Site saturation mutagenesis demonstrates a central role for cysteine 298 as proton donor to the catalytic site in CaHydA [FeFe]-hydrogenase.

[FeFe]-hydrogenases reversibly catalyse molecular hydrogen evolution by reduction of two protons. Proton supply to the catalytic site (H-cluster) is essential for enzymatic activity. Cysteine 298 is a highly conserved residue in all [FeFe]-hydrogenases; moreover C298 is structurally very close to the H-cluster and it is important for hydrogenase activity.