:
•Sliding window
•A table that gives a score for each amino acid match
•A graph that converts the score to a dot of certain density.
The higher the density the higher the score.
Region of
similarity
Single region on F12
is similar to two regions
on PLAT