Methanobacterium wolfei is a thermophilic methanogenic archaeon which requires tungsten or molybdenum for growth. We have found that the organism contains two formylmethanofuran dehydrogenases, one of which is a tungsten enzyme. Indirect evidence indicates that the other formylmethanofuran dehydrogenase is a molybdenum enzyme. The tungsten enzyme was purified and characterized. The native enzyme had an apparent molecular mass of 130 kDa. SDSjPAGE revealed a composition of three subunits of apparent molecular mass 35, 51 and 64 kDa, the Nterminal amino acid sequences of two of which were determined. 0.3 -0.4 mol tungsten/mol enzyme was found but no molybdenum. The pterin cofactor was identified as molybdopterin guanine dinucleotide. The purified enzyme exhibited a specific activity of 8.3 pmol . min-' . mg protein-' and an apparent K, for formylmethanofuran and methylviologen of 13 pM and 0.4 mM, respectively.The optimum temperature for activity was 65°C. At 40 -6O"C, the rate increased with a Qlo of 1.9; the activation energy of the reaction was 45 kJ/mol. The enzyme was found to require potassium ions for thermostability. The oxygen-sensitive enzyme was not inactivated by cyanide.