Studies on the carbon monoxide dehydrogenase enzyme complex present in acetate-grown Methanosarcina thermophila strain TM-1

The carbon monoxide dehydrogenase complex was purified from acetate-grown Methanosarcina thermophila. This complex made up greater than 10% of the cellular protein and the native enzyme formed aggregates with a Mr of approximately 1,000,000. The enzyme contained five subunits of different molecular weight suggesting a multifunctional enzyme complex. The electron paramagnetic resonance spectrum of CO-reduced enzyme at 113K contained g values of 2.073, 2.049, and 2.028. Isotope substitution with 61Ni, 57Fe, or 13CO resulted in broadening of the spectrum consistent with a Ni-Fe-C spin-coupled complex. Acetyl-CoA caused a perturbation of the signal that was not caused by acetyl-phosphate or mercaptoethanol indicating acetyl-CoA is a physiological substrate. Cell extracts from acetate-grown M. thermophila contained CO-oxidizing:H2-evolving activity 16-fold greater than extracts of methanol-grown cells. CO-oxidizing:H2-evolving activity was reconstituted upon combination of: (i) CO dehydrogenase complex, (ii) a ferredoxin, and (iii) purified membranes with associated hydrogenase and b-type cytochrome