Studies on Nudix hydrolase proteins and crosstalk between DNA repair pathways

Abstract

Mycobacterium tuberculosis, the causative agent of tuberculosis, has become a global health concern. This calls for a dire need to understand various aspects of mycobacterial physiology in order to design better strategies to control the infection. Inside the host macrophages, pathogen encounters high oxidative and nitrosative stress and their GC rich genomes, render them susceptible to exceptionally mutagenic base modifications like oxidation of guanine to 8-O-guanine and deamination of cytosine to uracil. To safeguard its DNA, the pathogen has evolved specialized mechanisms of DNA repair. MutT hydrolyzes 8-O-dGTP present in the nucleotide pool to its monophosphate form and eliminates chances of its misincorporation in the DNA. Even though 4 orthologs of MutT have been identified, the identity of a canonical MutT remains indeterminate in mycobacteria. The MutT proteins belong to Nudix hydrolase family of proteins. To further our understanding of MutT mediated 8-O-dGTP sanitization mechanisms in mycobacteria, we carried out biochemical and functional analysis of one of the mycobacterial Nudix hydrolase family proteins in the first part of the study. In the second part, we tested the functions of Nucleoside diphosphate kinase (NDK), known to maintain nucleotide pools, towards 8-O-dGTP using E. coli model system. In addition, Base Excision Repair (BER) and Nucleotide Excision Repair (NER) pathways are believed to play major roles in DNA repair in mycobacteria because of the absence of mismatch repair system and little contributions from RecA in eliciting the DNA damage response. In other organisms, NER has been observed to contribute in the repair of single nucleotide damage, facilitated by BER pathway specific proteins. In part III of the study, we have worked on a hypothesis that DNA damage repair by a uracil DNA glycosylase (UdgB) in mycobacteria invites NER pathway proteins to complete the repair.