Abstract
Mycobacterium tuberculosis (M. tuberculosis), the pathogen responsible for tuberculosis, detoxifies cytotoxic peroxides produced by activated macrophages. M. tuberculosis expresses alkyl hydroxyperoxide reductase E (AhpE), among other peroxiredoxins. So far the system that reduces AhpE was not known. We identified M. tuberculosis mycoredoxin-1 (MtMrx1) acting in combination with mycothiol and mycothiol disulfide reductase (MR), as a biologically relevant reducing system for MtAhpE. MtMrx1, a glutaredoxin-like, mycothiol-dependent oxidoreductase, directly reduces the oxidized form of MtAhpE, through a protein mixed disulfide with the N-terminal cysteine of MtMrx1 and the sulfenic acid derivative of the peroxidatic cysteine of MtAhpE. This disulfide is then reduced by the C-terminal cysteine in MtMrx1. Accordingly, MtAhpE catalyzes the oxidation of wt MtMrx1 by hydrogen peroxide but not of MtMrx1 lacking the C-terminal cysteine, confirming a dithiolic mechanism. Alternatively, oxidized MtAhpE forms a mixed disulfide with mycothiol, which in turn is reduced by MtMrx1 using a monothiolic mechanism. We demonstrated the H2O2-dependent NADPH oxidation catalyzed by MtAhpE in the presence of MR, Mrx1, and mycothiol. Disulfide formation involving mycothiol probably competes with the direct reduction by MtMrx1 in aqueous intracellular media, where mycothiol is present at millimolar concentrations. However, MtAhpE was found to be associated with the membrane fraction, and since mycothiol is hydrophilic, direct reduction by MtMrx1 might be favored. The results reported herein allow the rationalization of peroxide detoxification actions inferred for mycothiol, and more recently, for Mrx1 in cellular systems. We report the first molecular link between a thiol-dependent peroxidase and the mycothiol/Mrx1 pathway in Mycobacteria.
Highlights
Mycothiol, the major low-molecular weight thiol of Mycobacterium tuberculosis, is important for peroxide detoxification and virulence
Considering that alkyl hydroxyperoxide reductase E (AhpE)-SOH reacts with glutathione and N-acetylcysteine to form mixed disulfides, we hypothesized that protection was due to the formation of an MtAhpE-SS-M adduct according to Equation 5
We demonstrated the peroxidase activity of the one-cysteine peroxiredoxin from M. tuberculosis, MtAhpE, using different peroxides and artificial reducing substrates
Summary
The major low-molecular weight thiol of Mycobacterium tuberculosis, is important for peroxide detoxification and virulence. MtMrx, a glutaredoxin-like, mycothiol-dependent oxidoreductase, directly reduces the oxidized form of MtAhpE, through a protein mixed disulfide with the N-terminal cysteine of MtMrx and the sulfenic acid derivative of the peroxidatic cysteine of MtAhpE. MSH/Mrx Reduces AhpE from M. tuberculosis and the presence of millimolar concentrations of 1-D-myo-inosityl 2-(N-acetyl-L-cysteinyl)amido-2-deoxy-␣-D-glucopyranoside (mycothiol or MSH) [6,7] as the main low molecular weight thiol. Compensatory overexpression of an organic hydroperoxide resistance protein (Ohr) in M. smegmatis lacking MSH suggests the existence of a MSH-dependent organic hydroperoxide peroxidase [12] These data are only indirect evidence for the presence of a MSH-dependent peroxidase in these bacteria, the molecular link between MSH and peroxidase activity has not been clearly established yet, and purified peroxidases studied so far failed in using MSH as reducing substrate [13]. We propose a functional link between the mycothiol/mycoredoxin-1 pathway and the bacterial peroxide detoxification systems, which helps to rationalize the increased peroxide-dependent cytotoxicity in Mycobacteria with a reduced MSH content (9 –11)
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
