Structural dynamics behind variants in pyrazinamidase and pyrazinamide resistance

Abstract

Pyrazinamide (PZA) is an important component of first-line anti-tuberculosis (anti-TB) drugs. The anti-TB agent is activated into an active form, pyrazinoic acid (POA), by Mycobacterium tuberculosis (MTB) pncA gene encoding pyrazinamidase (PZase). The major cause of PZA-resistance has been associated with mutations in the pncA gene. We have detected several novel mutations including V131F, Q141P, R154T, A170P, and V180F (GeneBank Accession No. MH461111) in the pncA gene of PZA-resistant isolates during PZA drug susceptibility testing followed by pncA gene sequencing. Here, we investigated molecular mechanism of PZA-resistance by comparing the results of experimental and molecular dynamics. The mutants (MTs) and wild type (WT) PZase structures in apo and complex with PZA were subjected to molecular dynamic simulations (MD) at the 40 ns. Multiple factors, including root mean square deviations (RMSD), binding pocket, total energy, dynamic cross correlation, and root mean square fluctuations (RMSF) of MTs and WT were compared. The MTs attained a high deviation and fluctuation compared to WT. Binding pocket volumes of the MTs, were found, lower than the WT, and the docking scores were high than WT while shape complementarity scores were lower than that of the WT. Residual motion in MTs are seemed to be dominant in anti-correlated motion. Mutations at locations, V131F, Q141P, R154T, A170P, and V180F, might be involved in the structural changes, possibly affecting the catalytic property of PZase to convert PZA into POA. Our study provides useful information that will enhance the understanding for better management of TB. Abbreviations DST drug susceptibility testing Δelec electrostatic energy LJ Lowenstein–Jensen medium MGIT mycobacterium growth indicator tubes MTs mutants MD molecular dynamic simulations MTB Mycobacterium tuberculosis NALC–NaOH N-acetyl-l-cysteine–sodium hydroxide NIH National Institutes of Health NPT amount of substance (N), pressure (P) temperature (T) NVT moles (N), volume (V) temperature (T) PZase pyrazinamidase Δps polar solvation energy PTRL Provincial Tuberculosis Reference Laboratory RMSD root mean square deviations RMSF root mean square fluctuations ΔSASA solvent accessible surface area energy TB tuberculosis GTotal total binding free energy ΔvdW Van der Waals energy WT wild type Communicated by Ramaswamy H. Sarma