NADH FORMATION BY NA+-COUPLED REVERSED ELECTRON-TRANSFER IN KLEBSIELLA-PNEUMONIAE

Citrate is fermented by Klebsiella pneumoniae to 2 acetate, 0.5 formate and 1.2 CO2. The formation of < 1 formate and > 1 CO2 per citrate can be accounted for by the oxidation of formate to CO2 in order to provide reducing equivalents for the assimilation of citrate into cell carbon. A membrane-bound electron transport chain is apparently involved in NADH synthesis by these cells. The electrons from formate oxidation to CO2 are used to reduce ubiquinone to ubiquinol by membrane-bound formate dehydrogenase and ubiquinol further delivers its electrons to NAD+, if this endergonic reaction is powered by DELTA-mu approximately Na+. The endogenous NADH level of K. pneumoniae cells thus increased in the presence of formate in response to a DELTA-pNa+ > -100 mV. NADH formation was completely abolished in the presence of oxygen or after addition of hydroxyquinoline-N-oxide, a specific inhibitor of the Na+-translocating NADH:ubiquinone oxidoreductase. The increase of endogenous NADH was dependent on the DELTA-pNa+ applied to the cells. Inverted membrane vesicles of K. pneumoniae catalysed the reduction of NAD+ to NADH with formate as electron donor after application of DELTA-mu approximately Na+ of about 120 mV consisting of DELTA-pNa+ of 60 mV and DELTA-psi of the same magnitude. Neither the DELTA-pNa+ nor the DELTA-psi of this size alone was sufficient to drive the endergonic reaction. Strictly anaerobic conditions were required for NADH formation and hydroxyquinoline-N-oxide completely inactivated the reaction. It is suggested that NADH formation by reversed DELTA-mu approximately Na+-coupled electron transfer in these cells is an essential requirement for the synthesis of cell material from citrate.