CHARACTERIZATION OF A CO-HETERODISULFIDE OXIDOREDUCTASE SYSTEM FROM ACETATE-GROWN METHANOSARCINA-THERMOPHILA

During the methanogenic fermentation of acetate by Methanosarcina ther mophila, the CO dehydrogenase complex cleaves acetyl coenzyme A and ox idizes the carbonyl group (or CO) to CO2, followed by electron transfe r to coenzyme M (CoM)-S-S-coenzyme B (CoB) and reduction of this heter odisulfide to HS-CoM and HS-CoB (A. P. Clements, R. H. White, and J. G . Ferry, Arch. Microbiol. 159:296-300, 1993). The majority of heterodi sulfide reductase activity was present in the soluble protein fraction after French pressure cell lysis. A CO:CoM-S-S-CoB oxidoreductase sys tem from acetate-grown cells was reconstituted with purified CO dehydr ogenase enzyme complex, ferredoxin, membranes, and partially purified heterodisulfide reductase. Coenzyme F-420 (F-420) was not required, an d CO:F-420 oxidoreductase activity was not detected in cell extracts. The membranes contained cytochrome b that was reduced with CO and oxid ized with CoM-S-S-CoB. The results suggest that a novel CoM-S-S-CoB re ducing system operates during acetate conversion to CH4 and CO2. In th is system, ferredoxin transfers electrons from the CO dehydrogenase co mplex to membrane-bound electron carriers, including cytochrome b, tha t are required for electron transfer to the heterodisulfide reductase. The cytochrome b was purified from solubilized membrane proteins in a complex with six other polypeptides. The cytochrome was not reduced w hen the complex was incubated with H-2 or CO, and H-2 uptake hydrogena se activity was not detected; however, the addition of CO dehydrogenas e enzyme complex and ferredoxin enabled the CO-dependent reduction of cytochrome b.