Methane synthesis by membrane vesicles and a cytoplasmic cofactor isolated from Methanobacterium thermoautotrophicum.
Methanobacterium thermoautotrophicum when grown on ordinary culture medium has a tough cell wall which is lysozyme-resistant and difficult to disrupt by physical means. The cell wall, however, can be weakened by the addition of D-sorbitol to the growth medium and the organisms form protoplasts after lysozyme addition. This technique allowed the isolation of two types of intracellular small vesicles: (a) isolated by disruption of the total cell population (lysozyme-sensitive and lysozyme-resistant cells) by ultrafrequency sound and (b) isolated by osmotic lysis of protoplasts. For the first time, a small vesicle fraction isolated as in (a) was capable of synthesizing methane from CO2 and H2 without cytoplasm. There was, however, an absolute requirement for a small, heat-stable, oxygen-sensitive cofactor which was isolated from the cytoplasm. Methane synthesis with this vesicle fraction was inhibited by the detergent deoxycholate, and by the protonophores 2,4-dinitrophenol and carbonyl cyanide m-chlorophenylhydrazone. Mg2+-ATPase appeared to be located on the outer or cytoplasmic surface of the small vesicle fraction isolated as in (b). The results were consistent with a previously made suggestion [Sauer, Erfle & Mahadevan (1981) J. Biol. Chem. 256, 9843-9848] that the interior of the small intracellular vesicles becomes acid during methane synthesis.
D002458:Cell Fractionation; D003067:Coenzymes / Q000378:metabolism; D003470:Culture Media; D003593:Cytoplasm / Q000378:metabolism; D019605:Euryarchaeota / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D007425:Intracellular Membranes / Q000378:metabolism; D008697:Methane / Q000096:biosynthesis*; D008854:Microscopy, Electron; D013010:Sonication; D013012:Sorbitol; D014475:Uncoupling Agents / Q000494:pharmacology
D003067:Coenzymes; D003470:Culture Media; D014475:Uncoupling Agents; D013012:Sorbitol; D008697:Methane