Computational protocol: Prevalence of hepatitis C virus-resistant association substitutions to direct-acting antiviral agents in treatment-naïve hepatitis C genotype 1b-infected patients in western China

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Protocol publication

[…] Clinically relevant RASs to DAAs were summarized from the literature and are presented in . The NS3, NS5A, and NS5B nucleotide sequences were aligned with the HCV 1b reference sequence (GenBank accession AJ238799) and analyzed using BioEdit 7.0.9 software. A variant type was indicated by replacement of the consensus amino acid with a novel one, eg, Y93H and Y93N in the NS5A region were described as two variant types.A phylogenetic tree of each gene was created using the maximum-likelihood statistical method with the Jones–Taylor–Thornton two-parameter model in MEGA version 5.05. A γ-parameter of 0.5 was used to model differences in substitution rates among base sites. The reliability of the cluster descending from that node was assessed by a bootstrap analysis with 1,000 replicates. [...] Protein-homology modeling was performed with Swiss-Model (https://swissmodel.expasy.org), with an identity of more than 90% used to model the X-ray crystal structures of HCV NS3 protease, NS5A, protein and NS5B polymerase for GT 1b. Structural editing and analysis were done using PyMol (Schrödinger, New York, NY, USA). The three structures were loaded into PyMol to analyze the cartoon structures, the stick structures of RASs, and the drug-binding pocket.Sequences consisting of the NS3, NS5A, and NS5B genes were aligned and translated into amino acid sequences using BioEdit 7.09. Then, all possible amino-acid covariant pairs were determined by employing the observed-minus-expected-squared (OMES) algorithm, which can exclude invariant positions and spurious correlations generated by random differences, as described in other work. To identify the covarying pairs, we calculated an OMES score for every possible pair. An OMES score of 0.5 was used as the cutoff for selecting covariant amino-acid pairs for analyses. Finally, amino-acid covariant networks were visualized by Cytoscape (version 3.2.1). […]

Pipeline specifications

Software tools BioEdit, MEGA, SWISS-MODEL, PyMOL
Databases ExPASy
Applications Phylogenetics, Protein structure analysis
Organisms Homo sapiens
Diseases Hepatitis C