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Genetic basis for metabolism of methylated sulfur compounds in Methanosarcina species.

ABSTRACT: Methanosarcina acetivorans uses a variety of methylated sulfur compounds as carbon and energy sources. Previous studies implicated the mtsD, mtsF, and mtsH genes in catabolism of dimethylsulfide, but the genes required for use of other methylsulfides have yet to be established. Here, we show that a four-gene locus, designated mtpCAP-msrH, is specifically required for growth on methylmercaptopropionate (MMPA). The mtpC, mtpA, and mtpP genes encode a putative corrinoid protein, a coenzyme M (CoM) methyltransferase, and a major facilitator superfamily (MFS) transporter, respectively, while msrH encodes a putative transcriptional regulator. Mutants lacking mtpC or mtpA display a severe growth defect in MMPA medium but are unimpaired during growth on other substrates. The mtpCAP genes comprise a transcriptional unit that is highly and specifically upregulated during growth on MMPA, whereas msrH is monocistronic and constitutively expressed. Mutants lacking msrH fail to transcribe mtpCAP and grow poorly in MMPA medium, consistent with the assignment of its product as a transcriptional activator. The mtpCAP-msrH locus is conserved in numerous marine methanogens, including eight Methanosarcina species that we showed are capable of growth on MMPA. Mutants lacking the mtsD, mtsF, and mtsH genes display a 30% reduction in growth yield when grown on MMPA, suggesting that these genes play an auxiliary role in MMPA catabolism. A quadruple ?mtpCAP ?mtsD ?mtsF ?mtsH mutant strain was incapable of growth on MMPA. Reanalysis of mtsD, mtsF, and mtsH mutants suggests that the preferred substrate for MtsD is dimethylsulfide, while the preferred substrate for MtsF is methanethiol.Methylated sulfur compounds play pivotal roles in the global sulfur and carbon cycles and contribute to global temperature homeostasis. Although the degradation of these molecules by aerobic bacteria has been well studied, relatively little is known regarding their fate in anaerobic ecosystems. In this study, we identify the genetic basis for metabolism of methylmercaptopropionate, dimethylsulfide, and methanethiol by strictly anaerobic methanogens of the genus Methanosarcina. These data will aid the development of predictive sulfur cycle models and enable molecular ecological approaches for the study of methylated sulfur metabolism in anaerobic ecosystems.

SUBMITTER: Fu H

PROVIDER: S-EPMC4372740 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Background and purposeHomocystinurias are rare genetic defects characterized by altered fluxes of sulfur compounds including homocysteine and cysteine. We explored whether the severely perturbed sulfur amino acid metabolism in patients with homocystinurias affects the metabolism of hydrogen sulfide.Experimental approachWe studied 10 treated patients with a block in the conversion of homocysteine to cysteine due to cystathionine ?-synthase deficiency (CBSD) and six treated patients with remethylation defects (RMD) and an enhanced flux of sulfur metabolites via transsulfuration. Control groups for CBSD and RMD patients consisted of 22 patients with phenylketonuria on a low-protein diet and of 12 healthy controls respectively. Plasma and urine concentrations of selected sulfur compounds were analysed by HPLC and LC-MS/MS.Key resultsPatients with CBSD exhibited plasma concentrations of monobromobimane-detected sulfide similar to appropriate controls. Urinary homolanthionine and thiosulfate in CBSD were increased significantly 1.9 and 3 times suggesting higher hydrogen sulfide synthesis by ?-cystathionase and detoxification respectively. Surprisingly, patients with RMD had significantly lower plasma sulfide levels (53 and 64% of controls) with lower sulfite concentrations, and higher taurine and thiosulfate levels suggesting enhanced cysteine oxidation and hydrogen sulfide catabolism respectively.Conclusion and implicationsThe results from this study suggest that severe inherited defects in sulfur amino acid metabolism may be accompanied by only moderately perturbed hydrogen sulfide metabolism and lends support to the hypothesis that enzymes in the transsulfuration pathway may not be the major contributors to the endogenous hydrogen sulfide pool.Linked articlesThis article is part of a themed section on Chemical Biology of Reactive Sulfur Species. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v176.4/issuetoc.

Project description:UnlabelledPyruvate, a central intermediate in the carbon fixation pathway of methanogenic archaea, is rarely used as an energy source by these organisms. The sole exception to this rule is a genetically uncharacterized Methanosarcina barkeri mutant capable of using pyruvate as a sole energy and carbon source (the Pyr(+) phenotype). Here, we provide evidence that suggests that the Pyr(+) mutant is able to metabolize pyruvate by overexpressing pyruvate ferredoxin oxidoreductase (por) and mutating genes involved in central carbon metabolism. Genomic analysis showed that the Pyr(+) strain has two mutations localized to Mbar_A1588, the biotin protein ligase subunit of the pyruvate carboxylase (pyc) operon, and Mbar_A2165, a putative transcriptional regulator. Mutants expressing the Mbar_A1588 mutation showed no growth defect compared to the wild type (WT), yet the strains lacked pyc activity. Recreation of the Mbar_A2165 mutation resulted in a 2-fold increase of Por activity and gene expression, suggesting a role in por transcriptional regulation. Further transcriptomic analysis revealed that Pyr(+) strains also overexpress the gene encoding phosphoenolpyruvate carboxylase, indicating the presence of a previously uncharacterized route for synthesizing oxaloacetate in M. barkeri and explaining the unimpaired growth in the absence of Pyc. Surprisingly, stringent repression of the por operon was lethal, even when the media were supplemented with pyruvate and/or Casamino Acids, suggesting that por plays an unidentified essential function in M. barkeri.ImportanceThe work presented here reveals a complex interaction between anabolic and catabolic pathways involving pyruvate metabolism in Methanosarcina barkeri Fusaro. Among the unexpected findings were an essential role for the enzyme pyruvate-ferredoxin oxidoreductase and an alternate pathway for synthesis of oxaloacetate. These results clarify the mechanism of methanogenic catabolism of pyruvate and expand our understanding of carbon assimilation in methanogens.

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Genetic basis for metabolism of methylated sulfur compounds in Methanosarcina species.

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