METHANOGENESIS FROM ACETATE IN CELL-EXTRACTS OF METHANOSARCINA-BARKERI - ISOTOPE EXCHANGE BETWEEN CO2 AND THE CARBONYL GROUP OF ACETYL-COA, AND THE ROLE OF H-2

From our previous studies on the mechanism of methane formation from acetate it was known that cell extracts of acetate-grown Methanosarcina barkeri (100 000 × g supernatant) catalyze the conversion of acetyl-CoA plus tetrahydromethanopterin (=H4MPT) to methyl-H4MPT, CoA, CO2 and presumably H2. We report here that these extracts, in the absence of H4MPT, mediated an isotope exchange between CO2 ([S]0.5 v=0.2% in the gas phase) and the carbonyl group of acetyl-CoA at almost the same specific rate as the above conversion (10 nmol · min-1 · mg protein-1). Both the exchange and the formation of methyl-H4MPT were inhibited by N2O, suggesting that a corrinoid could be the primary methyl group acceptor in the acetyl-CoA C-C-cleavage reaction. Both activities were dependent on the presence of H2 (E0′=-414 mV). Ti(III)citrate (E0′=-480 mV) was found to substitute for H2, indicating a reductive activation of the system. In the presence of Ti(III)citrate it was shown that the formation of CO2 from the carbonyl group of acetyl-CoA is associated with a 1:1 stoichiometric generation of H2. Free CO, a possible intermediate in CO2 and H2 formation, was not detected. © 1990 Springer-Verlag.