Oscillations of nitric oxide concentration in the perturbed denitrification pathway of Paracoccus denitrificans

The metabolism of nitric oxide in Paracoccus denitrificans has been studied using a Clark-type electrode. The uncoupler carbonyl cyanide m-chlorophenylhydrazone (CCCP) and the SH reagent N-ethylmaleimide, both of which released nitric oxide from cells respiring nitrite, were found to be efficient inhibitors of nitric oxide reductase activity. Control experiments with another uncoupler, pentachlorophenol, showed that the inhibitory effect of CCCP was not the result of a decrease in membrane potential. The denitrification pathway in cells with partly inhibited nitric oxide reductase, or in a reconstituted system containing purified nitric reductase and membrane vesicles, exhibited marked sustained oscillations of nitric oxide concentration. The occurrence of the oscillations was strictly dependent on the initial concentration of nitrite. The observed oscillatory kinetics is considered to reflect two regulatory signals destabilizing the denitrification pathway, namely the inhibition of nitric oxide reductase by nitric oxide and/or by nitrite.