Task 5: Mapping point mutations
- 1 Task description
- 2 SNP databases
- 3 Comparing the annotation of HGMD and SNPdb
- 4 Discussion
A detailed task description can be found here: Mapping point mutations
- Searched for PAH
- 429 Missense/Nonsense mutations known by HGMD Professional
There are several mutation types known for PAH:
- Small deletions
- Small insertions
- Small indels
- Gross deletions
- Gross insertions/duplications
- Complex rearrangements
One additional category of mutation is known, but is not recorded for PAH
- Repeat variations
The reference sequence is given by the accession number NM_000277.1, whose entry contains the following amino acid sequence:
MSTAVLENPGLGRKLSDFGQETSYIEDNCNQNGAISLIFSLKEEVGALAKVLRLFEEN DVNLTHIESRPSRLKKDEYEFFTHLDKRSLPALTNIIKILRHDI GATVHELSRDKKKDTVPWFPRTIQELDRFANQILSYGAELDADHPGFKDPVYRARRKQ FADIAYNYRHGQPIPRVEYMEEEKKTWGTVFKTLKSLYKTHACYEYNHIFPLLEKYCG FHEDNIPQLEDVSQFLQTCTGFRLRPVAGLLSSRDFLGGLAFRVFHCTQYIRHGSKPM YTPEPDICHELLGHVPLFSDRSFAQFSQEIGLASLGAPDEYIEKLATIYWFTVEFGLC KQGDSIKAYGAGLLSSFGELQYCLSEKPKLLPLELEKTAIQNYTVTEFQPLYYVAESF NDAKEKVRNFAATIPRPFSVRYDPYTQRIEVLDNTQQLKILADSINSEIGILCSALQK IK
We searched in dbSNP for silent mutations in coding regions. This means we only considered those SNPs which alter the triplet but not the amino acid.
To do so we used the Entrez interface of NCBI which is accessible under this URL:
The advantage of this Entrez interface is that we can construct arbitrary complex queries to restrict our result set.
We constructed the following query to search for SNPs which are considered silent in the coding regions of the human PAH gene (see figure 1):
- "synonymous-codon"[Function_Class] AND PAH[GENE] AND "human"[ORGN] AND "snp"[SNP_CLASS]
Results of this query can be accessed directly via the following URL:
We decided to download the results as FlatFile. This seemed to be the most simple format to process and contains almost all information we need.
To parse the important information out of the FlatFile we downloaded in the previous step we wrote a Perl script. In the current version the following information is parsed out of the FlatFile: identifier, triplet reference/mutated, allele reference/mutated , frame of the mutation, residue reference/mutated and residue position.
All annotations we retrieved are annotated for the mRNA sequence with the GenBank accession number NM_000277.1 which contains the coding sequences of our PAH protein with the accession number NP_000268.1
We could find the following silent mutations in dbSNP:
|Identifier||AA-Position||Reference Triplet||Mutated Triplet||Reference Allele||Mutated Allele||Frame||Reference Residue||Mutated Residue|
Comparing the annotation of HGMD and SNPdb
Alignment of the reference sequences
We decided to use the sequence of PAH of Uniprot (see UniProt).
Alignment with the reference sequence used in HGMD
The resulting alignment shows a 100% identity without any gaps. Therefore it is a "self-alignment".