Project description:Methane-producing archaea are key organisms in the anaerobic carbon cycle. These organisms, also called methanogens, grow by converting substrate to methane gas in a process called methanogenesis. Previous research showed that reduction of the terminal electron acceptor is the rate-limiting step in methanogenesis by Methanosarcina acetivorans. In order to gain insight into how the cells sense and respond to availability of the terminal electron acceptor, we designed an experiment to deplete cells of the essential terminal oxidase enzyme, HdrED. We found that depletion of HdrED in vivo results in higher abundance of transcripts for methyltransferases (mtaC2, mtaB3, mtaC3), coenzyme B biosynthesis, C1 metabolism, and pyrimidine compounds. In most cases, these changes were distinct from transcript abundance changes observed during the transition from exponential growth to stationary phase cultures. These data implicate MsrC (MA4383) in CoM-S-S-CoB heterodisulfide sensing and indicate cells have a specific mechanism to sense intracellular ratio of CoM-S-S-CoB, coenzyme M and coenzyme B thiols and further suggest transcripts encoding translation and methanogenesis functions are controlled by feed-forward regulation depending on substrate availability.
Project description:Our goal is to convert methane efficiently into liquid fuels that may be more readily transported. Since aerobic oxidation of methane is less efficient, we focused on anaerobic processes to capture methane, which are accomplished by anaerobic methanotrophic archaea (ANME) in consortia. However, no pure culture capable of oxidizing and growing on methane anaerobically has been isolated. In this study, Methanosarcina acetivorans, an archaeal methanogen, was metabolically engineered to take up methane, rather than to generate it. To capture methane, we cloned the DNA coding for the enzyme methyl-coenzyme M reductase (Mcr) from an unculturable archaeal organism from a Black Sea mat into M. acetivorans to effectively run methanogenesis in reverse. The engineered strain produces primarily acetate, and our results demonstrate that pure cultures can grow anaerobically on methane.
