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The redox cofactor F420 protects mycobacteria from diverse antimicrobial compounds and mediates a reductive detoxification system.

ABSTRACT: A defining feature of mycobacterial redox metabolism is the use of an unusual deazaflavin cofactor, F420 This cofactor enhances the persistence of environmental and pathogenic mycobacteria, including following antimicrobial treatment, though the molecular basis for this remains to be understood. In this work, we explored our hypothesis that F420 enhances persistence by serving as a cofactor in antimicrobial-detoxifying enzymes. To test this, we performed a series of phenotypic, biochemical, and analytical chemistry studies in relation to the model soil bacterium Mycobacterium smegmatis Mutant strains unable to synthesize or reduce F420 were found to be more susceptible to a wide range of antibiotic and xenobiotic compounds. Compounds from three classes of antimicrobial compounds traditionally resisted by mycobacteria inhibited growth of F420 mutant strains at subnanomolar concentrations, namely furanocoumarins (e.g. methoxsalen), arylmethanes (e.g. malachite green), and quinone analogues (e.g. menadione). We demonstrated that promiscuous F420H2-dependent reductases directly reduce these compounds by a mechanism consistent with hydride transfer. Moreover, M. smegmatis strains unable to make F420H2 lost the capacity to reduce and detoxify representatives of the furanocoumarin and arylmethane compound classes in whole-cell assays. By contrast, mutant strains were only slightly more susceptible to clinical antimycobacterials and this appeared to be due to indirect effects of F420 loss-of-function (e.g. redox imbalance) rather than loss of a detoxification system. Together, these data show that F420 enhances antimicrobial resistance in mycobacteria and suggest that one function of the F420H2-dependent reductases is to broaden the range of natural products that mycobacteria and possibly other environmental actinobacteria can reductively detoxify.This study reveals that a unique microbial cofactor, F420, is critical for antimicrobial resistance in the environmental actinobacterium Mycobacterium smegmatis We show that a superfamily of redox enzymes, the F420H2-dependent reductases, can reduce diverse antimicrobials in vitro and in vivo M. smegmatis strains unable to make or reduce F420 become sensitive to inhibition by these antimicrobial compounds. This suggests that mycobacteria have harnessed the unique properties of F420 to reduce structurally diverse antimicrobials as part of the antibiotics arm race. The F420H2-dependent reductases that facilitate this process represent a new class of antimicrobial-detoxifying enzymes with potential applications in bioremediation and biocatalysis.

SUBMITTER: Jirapanjawat T

PROVIDER: S-EPMC5103081 | biostudies-other | 2016 Sep

REPOSITORIES: biostudies-other

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The Redox Cofactor F420 Protects Mycobacteria from Diverse Antimicrobial Compounds and Mediates a Reductive Detoxification System.

Jirapanjawat Thanavit T Ney Blair B Taylor Matthew C MC Warden Andrew C AC Afroze Shahana S Russell Robyn J RJ Lee Brendon M BM Jackson Colin J CJ Oakeshott John G JG Pandey Gunjan G Greening Chris C

Applied and environmental microbiology 20160916 23

A defining feature of mycobacterial redox metabolism is the use of an unusual deazaflavin cofactor, F420 This cofactor enhances the persistence of environmental and pathogenic mycobacteria, including after antimicrobial treatment, although the molecular basis for this remains to be understood. In this work, we explored our hypothesis that F420 enhances persistence by serving as a cofactor in antimicrobial-detoxifying enzymes. To test this, we performed a series of phenotypi ...[more]

PMID: 27637879

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Project description:Cells receive growth and survival stimuli through their attachment to an extracellular matrix (ECM). Overcoming the addiction to ECM-induced signals is required for anchorage-independent growth, a property of most malignant cells. Detachment from ECM is associated with enhanced production of reactive oxygen species (ROS) owing to altered glucose metabolism. Here we identify an unconventional pathway that supports redox homeostasis and growth during adaptation to anchorage independence. We observed that detachment from monolayer culture and growth as anchorage-independent tumour spheroids was accompanied by changes in both glucose and glutamine metabolism. Specifically, oxidation of both nutrients was suppressed in spheroids, whereas reductive formation of citrate from glutamine was enhanced. Reductive glutamine metabolism was highly dependent on cytosolic isocitrate dehydrogenase-1 (IDH1), because the activity was suppressed in cells homozygous null for IDH1 or treated with an IDH1 inhibitor. This activity occurred in absence of hypoxia, a well-known inducer of reductive metabolism. Rather, IDH1 mitigated mitochondrial ROS in spheroids, and suppressing IDH1 reduced spheroid growth through a mechanism requiring mitochondrial ROS. Isotope tracing revealed that in spheroids, isocitrate/citrate produced reductively in the cytosol could enter the mitochondria and participate in oxidative metabolism, including oxidation by IDH2. This generates NADPH in the mitochondria, enabling cells to mitigate mitochondrial ROS and maximize growth. Neither IDH1 nor IDH2 was necessary for monolayer growth, but deleting either one enhanced mitochondrial ROS and reduced spheroid size, as did deletion of the mitochondrial citrate transporter protein. Together, the data indicate that adaptation to anchorage independence requires a fundamental change in citrate metabolism, initiated by IDH1-dependent reductive carboxylation and culminating in suppression of mitochondrial ROS.

Dataset Information

The redox cofactor F420 protects mycobacteria from diverse antimicrobial compounds and mediates a reductive detoxification system.

Publications

The Redox Cofactor F<sub>420</sub> Protects Mycobacteria from Diverse Antimicrobial Compounds and Mediates a Reductive Detoxification System.

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