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Detoxification of glutathione and nitrosoglutathione by thioredoxin system of Mycobacterium tuberculosis Attarian, Rodgoun

Abstract

Tuberculosis is the leading cause of mortality due to a single pathogenic infection. Its etiological agent, Mycobacterium tuberculosis infects, resides and multiplies within human alveolar macrophages. It is exposed to reactive oxygen intermediates and reactive nitrogen intermediates (RNI) such as nitric oxide (NO) produced within phagosomes and granulomas against invading pathogens. Therefore, proliferation of M. tuberculosis within the host depends on its strategies to counteract the onslaught of these intermediates. One example is recruitment of the thioredoxin system as one of the most proficient pathways for protection against oxidative stress, since it dominates peroxide detoxification pathways. A burst of NO within macrophages parallels the production of glutathione (GSH) to protect the host against NO toxicity. The macrophage GSH pool reduces NO to S-nitrosoglutathione (GSNO). Both glutathione disulphide (GSSG) and GSNO possess mycobactericidal activities in vitro. This thesis is focused on characterizing the role of M. tuberculosis thioredoxin system in detoxification of antimycobacterial compounds produced within the host such as GSSG and GSNO, due to the intrinsic capacity of the system to universally reduce disulfide bonds and reduce GSNO in humans. By performing NADPH oxidation assays and HPLC analysis we demonstrate that M. tuberculosis thioredoxin redox cascade is a general reduction system able to efficiently reduce the low molecular weight disulfides GSSG and MSSM, and dissimilate GSNO. We also investigated the cellular pathways in which thioredoxin of M. tuberculosis participates. Here, we present an analysis of the thioredoxin-linked M. tuberculosis proteome by using a substrate trapping assay and mass spectrometry. We have identified eleven proteins associated with TrxC, implicating the involvement of thioredoxin in distinct cellular processes in this pathogen. The findings described in this thesis elucidate a novel function for the thioredoxin system of M. tuberculosis. We demonstrate that this system serves as a detoxification pathway against mycobactericidal compounds such as GSSG and GSNO. Overall, the data presented here establishes that M. tuberculosis thioredoxin has pleiotropic roles and is involved in a spectrum of processes from metabolic pathways to gene expression and signal transduction.

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