Difference between revisions of "Sequence Alignments"
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To illustrate the overlap of the returned sequences for the sequences searches, Venn Diagrams were drawn. |
To illustrate the overlap of the returned sequences for the sequences searches, Venn Diagrams were drawn. |
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| − | PSI1 = PSI-BLAST run with 3 iterations, E-value cutoff 0.005 |
+ | [[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)]] |
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| + | 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]): |
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| + | [[Image:vennDiagram.png|none|300px]] |
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| + | 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. |
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| − | PSI2 = PSI-BLAST run with 5 iterations, E-value cutoff 0.005 |
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| + | 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. |
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| − | |||
| + | All in all one could say, that the Search algorithms returned nearly the same sequences matching our query. |
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| − | PSI3 = PSI-BLAST run with 3 iterations, E-value cutoff 10E6 |
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| − | |||
| − | PSI4 = PSI-BLAST run with 5 iterations, E-value cutoff 10E6 |
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| − | |||
| − | [[Image:comparePSI10E6.png|left|300px]] |
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| − | This image shows the overlap of returned sequences for the two PSI-BLAST runs with E-value cutoff 10E6. |
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| − | |||
| − | [[Image:comparePSI0.005.png|left|300px]] |
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| − | The overlap of returned sequences for the two PSI-BLAST runs with E-value cutoff 0.005 is identical to the picture above. This indicates, that PSI-BLAST with 5 iterations returns 6 sequences more, as it may have more time to search for additional matching hits. |
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| − | |||
| − | [[Image:comparePSI3.png|left|300px]] |
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| − | The PSI-BLAST runs with each 3 iterations returned absolutely the same sequences match our query BCKDHA. From the 3 shown pictures the following conclusion can be drawn: both PSI-BLAST runs with each 5 iterations also returned absolutely the same results. |
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| − | |||
| − | [[Image:compAll.png|left|100px]] |
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| − | [[Image:compAll2.png|left|100px]] |
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| + | === Functionally important residues === |
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| − | |||
| + | The functionally important sites are according to [http://www.uniprot.org/uniprot/P12694] the following sites: |
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| − | |||
| + | * Metal binding site, 206 |
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| + | * Metal binding site, 211 |
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| + | * Metal binding site, 212 |
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=== References === |
=== References === |
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[http://www.uniprot.org/uniprot/P12694 Secondary structure information] |
[http://www.uniprot.org/uniprot/P12694 Secondary structure information] |
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| + | back to [[Reference_Sequence_BCKDHA|Reference Sequence of BCKDHA]] |
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| − | |||
back to [[Maple syrup urine disease]] main page |
back to [[Maple syrup urine disease]] main page |
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Latest revision as of 11:13, 21 May 2011
Contents
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.
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]):
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
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
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
| 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
| 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
| 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
| 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
| 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
