Summary: Initial attempts to grow N-fixing autotrophically on H, CO, O and N in a closed system yielded variable results. Poor growth was found to be due to rapid O depletion and the requirement for an agar surface. In a closed system, CH reduction assays could not be carried out due to complete consumption of H. Hence a flow-through culturing technique was developed to supply gases at a constant partial pressure and to perform CH reduction assays in a continuous flow system. Hydrogenase of autotrophic was not inhibited by CH, even at 0·5 atm, and the of hydrogenase for H was approximately 0·15 atm. The effects of O and H on CH reduction were examined, using the flow-through assay system. The rate of CH reduction decreased below 0·074 atm H, suggesting that ATP and reductant supply were limiting the nitrogenase activity.


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