title: Biochemical Characterization and Computational Identification of Mycobacterium tuberculosis Pyrazinamidase in Some Pyrazinamide-Resistant Isolates of Iran.
abstract: Pyrazinamide (PZA) is one the first line anti-tuberculosis drugs that require activation by the pyrazinamidase (PZase). Most PZA-resistant Mycobacterium tuberculosis strains have mutations in the pncA gene which encoding PZase that result in the reduction or loss of the enzyme activity. Herein, we have examined how various mutations, which have been found from the PZA-resistant M. tuberculosis strains in Iran, modify the PZase activity. To elucidate the possible role of these mutations, namely A143T (MUT1), L151S (MUT2), A143T/T168A/E173K (MUT3), in the bioactivity of the enzyme, the PZase and mutant genes were cloned, functionally expressed and biochemically and computationally characterized. In comparison to the PZase enzyme, the enzymatic efficiency of mutant enzymes was decreased, with MUT2 indicating the largest enzymatic efficiency reduction. Homology models of mutants were constructed based on the PZase X-ray crystal structure. Molecular modeling and substrate docking revealed that the wild-type has much stronger binding affinity to PZA than the mutants whereas MUT2 has the weakest binding affinity. In addition, the molecular dynamics simulations and the essential dynamics results illustrated that the positions of the 51st to 71st residues were more dynamics in MUT2 as compared to the other atoms in PZase, MUT1 and MUT3 which could decrease the K(m) and k(cat) values of the enzymes.
authors.affiliation: Infectious Diseases and Tropical Medicine Research Center, Shahid Beheshti University of Medical Sciences, P. O. Box 1985717443, Tehran, Iran.
authors.forename: Farahnoosh
authors.initials: F
authors.lastname: Doustdar
authors.forename: Mohammad
authors.initials: M
authors.lastname: Pazhang
authors.forename: Faramarz
authors.initials: F
authors.lastname: Mehrnejad
authors.forename: Mehrnoosh
authors.initials: M
authors.lastname: Safarzadeh
authors.forename: Davod
authors.initials: D
authors.lastname: Rabiei
authors.forename: Nader
authors.initials: N
authors.lastname: Chaparzadeh
authors.forename: Hanieh
authors.initials: H
authors.lastname: Falahati
authors.forename: Mohammad
authors.initials: M
authors.lastname: Mir-Derikvand
mesh_terms: D000581:Amidohydrolases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D000595:Amino Acid Sequence; D000995:Antitubercular Agents / Q000494:pharmacology; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D024881:Drug Resistance, Bacterial; D006801:Humans; D006860:Hydrogen Bonding; D007492:Iran; D056004:Molecular Dynamics Simulation; D008969:Molecular Sequence Data; D009154:Mutation; D009169:Mycobacterium tuberculosis / Q000187:drug effects* / Q000201:enzymology* / Q000235:genetics; D011718:Pyrazinamide / Q000494:pharmacology*; D016415:Sequence Alignment; D014376:Tuberculosis / Q000382:microbiology*
chemical_list: D000995:Antitubercular Agents; D001426:Bacterial Proteins; D011718:Pyrazinamide; D000581:Amidohydrolases; C022490:pyrazinamide deamidase