Kinetics of mitochondrial calcium transport. II. A kinetic description of the sodium-dependent calcium efflux mechanism of liver mitochondria and inhibition by ruthenium red and by tetraphenylphosphonium

Abstract

Sodium-dependent calcium efflux from rat liver mitochondria has been studied as a function of mitochondrial calcium loads (2 to 40 nmol/mg) and extramitochondrial sodium concentrations (5 to 40 mM). The resulting data can be fit to a terreactant model which exhibits simultaneous kinetics (i.e. both sodium and calcium must be bound simultaneously for transport to occur). The Hill coefficients for the calcium and sodium dependences were 1.0 +/- 0.1 and 2.0 +/- 0.2, respectively. The cooperativity of the sodium dependence allows the terreactant model to be reduced to a bireactant model in which the sodium concentration only appears mathematically as the square of the sodium concentration. The data then fit the relationship (Formula: see text) The experimentally determined value of Vmax is found to be 2.6 +/- 0.5 nmol/mg/min, and the load of calcium (KCa) and concentration of sodium (KNa) necessary to stimulate the efflux to half its maximal calcium-dependent activity and sodium-dependent activity, respectively, were 8.1 +/- 1.4 nmol of Ca2+/mg and 9.4 +/- 0.6 mM Na+. This sodium-dependent calcium efflux from liver mitochondria was inhibited by magnesium, by ruthenium red, and by tetraphenylphosphonium. Fifty percent inhibition was obtained at 1.0-1.5 mM magnesium, at 12 nmol of ruthenium red/mg of protein, and at 0.2 microM tetraphenylphosphonium.