Active [FeFe] hydrogenases can be obtained by expressing the unmaturated enzyme in Escherichia coli followed by incubation with a synthetic precursor of the binuclear [2Fe] subcluster, namely: [NEt4]2[Fe2(adt)(CO)4(CN)2] (adt = [S-CH2-NH-CH2-S](2-)). The binuclear subsite Fe2(adt)(CO)3(CN)2 is attached through a bridging cysteine side chain to a [4Fe-4S] subcluster already present in the unmaturated enzyme thus yielding the intact native "H-cluster". We present FTIR electrochemical studies of the [FeFe] hydrogenase from Chlamydomonas reinhardtii, CrHydA1, maturated with the precursor of the native cofactor [Fe2(adt)(CO)4(CN)2](2-) as well as a non-natural variant [Fe2(pdt)(CO)4(CN)2](2-) in which the bridging amine functionality is replaced by CH2. The obtained active enzyme CrHydA1(adt) shows the same redox states in the respective potential range as observed for the native system (E(ox/red) = -400 mV, E(red/sred) = -470 mV). For the Hox → Hred transition the reducing equivalent is stored on the binuclear part, ([4Fe-4S](2+)Fe(II)Fe(I) → [4Fe-4S](2+)Fe(I)Fe(I)), while the Hred → Hsred transition is characterized by a reduction of the [4Fe-4S] part of the H-cluster ([4Fe-4S](2+)Fe(I)Fe(I) → [4Fe-4S](+)Fe(I)Fe(I)). A similar transition is reported here for the CO inhibited state of the H-cluster: ([4Fe-4S](2+)Fe(I)Fe(II)CO → [4Fe-4S](+)Fe(I)Fe(II)CO). An FTIR electrochemical study of the inactive variant with the pdt ligand, CrHydA1(pdt), identified two redox states H(pdt)-ox and H(pdt)-"red". Both EPR and FTIR spectra of H(pdt)-ox are virtually identical to those of the H(adt)-ox and the native Hox state. The H(pdt)-"red" state is also characterized by a reduced [4Fe-4S] subcluster. In contrast to CrHydA1(adt), the H(pdt)-ox state of CrHydA1(pdt) is stable up to rather high potentials (+200 mV). This study demonstrates the distinct redox coupling between the two parts of the H-cluster and confirms that the [4Fe-4S]H subsite is also redox active and as such an integral part of the H-cluster taking part in the catalytic cycle.