Computational protocol: Study on Phylogenetic Relationships, Variability, and Correlated Mutations in M2 Proteins of Influenza Virus A

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[…] The template protein used in the study was the matrix protein 2 from Influenza A virus (A/Udorn/307/1972(H3N2)) . The amino acid sequence and structure of this protein was obtained from Protein Data Bank ( (pdb code 2RLF).The virus M2 protein sequences were taken from the UniProtKB/Swiss-Prot protein sequence database ( The M2 sequences revealing significant identity/similarity scores to the 2RLF were selected by protein BLAST (blastp) – at the default values of search.For the preliminary multiple sequence alignment ClustalX – was used, then the alignment was verified and corrected if necessary by manual analysis following the algorithm of genetic semihomology –. The verification concerned the possible genetic relationship between compared positions which were non-identical (possible replacements by single transition/transversion). The consensus sequence of the aligned M2 proteins was constructed with the aid of Consensus Constructor . [...] The phylogenetic trees were constructed on the basis of different algorithms of distance calculation and with the aid of different applications. The programs used for cladogram and/or phylogram construction were: ClustalX –, SSSSg (freely accessible at:, ConSurf – and Phylip . The phylogenetic trees were constructed on the basis of the aligned complete sequences and of the 43 residue fragment corresponding to 2RLF. [...] The mutational variability/conservativity of the aligned homologous positions and regions was analysed with the use of ConSurf – and Talana (available at: The variability pattern plotted on the 2RLF structure was visualised by Rastop2.2 ( […]

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