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Abstract
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Pyrazinamide(PZA) is one the first line anti-tuberculosisdrugs that require activation by the pyrazinamidase(PZase).Most PZA-resistantMycobacterium tuberculosisstrains have mutations in the pncAgene whichencoding PZase that result in thereduction or loss of the enzyme activity. Herein, we have examined how various mutations, which have been found from the PZA-resistantM. tuberculosis strains in Iran, modify the PZase activity.To elucidate thepossible 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 enzymeswasdecreased, 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 illustratedthat the positions of the 51stto 71st residues were more dynamics in MUT2 as compared to the other atoms in PZase, MUT1 and MUT3 which could decrease the Km and kcat values of the enzymes.
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