Fig. 4.

Experimental transport kinetics for wild-type NCX_Mj and D240 mutants. To characterize the exchange of Na⁺/Ca²⁺ and Ca²⁺/Ca²⁺ mediated by NCX_Mj, E. coli-derived vesicles in which the protein had been overexpressed were loaded with either Na⁺ (160 mM) or Ca²⁺ (250 µM), and diluted in an assay medium containing 20–2,000 µM ⁴⁵CaCl₂. Initial ⁴⁵Ca²⁺-uptake rates (t = 5 s) were measured as described in Methods. (A) V_max and (B) K_m values for Na⁺/Ca²⁺ and Ca²⁺/Ca²⁺ exchange were obtained by fitting the experimental data to the Michaelis equation. The bars represent mean ± SE values obtained from three independent experiments.