Kinetics and ion specificity of Na(+)/Ca(2+) exchange mediated by the reconstituted beef heart mitochondrial Na(+)/Ca(2+) antiporter.

Abstract

The Na(+)/Ca(2+) antiporter was purified from beef heart mitochondria and reconstituted into liposomes containing fluorescent probes selective for Na(+) or Ca(2+). Na(+)/Ca(2+) exchange was strongly inhibited at alkaline pH, a property that is relevant to rapid Ca(2+) oscillations in mitochondria. The effect of pH was mediated entirely via an effect on the K(m) for Ca(2+). When present on the same side as Ca(2+), K(+) activated exchange by lowering the K(m) for Ca(2+) from 2 to 0.9 microM. The K(m) for Na(+) was 8 mM. In the absence of Ca(2+), the exchanger catalyzed high rates of Na(+)/Li(+) and Na(+)/K(+) exchange. Diltiazem and tetraphenylphosphonium cation inhibited both Na(+)/Ca(2+) and Na(+)/K(+) exchange with IC(50) values of 10 and 0.6 microM, respectively. The V(max) for Na(+)/Ca(2+) exchange was increased about fourfold by bovine serum albumin, an effect that may reflect unmasking of an autoregulatory domain in the carrier protein.