Insights Into Mutations Induced Conformational Changes and Rearrangement of Fe2+ Ion in pncA Gene of Mycobacterium tuberculosis to Decipher the Mechanism of Resistance to Pyrazinamide

Asma Sindhoo Nangraj; Abbas Khan; Shaheena Umbreen; Sana Sahar; Maryam Arshad; Saba Younas; Sajjad Ahmad; Shahid Ali; Syed Shujait Ali; Liaqat Ali; Dong-Qing Wei

doi:10.3389/fmolb.2021.633365

Insights Into Mutations Induced Conformational Changes and Rearrangement of Fe²⁺ Ion in pncA Gene of Mycobacterium tuberculosis to Decipher the Mechanism of Resistance to Pyrazinamide

Front Mol Biosci. 2021 May 20:8:633365. doi: 10.3389/fmolb.2021.633365. eCollection 2021.

Authors

Asma Sindhoo Nangraj¹, Abbas Khan¹, Shaheena Umbreen², Sana Sahar³, Maryam Arshad⁴, Saba Younas⁵, Sajjad Ahmad⁶, Shahid Ali⁷, Syed Shujait Ali⁷, Liaqat Ali⁸, Dong-Qing Wei^{1

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Affiliations

¹ Department of Bioinformatics and Biological Statistics, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai, China.
² Department of Botany, University of Okara, Okara, Pakistan.
³ The Islamia University of Bahawalpur, Bahawalpur, Pakistan.
⁴ Government College University Faisalabad, Sahiwal, Pakistan.
⁵ University of Education, Lahore, Pakistan.
⁶ Department of Health and Biological Sciences, Abasyn University, Peshawar, Pakistan.
⁷ Center for Biotechnology and Microbiology, University of Swat, Swat, Pakistan.
⁸ Department of Biological Sciences, National University of Medical Sciences, Islamabad, Pakistan.
⁹ Peng Cheng Laboratory, Shenzhen, China.
¹⁰ State Key Laboratory of Microbial Metabolism, Shanghai-Islamabad-Belgrade Joint Innovation Center on Antibacterial Resistances, Joint International Research Laboratory of Metabolic and Developmental Sciences, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai, China.

Abstract

Pyrazinamide (PZA) is the first-line drug commonly used in treating Mycobacterium tuberculosis (Mtb) infections and reduces treatment time by 33%. This prodrug is activated and converted to an active form, Pyrazinoic acid (POA), by Pyrazinamidase (PZase) enzyme. Mtb resistance to PZA is the outcome of mutations frequently reported in pncA, rpsA, and panD genes. Among the mentioned genes, pncA mutations contribute to 72-99% of the total resistance to PZA. Thus, considering the vital importance of this gene in PZA resistance, its frequent mutations (D49N, Y64S, W68G, and F94A) were investigated through in-depth computational techniques to put conclusions that might be useful for new scaffolds design or structure optimization to improve the efficacy of the available drugs. Mutants and wild type PZase were used in extensive and long-run molecular dynamics simulations in triplicate to disclose the resistance mechanism induced by the above-mentioned point mutations. Our analysis suggests that these mutations alter the internal dynamics of PZase and hinder the correct orientation of PZA to the enzyme. Consequently, the PZA has a low binding energy score with the mutants compared with the wild type PZase. These mutations were also reported to affect the binding of Fe²⁺ ion and its coordinated residues. Conformational dynamics also revealed that β-strand two is flipped, which is significant in Fe²⁺ binding. MM-GBSA analysis confirmed that these mutations significantly decreased the binding of PZA. In conclusion, these mutations cause conformation alterations and deformities that lead to PZA resistance.

Keywords: PCA; PZA; free energy; mutations; simulation.