Ech hydrogenase fromMethanosarcina barkeriis a member of a distinct group of membrane-bound [NiFe] hydrogenases with sequence similarity to energy-conserving NADH:quinone oxidoreductase (complex I). The sequence of the enzyme predicts the binding of three [4Fe-4S] clusters, one by subunit EchC and two by subunit EchF. Previous studies had shown that two of these clusters could be fully reduced under 10 5 Pa of H2at pH 7 giving rise to two distinct S½ electron paramagnetic resonance (EPR) signals, designated as the g¼ and theg¼ signal
