c Ergothioneine (ERG) and mycothiol (MSH) are two low-molecular-weight thiols synthesized by mycobacteria. The role of MSH has been extensively investigated in mycobacteria; however, little is known about the role of ERG in mycobacterial physiology. In this study, quantification of ERG at various points in the growth cycle of Mycobacterium smegmatis revealed that a significant portion of ERG is found in the culture media, suggesting that it is actively secreted. A mutant of M. smegmatis lacking egtD (MSMEG_6247) was unable to synthesize ERG, confirming its role in ERG biosynthesis. Deletion of egtD from wild-type M. smegmatis and an MSH-deficient mutant did not affect their susceptibility to antibiotics tested in this study. The ERG-and MSH-deficient double mutant was significantly more sensitive to peroxide than either of the single mutants lacking either ERG or MSH, suggesting that both thiols play a role in protecting M. smegmatis against oxidative stress and that ERG is able to partly compensate for the loss of MSH. G lutathione (GSH) is a thiol known for its efficient detoxification of reactive oxygen species, reactive nitrogen species, and free radicals in eukaryotes. Mycobacteria do not synthesize GSH but produce two low-molecular-weight thiols,and ergothioneine (2-mercaptohistidine trimethylbetaine) (ERG) (4, 5). Four genes are involved in MSH biosynthesis in mycobacteria, namely, mshA, mshB, mshC, and mshD, and mutants harboring deletions in mshB, mshC, and mshD produce different levels of MSH due to the ability of other enzymes to partially compensate for their loss (6, 7). MSH-deficient mutants of Mycobacterium smegmatis show increased sensitivity to oxidative stress, alkylating agents, and a range of antibiotics, including erythromycin, azithromycin, vancomycin, penicillin G, streptomycin, and rifampin, but exhibit increased resistance to isoniazid (INH) and ethionamide (ETH) (8, 9). The MSH-deficient ⌬mshA mutant of Mycobacterium tuberculosis requires catalase during in vitro growth, implicating MSH in detoxifying reactive oxygen species (10).ERG biosynthetic genes (egtA, egtB, egtC, egtD, and egtE) were recently identified in M. smegmatis (11). Although several lines of evidence support the cytoprotective and antioxidative role of ERG in eukaryotes (12), bacteria (13), and, recently, fungi (14), nothing is known of its role in mycobacteria. ERG has also been implicated in modulating the immune response (15) and in the inhibition of metalloenzymes, preventing the copper-induced oxidation of DNA and protein due to its metal-chelating properties (16,17). Eukaryotes obtain ERG from their diet, and its accumulation in cells is dependent on the activity of a highly specific transporter, OCTN1, since the zwitterionic nature of ERG prevents it from crossing the plasma membrane (18,19). In an M. smegmatis ⌬mshA mutant, which is MSH deficient, the levels of ERG and the organic hydroperoxide resistance (Ohr) protein are elevated, suggesting that ERG may partly compensate for the loss of MSH (20). This may exp...

No abstract

The high acquisition rate of drug resistance by necessitates the ongoing search for new drugs to be incorporated in the tuberculosis (TB) regimen. Compounds used for the treatment of other diseases have the potential to be repurposed for the treatment of TB. In this study, a high-throughput screening of compounds against thiol-deficient strains and subsequent validation with thiol-deficient strains revealed that and mutants had increased susceptibility to azaguanine (Aza) and sulfaguanidine (Su); and mutants had increased susceptibility to bacitracin (Ba); and, , and mutants had increased susceptibility to fusaric acid (Fu). Further analyses revealed that some of these compounds were able to modulate the levels of thiols and oxidative stress in This study reports the activities of Aza, Su, Fu, and Ba against and provides a rationale for further investigations.

No abstract