3bsqA Methods & Materials

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Multiple sequence alugnment was made using clustalX from the DVD. The crystal structure of all proteins involved (ASA, ASK and STS) were viewed using the protein data bank text file, downloaded to the PyMol (Downloaded version in 'colaborative learning center' (CLC)). The structural alignment was done using the DALI server with PDB identifiers. Protein interaction of ASK were searched with the programme 'STRING'which is available online. The protein name or PDB identifier was not detected by 'STRING', therefore the aminoacid sequence was used. The function of highly related proteins were found using ProFunc.

'Two sequence alignment' of ASK and STS was done with the programme SIM. The method used was 'BLOSUM62'. A 'gap penalty' of 5 and 'gap extension penalty' of 2 were used to optimise the alignment according to what was observed in 'clustalX'.The pylogenic tree was made using the 'clusalX' alignment and viewed with Treeview from the DVD.

'BLAST' search performed using the crystal structure of a putative sulfatase found from to Bacteroides Thetaiotaomicron (gi|160286517) to determine all the related species. Multiple sequence alignment was found using 'ClustalX', which was used to align the gaps among all the species sequences. 'TreeView' constructed a phylogeny tree using taxonomy to name and group all the species and illustrate the divergence and nature of the common ancestors. Phylogenetic analyses were conducted in 'MEGA4', including determining the bootstrap values. The evolutionary history was inferred using the Neighbor-Joining method. The percentage of replicate trees in which the associated taxa clustered together in the bootstrap test (500 replicates) are shown next to the branches. All positions containing alignment gaps and missing data were eliminated only in pair-wise sequence comparisons. There were a total of 1222 positions in the final dataset.