The effect of inhibitors on the oxygen kinetics of terminal oxidases of Acanthamoeba castellanii

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Abstract
Respiration of growing cultures of A. castellanii is inhibited less than 60% by azide (35 mM); the respiration of early-exponential-phase cultures differs from that of late-exponential-phase cultures in being stimulated by up to 120% by low concentrations (< 1 mM) of this inhibitor. Azide (0.5 mM) plus 1 mM-salicylhydroxamic acid gives 80% inhibition of respiration in early- or late-exponential-phase cultures. Lineweaver-Burk plots of 1/v against 1/[O2] for growing and stationary-phase cultures give values of < 1 .mu.M for the apparent Km for O2. These values are not significantly altered when determined in the presence of 1 mM-salicylhydroxamic acid. Higher values (> 7 .mu.M) for apparent Km values for O2 were obtained in the presence of azide, which gives nonlinear Lineweaver-Burk plots. Competitive inhibition of respiration by CO occurs with Ki 2.4 .mu.M. The results are discussed in terms of the presence of 3 terminal oxidases in this organism, namely 2 oxidases with high affinities for O2 (cytochrome c oxidase of the main phosphorylating electron-transport chain and the salicylhydroxamic acid-sensitive oxidase) and a 3rd oxidase with a low affinity for O2, sensitive to inhibition by cyanide but not by azide or salicylhydroxamic acid. The relative contributions to O2 utilization by these oxidases change during the growth of a batch culture.