Abstract 1. The oxyhydrogen reaction of Anacystis nidulans was studied manometrically and polarographically in whole cells and in cell-free preparations; the activity was found to be associated with the particulate fraction. 2. Besides O 2, the isolated membranes reduced artificial electron acceptors of positive redox potential; the reactions were unaffected by O 2 levels <10–15%; aerobically the artificial acceptors were reduced simultaneously with O 2. 3. H 2-supported O 2 uptake was inhibited by CO, KCN and 2- n-heptyl-8-hydroxyquinoline- N-oxide. Inhibition by CO was partly reversed by strong light. Uncouplers stimulated the oxyhydrogen reaction. 4. The kinetic properties of O 2 uptake by isolated membranes were the same in presence of H 2 and of other respiratory substrates. 5. Low rates of H 2 evolution by the membrane preparations were found in presence of dithionite; methyl viologen stimulated the reaction. 6. The results indicate that under certain growth conditions Anacystis synthesizes a membrane-bound hydrogenase which appears to be involved in phosphorylative electron flow from H 2 to O 2 through the respiratory chain.