Large quantities of purified carbon monoxide dehydrogenase (CODH) have been obtained from acetate-grown cells of Methanosarcina barkeri by scaling up the purification procedure which we developed in 1985. Studies on the pure enzyme demonstrated that it has an alpha 2 beta 2 oligomeric structure composed of subunits with molecular weights of 19,700 and 84,500. Analysis of the metal ion content indicated 1.3 plus or minus 0.3 (n=4) Ni and 15.6 plus or minus 5.6 (n=5) Fe per mol alpha 2 beta 2. Dialysis against EDTA did not reduce the content of these metals. The enzyme did not contain Co or Mo. The amino acid compositions of the subunits were determined. The enzyme did not catalyze isotopic exchange between (1- C14)acetyl-CoA and Co. Ferredoxin, FAD, FMN, 2,3,5- triphenyltetrazolium chloride (TTC), methyl viologen, and phenazine methosulfate served as electron acceptors; however, the enzyme did not reduce NAD, NADP, or the 8-hydroxy-5- deazaflavin factor F-420. A higher degree of thermostability was noted in the absence than in the presence of CO. Pure CODH from Methanococcus vannielii was also found to be an alpha 2 beta 2 oligomer composed of subunits which were similar in molecular weights to the subunits of M. barkeri CODH.
