title: Sequence and structural characterization of tbnat gene in isoniazid-resistant Mycobacterium tuberculosis: identification of new mutations.
abstract: The present study was carried out to investigate the presence of polymorphism in the N-acetyltransferase gene of 41 clinical isolates of Mycobacterium tuberculosis, that were resistant to isoniazid (INH) with no mutations in the hot spots of the genes previously described to be involved in INH resistance (katG, inhA and ahpC). We observed single nucleotide polymorphisms (SNPs) in ten of these, including the G619A SNP in five isolates and an additional four so far un-described mutations in another five isolates. Among the latter SNPs, two were synonymous (C276T, n=1 and C375G, n=3), while two more non-synonymous SNPs were composed of C373A (Leu→Met) and T503G (Met→Arg) were observed in respectively one and two isolates. Molecular modeling and structural analysis based in a constructed full length 3D models of wild type TBNAT (TBNAT_H37Rv) and the isoforms (TBNAT_L125M and TBNAT_M168R) were also performed. The refined models show that, just as observed in human NATs, the carboxyl terminus extends deep within the folded enzyme, into close proximity to the buried catalytic triad. Analysis of tbnat that present non-synonymous mutations indicates that both substitutions are plausible to affect enzyme specificity or acetyl-CoA binding capacity. The results contribute to a better understanding of structure-function relationships of NATs. However, further investigation including INH-sensitive strains as a control group is needed to get better understanding of the possible role of these new mutations on tuberculosis control.
authors.affiliation: Post-Graduation Program in Applied Genetics and Toxicology Program, Lutheran University of Brazil (PPGGTA/ULBRA), Canoas, RS, Brazil.
authors.forename: Millene Borges
authors.initials: MB
authors.lastname: Coelho
authors.forename: Elis Regina Dalla
authors.initials: ER
authors.lastname: Costa
authors.forename: Sidra Ezídio Gonçalves
authors.initials: SE
authors.lastname: Vasconcellos
authors.forename: Natali
authors.initials: N
authors.lastname: Linck
authors.forename: Ricardo Martins
authors.initials: RM
authors.lastname: Ramos
authors.forename: Hermes Luís Neubauer de
authors.initials: HL
authors.lastname: Amorim
authors.forename: Philip Noel
authors.initials: PN
authors.lastname: Suffys
authors.forename: Adalberto Rezende
authors.initials: AR
authors.lastname: Santos
authors.forename: Pedro Eduardo Almeida da
authors.initials: PE
authors.lastname: Silva
authors.forename: Daniela Fernandes
authors.initials: DF
authors.lastname: Ramos
authors.forename: Márcia Susana Nunes
authors.initials: MS
authors.lastname: Silva
authors.forename: Maria Lucia Rosa
authors.initials: ML
authors.lastname: Rossetti
mesh_terms: D000995:Antitubercular Agents / Q000494:pharmacology*; D001191:Arylamine N-Acetyltransferase / Q000235:genetics*; D024881:Drug Resistance, Bacterial; D005798:Genes, Bacterial; D007527:Isoenzymes / Q000235:genetics; D007538:Isoniazid / Q000494:pharmacology*; D008826:Microbial Sensitivity Tests; D008958:Models, Molecular; D009154:Mutation; D009169:Mycobacterium tuberculosis / Q000235:genetics; D020641:Polymorphism, Single Nucleotide
chemical_list: D000995:Antitubercular Agents; D007527:Isoenzymes; D001191:Arylamine N-Acetyltransferase; D007538:Isoniazid