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Sequence analysis course

Sequence analysis course. Lecture 6 Multiple sequence alignment 2 of 3 Multiple alignment methods. Biological definitions for related sequences.

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Sequence analysis course

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  1. Sequence analysis course Lecture 6 Multiple sequence alignment 2 of 3 Multiple alignment methods Sequence analysis 2005 - lecture 6

  2. Biological definitions for related sequences • Homologues are similar sequences in two different organisms that have been derived from a common ancestor sequence. Homologues can be described as either orthologues or paralogues. • Orthologues are similar sequences in two different organisms that have arisen due to a speciation event. Orthologs typically retain identical or similar functionality throughout evolution. • Paralogues are similar sequences within a single organism that have arisen due to a gene duplication event. • Xenologues are similar sequences that do not share the same evolutionary origin, but rather have arisen out of horizontal transfer events through symbiosis, viruses, etc. Sequence analysis 2005 - lecture 6

  3. So this means … Source: http://www.ncbi.nlm.nih.gov/Education/BLASTinfo/Orthology.html Sequence analysis 2005 - lecture 6

  4. Multiple sequence alignment • Sequences can be conserved across species and perform similar or identical functions.> hold information about which regions have high mutation rates over evolutionary time and which are evolutionarily conserved;> identification of regions or domains that are critical to functionality. • Sequences can be mutated or rearranged to perform an altered function.> which changes in the sequences have caused a change in the functionality. Multiple sequence alignment: the idea is to take three or more sequences and align them so that the greatest number of similar characters are aligned in the same column of the alignment. Sequence analysis 2005 - lecture 6

  5. What to ask yourself • How do we get a multiple alignment?(three or more sequences) • What is our aim?– Do we go for max accuracy, least computational time or the best compromise? • What do we want to achieve each time Sequence analysis 2005 - lecture 6

  6. Sequence-sequence alignment sequence sequence Sequence analysis 2005 - lecture 6

  7. Multiple alignment methods • Multi-dimensional dynamic programming> extension of pairwise sequence alignment. • Progressive alignment> incorporates phylogenetic information to guide the alignment process • Iterative alignment> correct for problems with progressive alignment by repeatedly realigning subgroups of sequence Sequence analysis 2005 - lecture 6

  8. Simultaneous multiple alignmentMulti-dimensional dynamic programming The combinatorial explosion • 2 sequences of length n • n2 comparisons • Comparison number increases exponentially • i.e. nN where n is the length of the sequences, and N is the number of sequences • Impractical for even a small number of short sequences Sequence analysis 2005 - lecture 6

  9. Multi-dimensional dynamic programming(Murata et al., 1985) Sequence 1 Sequence 3 Sequence 2 Sequence analysis 2005 - lecture 6

  10. The MSA approach • MSA (Lipman et al., 1989, PNAS 86, 4412) • MSA restricts the amount of memory by computing bounds that approximate the centre of a multi-dimensional hypercube. • Calculate all pair-wise alignment scores. • Use the scores to to predict a tree. • Calculate pair weights based on the tree (lower bound). • Produce a heuristic alignment based on the tree. • Calculate the maximum weight for each sequence pair (upper bound). • Determine the spatial positionsthat must be calculated to obtain the optimal alignment. • Perform the optimal alignment. • Report the weight found comparedto the maximum weight previouslyfound (measure of divergence). • Extremely slow and memory intensive. • Max 8-9 sequences of ~250 residues. Sequence analysis 2005 - lecture 6

  11. The DCA approach • DCA (Stoye et al., 1997, Appl. Math. Lett. 10(2), 67-73) • Each sequence is cut in two behinda suitable cut position somewhere close to its midpoint. • This way, the problem of aligningone family of (long) sequences is divided into the two problems of aligning two families of (shorter) sequences. • This procedure is re-iterated untilthe sequences are sufficiently short. • Optimal alignment by MSA. • Finally, the resulting short alignments are concatenated. Sequence analysis 2005 - lecture 6

  12. So in effect … Sequence 1 Sequence 3 Sequence 2 Sequence analysis 2005 - lecture 6

  13. Multiple alignment methods • Multi-dimensional dynamic programming> extension of pairwise sequence alignment. • Progressive alignment> incorporates phylogenetic information to guide the alignment process • Iterative alignment> correct for problems with progressive alignment by repeatedly realigning subgroups of sequence Sequence analysis 2005 - lecture 6

  14. The progressive alignment method • Underlying idea: usually we are interested in aligning families of sequences that are evolutionary related. • Principle: construct an approximate phylogenetic tree for the sequences to be aligned and than to build up the alignment by progressively adding sequences in the order specified by the tree. • But before going into details, some notices of multiple alignment profiles … Sequence analysis 2005 - lecture 6

  15. How to represent a block of sequences? • Historically: consensus sequence – single sequence that best represents the amino acids observed at each alignment position. • Modern methods: Alignment profile – representation that retains the information about frequencies of amino acids observed at each alignment position. Sequence analysis 2005 - lecture 6

  16. Multiple alignment profiles (Gribskov et al. 1987) • Gribskov created a probe: group of typical sequences of functionally related proteins that have been aligned by similarity in sequence or three-dimensional structure (in his case: globins & immunoglobulins). • Then he constructed a profile, which consists of a sequence position-specific scoring matrix M(p,a) composed of 21 columns and N rows (N = length of probe). • The first 20 columns of each row specify the score for finding, at that position in the target, each of the 20 amino acid residues. An additional column contains a penalty for insertions or deletions at that position (gap-opening and gap-extension). Sequence analysis 2005 - lecture 6

  17. Multiple alignment profiles Core region Gapped region Core region i A C D    W Y fA.. fC.. fD..    fW.. fY.. fA.. fC.. fD..    fW.. fY.. fA.. fC.. fD..    fW.. fY.. - Gapo, gapx Gapo, gapx Gapo, gapx Position dependent gap penalties Sequence analysis 2005 - lecture 6

  18. Profile building • Example: each aa is represented as a frequency penalties as weights. i A C D    W Y 0.5 0 0    0 0.5 0.3 0.1 0    0.3 0.3 0 0.5 0.2    0.1 0.2 Gap penalties 1.0 0.5 1.0 Position dependent gap penalties

  19. Profile-sequence alignment sequence ACD……VWY

  20. Sequence to profile alignment A A V V L 0.4 A 0.2 L 0.4 V Score of amino acid L in sequence that is aligned against this profile position: Score = 0.4 * s(L, A) + 0.2 * s(L, L) + 0.4 * s(L, V)

  21. Profile-profile alignment profile A C D . . Y profile ACD……VWY

  22. Profile to profile alignment 0.75 G 0.25 S 0.4 A 0.2 L 0.4 V Match score of these two alignment columns using the a.a frequencies at the corresponding profile positions: Score = 0.4*0.75*s(A,G) + 0.2*0.75*s(L,G) + 0.4*0.75*s(V,G) + + 0.4*0.25*s(A,S) + 0.2*0.25*s(L,S) + 0.4*0.25*s(V,S) s(x,y) is value in amino acid exchange matrix (e.g. PAM250, Blosum62) for amino acid pair (x,y)

  23. So, for scoring profiles … • Think of sequence-sequence alignment. • Same principles but more information for each position. Reminder: • The sequence pair alignment score S comes from the sum of the positional scores M(aai,aaj) (i.e. the substitution matrix values at each alignment position minus penalties if applicable) • Profile alignment scores are exactly the same, but the positional scores are more complex Sequence analysis 2005 - lecture 6

  24. Scoring a profile position Profile 1 Profile 2 A C D . . Y A C D . . Y • At each position (column) we have different residue frequencies for each amino acid (rows) SO: • Instead of saying S=M(aa1, aa2) (one residue pair) • For frequency f>0 (amino acid is actually there)we take: Sequence analysis 2005 - lecture 6

  25. Log-average score • Remember the substitution matrix formula? • In log-average scoring (von Ohsen et al, 2003) • Why is this so important? Think about it… Sequence analysis 2005 - lecture 6

  26. Progressive alignment • Perform pair-wise alignments of all of the sequences; • Use the alignment scores to produces a dendrogram using neighbour-joining methods (guide-tree); • Align the sequences sequentially, guided by the relationships indicated by the tree. • Biopat (first method ever) • MULTAL (Taylor 1987) • DIALIGN (1&2, Morgenstern 1996) • PRRP (Gotoh 1996) • ClustalW (Thompson et al 1994) • PRALINE (Heringa 1999) • T Coffee (Notredame 2000) • POA (Lee 2002) • MUSCLE (Edgar 2004) Sequence analysis 2005 - lecture 6

  27. Progressive multiple alignment 1 Score 1-2 2 1 Score 1-3 3 4 Score 4-5 5 Scores Similarity matrix 5×5 Scores to distances Iteration possibilities Guide tree Multiple alignment

  28. General progressive multiple alignment technique(follow generated tree) d 1 3 1 3 2 5 1 3 2 5 1 root 3 2 5

  29. PRALINE progressive strategy d 1 3 1 3 2 1 3 2 5 4 1 3 2 5 4

  30. There are problems … Accuracy is very important !!!! • Alignment errors during the construction of the MSA cannot be repaired anymore: propagated into the progressive steps. • The comparisons of sequences at early steps during progressive alignments cannot make use of information from other sequences. • It is only later during the alignment progression that more information from other sequences (e.g. through profile representation) becomes employed in the alignment steps. “Once a gap, always a gap” Feng & Doolittle, 1987

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