Insight into novel clinical mutants of RpsA-S324F, E325K, and G341R of Mycobacterium tuberculosis associated with pyrazinamide resistance

Dong-Qing Wei,Shaukat Iqbal Malik,Muhammad Tahir Khan,Ashfaq Ur Rehaman,Muhammad Junaid

doi:10.1016/j.csbj.2018.09.004

Abstract

Pyrazinamide (PZA) is an important component of first-line anti-tuberculosis drugs which is converted into active form, pyrazinoic acid (POA), by Mycobacterium tuberculosis (MTB) pncA gene encoded, pyrazinamidase (PZase). Mutations in pncA are detected in >70% of PZA resistant isolates but, noticeably, not in all. In this study, we selected 18 PZA-resistant but wild type pncA (pncAWT) MTB isolates. Drug susceptibility testing (DST) of all the isolates were repeated at the critical concentration of PZA drug. All these PZA-resistance but pncAWT isolates were subjected to RpsA sequencing. Fifteen different mutations were identified in eleven isolates, where seven were present in a conserved region including, Ser324Phe, Glu325Lys, Gly341Arg. As the molecular mechanism of resistance behind these variants has not been reported earlier, we have performed multiple analysis to unveil the mechanisms of resistance behind mutations S324F, E325K, and G341R. The mutant and wild type RpsA structures were subjected to comprehensive computational molecular dynamic simulations at 50 ns. Root mean square deviation (RMSD), Root mean square fluctuation (RMSF), and Gibbs free energy of mutants were analyzed in comparison with wild type. Docking score of wild type-RpsA has been found to be maximum, showing a strong binding affinity in comparison with mutants. Pocket volume, RMSD and RMSF have also been found to be altered, whereas total energy, folding effect (radius of gyration) and shape complimentarily analysis showed that variants S324F, E325K, and G341R have been playing a significant role behind PZA-resistance. The study offers valuable information for better management of drug resistance tuberculosis.

Highlights

Pyrazinamide (PZA) is a first-line drug used in combination with rifampin and isoniazid that kills persister bacilli, and was found to be effective in shortening the duration of TB therapy [1,2]
The emergence of first and second-line drug resistance is a major hurdle towards WHO global end TB strategy 2020
Previous studies reported that resistance to PZA develops due to mutations in pncA, but resistance may develop due to mutations in ribosomal protein S1 (RpsA) and aspartate decarboxylase genes [8,35,36,37]

Summary

Introduction

Pyrazinamide (PZA) is a first-line drug used in combination with rifampin and isoniazid that kills persister bacilli, and was found to be effective in shortening the duration of TB therapy [1,2]. The prodrug is converted into an active state, pyrazinoic acid (POA), by Mycobacterium tuberculosis (MTB) pyrazinamidase (PZase) encoded by pncA. The major targets of POA that have so far been identified are ribosomal protein S1 (RpsA), which is involved in trans-translation, and aspartate. Q. Wei, Department of Bioinformatics and Biostatistics College of Life Sciences and Biotechnology, The State Key Laboratory of Microbial Metabolism, Shanghai Jiao Tong University, 800 Dongchuan Road Shanghai, Minhang District, China, 200240. I. Malik, Department of Bioinformatics and Biosciences Capital University of Science and Technology, 44000 Islamabad Expressway, Kahuta Road, ZoneV, Islamabad

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