DEMONSTRATION OF CARBON-CARBON BOND-CLEAVAGE OF ACETYL COENZYME-A BY USING ISOTOPIC EXCHANGE CATALYZED BY THE CO DEHYDROGENASE COMPLEX FROM ACETATE-GROWN METHANOSARCINA-THERMOPHILA

The purified nickel-containing CO dehydrogenase complex isolated from methanogenic Methanosarcina thermophila grown on acetate is able to catalyze the exchange of [1-C-14] acetyl-coenzyme A (CoA) (carbonyl group) with CO12 as well as the exchange of [3'-P-32]CoA with acetyl-CoA. Kinetic parameters for the carbonyl exchange have been determined: K(m) (acetyl-CoA) = 200 mu-M, V(max) = 15 min-1. CoA is a potent inhibitor of this exchange (K(i) = 25-mu-M) and is formed under the assay conditions because of a slow but detectable acetyl-CoA hydrolase activity of the enzyme. Kinetic parameters for both exchanges are compared with those previously determined for the acetyl-CoA synthase/CO dehydrogenase from the acetogenic Clostridium thermoaceticum. Collectively, these results provide evidence for the postulated role of CO dehydrogenase as the key enzyme for acetyl-CoA degradation in acetotrophic bacteria.