The [FeFe]-hydrogenases catalyze the uptake and evolution of hydrogen with unmatched speed at low over-potential. However, oxygen induces the degradation of the unique [6Fe-6S] cofactor within the active site, termed the H-cluster. We used X-ray structural analyses to determine possible modes of irreversible oxygen-driven inactivation. To this end, we exposed crystals of the [FeFe]-hydrogenase CpI from Clostridium pasteurianum to oxygen and quantitatively investigated the effects on the H-cluster structure over several time points using multiple data sets, while correlating it to decreases in enzyme activity. Our results reveal the loss of specific Fe atoms from both the diiron (2Fe(H)) and the [4Fe-4S] subcluster (4Fe(H)) of the H-cluster. Within the 2Fe(H), the Fe atom more distal to the 4Fe(H) is strikingly more affected than the more proximal Fe atom. The 4Fe(H) interconverts to a [2Fe-2S] cluster in parts of the population of active CpIADT, but not in crystals of the inactive apoCpI initially lacking the 2Fe(H). We thus propose two parallel processes: dissociation of the distal Fe atom and 4Fe(H) interconversion. Both pathways appear to play major roles in the oxidative damage of [FeFe]-hydrogenases under electron-donor deprived conditions probed by our experimental setup.