The Study of mutations effect on the inactivation of pyrazinamidase by molecular dynamics simulations

Pyrazinamide is one of the most important drugs in the treatment of latent tuberculosis infection. PncA gene encodes Pyrazinamidase (PZase) enzyme of Mycobacterium tuberculosis that is responsible for conversion of Pyrazinamide (PZA) to Pyrazinoic acid (active form). In spite of the role of this drug in shortening of the treatment period from 9 months to 6, the emergence of strains resistant to PZA represents an important public health problem. Pyrazinamidase mutations are associated to PZA – resistant phenotype. However, the relationship between mutations and structural changes that inactive the enzyme, has not been known very well. In this study, the PZase gene from the H37Rv strain (wild type) and two resistant strain of Mycobacterium tuberculosises to PZA were cloned, expressed and their activity were investigated by the method based on Wayne test. The structures of wild type enzyme and mutants (D63G/W119C and T160P) were modeled using homology modeling and single amino acid replacement, then structural parameters were calculated. Enzyme activity determination results revealed that the mutants lose their activity, completely. Computational results also confirmed that mutations affect the secondary structure of the enzyme and induce structural changes. These changes can alter the structure of the enzyme active site in D63G/W119C or shorten opening of the substrate binding site in T160P.