Difference between revisions of "Sequence Alignments"

From Bioinformatikpedia
(→‎T-Coffee)
(→‎References)
 
(38 intermediate revisions by 2 users not shown)
Line 13: Line 13:
 
* HHSearch
  
* HHSearch
 
hhsearch -i query -d database -o output.txt
 
hhsearch -i query -d database -o output.txt
−  
   
 
database = /data/blast/nr/nr
  
database = /data/blast/nr/nr
  +
  +
  +
'''Result Statistics'''
  +
  +
* '''Overlap'''
  +
To illustrate the overlap of the returned sequences for the sequences searches, Venn Diagrams were drawn.
  +
  +
[[Image:comparePSI3.png|thumb|right|Comparisons of the results for the PSIBLAST runs with each 3 iteraions. (PSI1 = PSI-BLAST run with 3 iterations, E-value cutoff 0.005, PSI3 = PSI-BLAST run with 3 iterations, E-value cutoff 10E6)]]
  +
[[Image:comparePSI5.png|thumb|right|Comparisons of the results for the PSIBLAST runs with each 5 iteraions. (PSI2 = PSI-BLAST run with 5 iterations, E-value cutoff 0.005, PSI4 = PSI-BLAST run with 5 iterations, E-value cutoff 10E6)]]
  +
The PSI-BLAST runs with each 3 iterations returned absolutely the same sequences match our query BCKDHA. The same is true for the PSI-BLAST runs with each 5 iterations. This fact was used to combine their results for the following Venn Diagramm (created with [http://bioinfogp.cnb.csic.es/tools/venny/index.html]):
  +
[[Image:vennDiagram.png|none|300px]]
  +
  +
Including BLAST and FASTA, the most interesting fact is that FASTA found more than 2600 more results, that were not returned by any of the other search algorithms. This may be due to no restriction concerning the E-value using the default FASTA search.
  +
BLAST returned only one additional Hit, that was not found in any of the PSI-BLAST searches, but which is also included in the FASTA results. Any PSI-BLAST results were also detected by FASTA. Using PSIBLAST with 5 iterations, 6 more Hits were returned than using PSIBLAST with only 3 iterations. This may be due to a longer search time.
  +
All in all one could say, that the Search algorithms returned nearly the same sequences matching our query.
  +
  +
  +
* '''Identity Distribution'''
  +
[[Image:IdentityDistribution.png|none|500px|Identity Distribution for the results from the Sequence Searches]]
  +
As the PSIBlast runs with each 3 iterations resulted in the same Hits, as well as the PSI-BLAST runs with 5 iterations, the identity distributions for those runs were pooled using the same colour.
  +
Remarkable is the identity distribution for the FASTA run, which returned a lot more hits with little identity, that the other runs. All in all, FASTA returned almost 3000 Hits (default parameter search), while all the BLAST/PSIBLAST runs returned not more than 300 Hits each. Therefore a lot of the FASTA 'Hits' have little identity and a quite low E-value (see below), but still FASTA returned some good results.
  +
  +
  +
* '''Evalue Distribution'''
  +
[[Image:EvalueDistribution.png|none|500px| E-value Distribution for the results from the Sequence Searches]]
  +
The E-value distribution for the sequence searches is quite similar concerining the BLAST and PSI-BLAST results. Only FASTA has a lot wider E-value range, which can be explained by the fact, that FASTA returned about 10 times more Hits, among which a lot of sequences have little identity and therefore a quite high E-value.
  +
  +
  +
   
 
Sequences chosen for the multiple Alignment:
  
Sequences chosen for the multiple Alignment:
Line 94: Line 122:
   
 
./cobalt -i sequences.fasta -norps T > output.aln
  
./cobalt -i sequences.fasta -norps T > output.aln
  +
  +
   
 
=== Conservation and Gaps ===
  
=== Conservation and Gaps ===
Line 130: Line 160:
 
|71
  
|71
 
|91
  
|91
−
|
  
 
|-
  
|-
 
|T-Coffee (3D)
  
|T-Coffee (3D)
Line 162: Line 191:
 
|10
  
|10
 
|}
  
|}
  +
   
 
=== Gaps in secondary structure ===
  
=== Gaps in secondary structure ===
Line 239: Line 269:
 
==== T-Coffee 3d ====
  
==== T-Coffee 3d ====
   
−
[[Image:t-coffee_3d_gaps_structure.png|thumb|right|T-Coffee (3D) gaps and structure]]
 +
[[Image:t-coffee_3d_gaps_structure.png|thumb|right|T-Coffee (3D) gaps and structure [http://www.pdb.org/pdb/explore/remediatedSequence.do?structureId=1U5B]]]
   
 
{| border="1" style="text-align:center; border-spacing:0;"
  
{| border="1" style="text-align:center; border-spacing:0;"
Line 345: Line 375:
 
==== Cobalt ====
  
==== Cobalt ====
   
−
[[Image:cobalt_gaps_structure.png|thumb|right|Cobalt gaps and structure]]
 +
[[Image:cobalt_gaps_structure.png|thumb|right|Cobalt gaps and structure [http://www.pdb.org/pdb/explore/remediatedSequence.do?structureId=1U5B]]]
   
 
{| border="1" style="text-align:center; border-spacing:0;"
  
{| border="1" style="text-align:center; border-spacing:0;"
Line 395: Line 425:
 
==== Muscle ====
  
==== Muscle ====
   
−
[[Image:muscle_gaps_structure.png|thumb|right|Muscle gaps and structure]]
 +
[[Image:muscle_gaps_structure.png|thumb|right|Muscle gaps and structure [http://www.pdb.org/pdb/explore/remediatedSequence.do?structureId=1U5B]]]
   
 
{| border="1" style="text-align:center; border-spacing:0;"
  
{| border="1" style="text-align:center; border-spacing:0;"
Line 431: Line 461:
 
|}
  
|}
   
  +
=== Functionally important residues ===
  +
The functionally important sites are according to [http://www.uniprot.org/uniprot/P12694] the following sites:
  +
* Metal binding site, 206
  +
* Metal binding site, 211
  +
* Metal binding site, 212
  +
  +
=== References ===
  +
  +
[http://www.uniprot.org/uniprot/P12694 Secondary structure information]
  +
  +
back to [[Reference_Sequence_BCKDHA|Reference Sequence of BCKDHA]]
 
back to [[Maple syrup urine disease]] main page
  
back to [[Maple syrup urine disease]] main page

Latest revision as of 11:13, 21 May 2011

Sequence Alignments

Sequence searches

  • FASTA

../bin/fasta36 sequence.fasta database > FastaOutput.txt

  • BLAST

blastall -p blastp -d database -i sequence.fasta > BlastOutput.txt

  • PSIBLAST

blastpgp -i sequence.fasta -j iterations -h evalueCutoff -d database > PsiblastOutput.txt

  • HHSearch

hhsearch -i query -d database -o output.txt

database = /data/blast/nr/nr


Result Statistics

  • Overlap

To illustrate the overlap of the returned sequences for the sequences searches, Venn Diagrams were drawn.

Comparisons of the results for the PSIBLAST runs with each 3 iteraions. (PSI1 = PSI-BLAST run with 3 iterations, E-value cutoff 0.005, PSI3 = PSI-BLAST run with 3 iterations, E-value cutoff 10E6)
Comparisons of the results for the PSIBLAST runs with each 5 iteraions. (PSI2 = PSI-BLAST run with 5 iterations, E-value cutoff 0.005, PSI4 = PSI-BLAST run with 5 iterations, E-value cutoff 10E6)

The PSI-BLAST runs with each 3 iterations returned absolutely the same sequences match our query BCKDHA. The same is true for the PSI-BLAST runs with each 5 iterations. This fact was used to combine their results for the following Venn Diagramm (created with [6]):

VennDiagram.png

Including BLAST and FASTA, the most interesting fact is that FASTA found more than 2600 more results, that were not returned by any of the other search algorithms. This may be due to no restriction concerning the E-value using the default FASTA search. BLAST returned only one additional Hit, that was not found in any of the PSI-BLAST searches, but which is also included in the FASTA results. Any PSI-BLAST results were also detected by FASTA. Using PSIBLAST with 5 iterations, 6 more Hits were returned than using PSIBLAST with only 3 iterations. This may be due to a longer search time. All in all one could say, that the Search algorithms returned nearly the same sequences matching our query.


  • Identity Distribution
Identity Distribution for the results from the Sequence Searches

As the PSIBlast runs with each 3 iterations resulted in the same Hits, as well as the PSI-BLAST runs with 5 iterations, the identity distributions for those runs were pooled using the same colour. Remarkable is the identity distribution for the FASTA run, which returned a lot more hits with little identity, that the other runs. All in all, FASTA returned almost 3000 Hits (default parameter search), while all the BLAST/PSIBLAST runs returned not more than 300 Hits each. Therefore a lot of the FASTA 'Hits' have little identity and a quite low E-value (see below), but still FASTA returned some good results.


  • Evalue Distribution
E-value Distribution for the results from the Sequence Searches

The E-value distribution for the sequence searches is quite similar concerining the BLAST and PSI-BLAST results. Only FASTA has a lot wider E-value range, which can be explained by the fact, that FASTA returned about 10 times more Hits, among which a lot of sequences have little identity and therefore a quite high E-value.



Sequences chosen for the multiple Alignment:

SeqIdentifier Seq Identity source
99-90% Sequence Identity
56967006|pdb|1X7Z 99% PSI BLAST, 3 iterations, E-value cutoff 0.005
7546384|pdb|1DTW 95% BLAST
34810149|pdb|1OLU 99% PSI BLAST, 3 iterations, E-value cutoff 10E-6
13277798|gb|AAH03787.1 95% PSI BLAST, 3 iterations, E-value cutoff 10E-6
148727347|ref|NP_001092034.1 95% BLAST
89-60% Sequence Identity
196011048|ref|XP_002115388.1 66% PSI BLAST, 3 iterations, E-value cutoff 0.005
149543950|ref|XP_001517857.1 67% BLAST
47227873|emb|CAG09036.1 82,5% FASTA
47196273|emb|CAF88112.1 81% PSI BLAST, 5 iterations, E-value cutoff 0.005
12964598|dbj|BAB32665.1 88% PSI BLAST, 5 iterations, E-value cutoff 10E-6
59-40% Sequence Identity
193290664|gb|ACF17640.1 47% BLAST
215431443|ref|ZP_03429362.1 40% FASTA
225557347|gb|EEH05633.1 51% PSI BLAST, 3 iterations, E-value cutoff 10E-6
58267618|ref|XP_570965.1 50% PSI BLAST, 5 iterations, E-value cutoff 0.005
162449842|ref|YP_001612209.1 41% PSI BLAST, 5 iterations, E-value cutoff 10E-6
39-20% Sequence Identity
56966700|pdb|1W85 31% PSI BLAST, 3 iterations, E-value cutoff 0.005
5822330|pdb|1QS0 38.1% FASTA
13516864|dbj|BAB40585.1 33% PSI BLAST, 3 iterations, E-value cutoff 10E-6
284166853|ref|YP_003405132.1 35% PSI BLAST, 5 iterations, E-value cutoff 0.005
76800932|ref|YP_325940.1 34% PSI BLAST, 5 iterations, E-value cutoff 10E-6

Sequences for the Multiple Sequences Alignment were downloaded via NCBI, the sequence id can be changed in the link to retrieve the fasta format: http://www.ncbi.nlm.nih.gov/protein/76800932?report=fasta

Multiple Alignments

clustalw sequences.fasta

t_coffee -seq sequences.fasta

t_coffee -seq sequences.fasta -mode expresso

muscle -in sequences.fasta -out output.aln

download cobalt

./cobalt -i sequences.fasta -norps T > output.aln


Conservation and Gaps

Alignment methods Gaps Conserved Columns
Gaps Avg Gap Length 100% cons >90% cons >80% cons >70% cons >60% cons >50% cons
ClustalW 12 3,75 24 136 53 60 71 91
T-Coffee 25 4,56 24 136 53 60 71 91
T-Coffee (3D) 56 4,75 21 326 83 92 67 71
Cobalt 19 3,26 24 128 68 56 101 72
Muscle 17 4,76 26 193 26 38 43 10


Gaps in secondary structure

ClustalW

ClustalW gaps and structure [1]
Gap position Gap length Secondary structure
109-110 4 Helix
142-143 1 Helix
235-236 1 Beta strand
276-277 11 Helix
294-295 1 Beta strand
394-395 5 Helix

T-Coffee

T-Coffee gaps and structure [2]
Gap position Gap length Secondary structure
141-142 1 Helix
232-233 1 Beta strand
275-276 11 Helix
310-311 1 Helix
369-370 5 Turn
395-396 18 Helix
398-399 5 Helix

T-Coffee 3d

T-Coffee (3D) gaps and structure [3]
Gap position Gap length Secondary structure
101-102 1 Helix
108-109 4 Helix
115-116 1 Helix
116-117 1 Helix
141-142 1 Helix
153-154 1 Beta strand
163-164 1 Helix
177-178 3 Helix
234-235 1 Beta strand
263-264 4 Beta strand
265-266 1 Beta strand
276-277 2 Helix
308-309 8 Helix
309-310 5 Helix
314-315 6 Helix
362-363 6 Helix
371-372 4 Turn
376-377 1 Helix
380-381 1 Helix
382-383 7 Helix
383-384 3 Helix
384-385 2 Helix
387-388 2 Helix
394-395 5 Helix

Cobalt

Cobalt gaps and structure [4]
Gap position Gap length Secondary structure
108-109 4 Helix
141-142 1 Helix
177-178 3 Helix
276-277 11 Helix
294-295 1 Beta strand
305-306 1 Helix
311-312 2 Helix
387-388 1 Helix
388-389 1 Helix
395-396 13 Helix

Muscle

Muscle gaps and structure [5]
Gap position Gap length Secondary structure
109-110 4 Helix
141-142 1 Helix
177-178 3 Helix
276-277 11 Helix
294-295 1 Beta strand
305-306 1 Helix
394-395 5 Helix

Functionally important residues

The functionally important sites are according to [7] the following sites:

  • Metal binding site, 206
  • Metal binding site, 211
  • Metal binding site, 212

References

Secondary structure information

back to Reference Sequence of BCKDHA back to Maple syrup urine disease main page

Retrieved from "https://i12r-studfilesrv.informatik.tu-muenchen.de/wiki/index.php?title=Sequence_Alignments&oldid=1018"