CELLULAR-REGULATION OF NITRATE UPTAKE IN DENITRIFYING FLEXIBACTER-CANADENSIS

Nitrate uptake and its regulation were investigated using an ion-speci fic nitrate electrode for denitrifying Flexibacter canadensis under an aerobic conditions. Glucose supported a greater rate of nitrate uptake than did glycerol, glutamate, lactose, cellobiose, or ethanol. Nitrat e uptake closely approximated Michaelis-Menten kinetics; the estimated K-s(glucose) and apparent K-m(nitrate) for nitrate uptake were 21 and 44 mu M, respectively. Nitrate disappearance was correlated with nitr ite accumulation, and nitrate had an inhibitory effect on nitrite redu ction. Oxygen inhibition of nitrate uptake increased as the percent ai r saturation increased, and reversed readily as the percent air satura tion decreased. The minimal air saturation showing inhibition of nitra te uptake was about 2-4%. Azide and cyanide completely inhibited nitra te uptake. No nitrate uptake was observed in cells grown in the presen ce of 1 or 5 mM tungstate (no added molybdate). When molybdate (100-20 0 mu M) was present in the medium, nitrate uptake was exhibited by org anisms grown with 1 mM, but not with 5 mM, tungstate, indicating that nitrate uptake was dependent on the presence of an active nitrate redu ctase, and that competition between tungsten and molybdenum occurred d uring the formation of nitrate reductase. Nitrite production from nitr ate by whole cells but not cell-free extracts was inhibited by 2,4-din itrophenol and carbonyl cyanide m-chlorophenylhydrazone, indicating th at nitrate and (or) nitrite transport depended upon the electrochemica l proton gradient.