Sequence Analysis

Introduction to
Alignment
Approaches
Pairwise Sequence Alignment - I

Alignment
• The process of
inexact matching
while keeping in
view the conserved
residues is called
sequence
“Alignment”
Introduction to Alignment Approaches

Background
• Sequences vary due
to insertions,
deletions and
substitutions

• To calculate the
similarity between
two sequences, we
need to align the
sequences
Introduction to Alignment Approaches

Introduction

• What are the

approaches to
aligning sequences

• Two approaches are:

• Global Alignment
• Local Alignment
Introduction to Alignment Approaches

Local Alignment
“Match one sequence with a portion
of the second sequence”
 Why Local Alignments?

Other Advantages
To calculate the
similarity between • Different length
two sequences, sequences can be
compared
we need to align
the sequences • Conserved domains
can be elicited from
proteins belonging
Local alignments have to different families
the power to detect • Common functional
small regions of high features can thus be
similarity between two identified
sequences
Introduction to Alignment Approaches

Global Alignment
“An end to end match of one sequence with the
second sequence”
 Pairwise Sequence Alignment - II

Types of pairwise alignments

Global Local
• Maximizing sequence • Find regions between
matches over the entire two sequences that have
length of two sequences, the strongest similarity,
by introducing gaps. excluding less similar
regions.

• Used to determine • Finding similar domains,

overall similarity, motifs, detecting distant
conservation. homology
Introduction to Alignment Approaches

• Local alignment
focuses on the
highly matching sub
regions within the
sequences

• Global alignment
creates an end-to-
end matching
between two
sequences

END
Pairwise Sequence Alignment -
Background
Pairwise sequence alignment is therefore
alignment of a pair of two sequences

• Given a pair of sequences (strings of characters)

• Pairwise alignment tells the similarity between the

sequences by maximizing the matches
 Pairwise Sequence Alignment - III

• Sequences can differ from each other in

three different ways:

• Substitutions ACGA  AGGA

• Insertions ACGA  ACCGA
• DeletionsACGA  AGA

• No change? match
• Insertions or deletions ("indels") result in
gaps in alignments
• Substitutions result in mismatches
 Pairwise Sequence Alignment - III

Optimal Alignment
Align 1 & 2
1: THIS IS A RATHER LONGER SENTENCE THAN THE NEXT.
2: THIS IS A SHORT SENTENCE.

1: THIS IS A RATHER LONGER SENTENCE THAN THE NEXT.

2: THIS IS A ...........SHORT…...SENTENCE……………………
OR

1: THIS IS A RATHER LONGER SENTENCE THAN THE NEXT.

2: THIS IS A ….SHORT…..SENT..EN……CE…………………….

Which alignment is optimal in biological context?

Pairwise Sequence Alignment - III

Conclusion
• Global and local
pairwise alignments
are two types of
alignment and they
can lead to different
results

• Indels lead to Gaps

in alignments

• Substitutions lead to
mismatches

END
Sequence Analysis

Multiple
Sequence
Alignment
Multiple Sequence Alignment

Introduction

• Multiple sequence
alignment involves
comparison of three
of more sequences

• Pairwise alignment
was either local or
global

• Multiple alignment is
mostly global
 Multiple Sequence Alignment

A block of multiply aligned sequences

MQVKLFTPLHDKSDHGKYH
MQVKIFTPLHDKS-HGKSH
MQVHLY-PLHDKS-TGKSH
MQVHLF-PLHDKSDTGKSH
MQVKLYTPLHDKSDHGKYH
Multiple Sequence Alignment

MSA vs Pairwise
• For pairwise
alignments, we used
Dynamic
Programming

• For MSA, Dynamic

Programming may
get very expensive,
computationally.

• Solution:
Progressive
Alignment
 Multiple Sequence Alignment

Conclusion
• MSA can help align
multiple sequences
• Dynamic
Programming
cannot be used as it
becomes expensive
• Progressive
alignment can help
perform MSA!
• CLUSTAL – Online
tool

END
More on Multiple Sequence Alignment

Application of
MSA

• Predict secondary
and tertiary
structures of new
protein sequences

• Evaluate
evolutionary order
of species or
“Phylogeny”
More on Multiple Sequence Alignment

Methodology

• Pairwise alignment
is the alignment of
two sequences

• MSA can be
performed by
repeated application
of pairwise
alignment
 More on Multiple Sequence Alignment

Find and list all

Compare all sequence in
sequences to be
a pairwise manner
compared

Perform clustering of
Create a hierarchy using
pairwise aligned
binary trees
sequences

Alignment of two best Process is repeated for

aligned sequences is next pair of best aligned
created sequences
More on Multiple Sequence Alignment
More on Multiple Sequence Alignment

Conclusion
• MSA can help align
multiple sequences
• Progressive
alignment can help
perform MSA!
• Need to remove
sequences with
>80% similarity

• CLUSTAL – Online
tool

END
Sequence Analysis

MSA - Example
MSA - Example

Background
• MSA involves
progressive
alignment of
sequences
• Doing so many
progressive
alignments can be
slow
• Let’s take a look at
an example
 MSA - Example

MSA on globin sequences

Hbb_human

Glb5_petma
Hba_human

Myg_phyca

Lgb2_lupla
Hbb_horse

Hba_horse
Hbb_human 1 -
Hbb_horse 2 .17 -
Hba_human 3 .59 .60 -
Hba_horse 4 .59 .59 .13 -
Myg_phyca 5 .77 .77 .75 .75 -
Glb5_petma 6 .81 .82 .73 .74 .80 -
Lgb2_lupla 7 .87 .86 .86 .88 .93 .90 -

Thompson et al, IGBMC

 MSA - Example

Progressive alignment using sequential branching

Hba_human
1
Hba_horse
2
Hbb_horse 3
Hbb_human 4
Glb5_petma 5
Myg_phyca 6
Lgb2_lupla

Thompson et al, IGBMC

 MSA - Example

Progressive alignment following a guide tree

.081
Hbb_human
.226
2.084
.061 Hbb_horse
3 .055
Hba_human
.015
4
.219
1.065
Hba_horse
.062 .398
5 Myg_phyca
6 .389
Glb5_petma
.442
Lgb2_lupla

Thompson et al, IGBMC

 MSA - Example

Alignment results

Thompson et al, IGBMC

 MSA - Example

Conclusion
• MSA can be better
performed using
clustering strategies
followed by
alignment of the
alignments later

• CLUSTAL is a free
online tool that does
all of this for us!

END