Abdul Halim, Mohd FaridDay, LeslieCosta, Kyle2020-12-012020-12-012020-12-01https://hdl.handle.net/11299/217228The files contain the raw mass spectromery data of independent samples (biological replicates) of Elution fraction from His-purification of wild-type, MvhA-His, or HdrB-His strains.Hydrogenotrophic methanogens produce CH4 using H2 as electron donor to reduce CO2. In the absence of H2, many are able to use formate or alcohols as alternate electron donors. Methanogens from the order Methanomicrobiales are capable of growth with H2, but many lack genes encoding hydrogenases that are typically found in other hydrogenotrophic methanogens. In an effort to better understand electron flow in methanogens from the Methanomicrobiales, we undertook a genetic and biochemical study of heterodisulfide reductase (Hdr) in Methanoculleus thermophilus. Hdr catalyzes an essential reaction by coupling the first and last steps of methanogenesis through flavin-based electron bifurcation. Hdr from M. thermophilus co-purified with formate dehydrogenase (Fdh) and only displayed activity when formate was supplied as an electron donor. We found no evidence of an Hdr associated hydrogenase, and H2 could not function as an electron donor, even with Hdr purified from cells grown on H2. We found that cells catalyze a formate hydrogenlyase activity that is likely essential for generating the formate needed for the Hdr reaction. Together, these results highlight the importance of formate as an electron donor for methanogenesis and suggest the ability to use formate is closely integrated into the methanogenic pathway in organisms from the order Methanomicrobiales.CC0 1.0 UniversalHeterodisulfide reductaseFormate dehydrogenaseMethanoculleus thermophilusDatasethttps://doi.org/10.13020/g0xp-gf10