DISTINCTION BETWEEN THE 2 BASIC MECHANISMS OF CATION-TRANSPORT IN THE CARDIAC NA+-CA-2+ EXCHANGE SYSTEM

In order to distinguish between the Ping-Pong and sequential mechanisms of cation transport in the cardiac Na+-Ca2+ exchange system, the initial rates of the Naidependent 45Ca uptake (t = 1 s) were measured in reconstituted proteoliposomes, loaded with a Ca chelator. Under “zero-trans” conditions ([Na]o = [Ca]i = 0) at a fixed [Na]i = 10–160 mM with varying [45Ca]o = 2.5–122 µM for each [Na]i, the Km and Vmax values increased from 7.7 to 33.5 µM and from 2.3 to 9.0 nmol•mg‒1•S‒1, respectively. The Vmax/Km, values show a ±2-10% deviation from the average value of 0.274 nmol•mg‒1•S‒1•µM‒1 over the whole range of [Na]i. These deviations are within the standard error of Vmax (±3-7%), Km(±l 1–17%), and Vmax/Km (±11-19%). This suggests that, under conditions in which Vmax and Km are [Na]i dependent and vary 4–5-fold, the Vmax/Km values are constant within the experimental error. In the presence of K+-valinomycin the Vmax/Km values are 0.85 ± 0.17 and 1.08 ± 0.18 nmol•mg‒1•S‒1•µM‒1 at [Na]i = 20 and 160 mM, respectively, suggesting that under conditions of “short circuit” of the membrane potential the Vmax/Km values still exhibit the [Na]i independence. At a very low fixed [45Ca]o = 1.1 µM with varying [Na]i = 10–160 mM, the initial rates were found to be [Na]i independent. At a high fixed [45Ca]o = 92 µM the initial rates show a sigmoidal dependence on the [Na]i with Vmax = 13.8 nmol•mg‒1•S‒1,KmNa = 21 mM, and Hill coefficient nH = 1.5. The presented data support a Ping-Pong (consecutive) mechanism of cation transport in the Na+-Ca2+ exchanger. © 1990, American Chemical Society. All rights reserved.