Cellular ATP levels and nitrogenase switchoff upon oxygen stress in chemostat cultures of Azotobacter vinelandii

When Azotobacter vinelandii, growing diazotrophically in chemostat culture, was subjected to sudden increases in the ambient oxygen concentration (oxygen stress), nitrogenase activity was switched off and cellular ATP pools decreased at rates depending on the stress level. Following a fast decrease, the ATP pool approached a lower level. When the stress was released, these effects were reversed. The reversible decrease of the ATP pool upon oxygen stress could also be observed with cultures assimilating ammonium and, at the same time, fixing dinitrogen because of growth at a high C/N ratio but not with cultures growing only at the expense of ammonium. When strains OP and UW136 of A. vinelandii were subjected to long-term increases in ambient oxygen, the sizes of cellular ATP pools eventually started to increase to the level before stress and diazotrophic growth resumed. The cytochrome d-deficient mutant MK5 of A. vinelandii, however, impaired in aerotolerant diazotrophic growth, was unable to recover from stress on the basis of its ATP pool. The results suggest that adaptation to higher ambient oxygen depends on increased ATP synthesis requiring increased electron flow through the entire respiratory chain, which is possible only in combination with the more active, yet possibly uncoupled, branch terminated by cytochrome d. It is proposed that the decrease of the cellular ATP level under oxygen stress resulted from the increased energy and electron donor requirement of nitrogenase in reacting with oxygen.