FIGURE 4.

Effect of vanadate on the level of calcium occluded by the PMCA. Occluded calcium was determined for increasing Ca²⁺ concentrations, with (triangles) or without (circles and squares) 50 μm (VO₄)³⁻ (see “Experimental Procedures” for details). Determinations where performed in the absence (A) and in the presence (B) of 2 mm ATP, except for a control experiment (closed squares), where media contained 25 μm ATP and 50 μm La^III. The continuous lines represent the fit of a Michaelis-Menten-like function of [Ca²⁺] to the experimental data. Best fitting values of the parameters were as follows. A, closed circles, Ca_occ,max = 0.058 ± 0.002 nmol of ⁴⁵Ca (mg of protein)⁻¹, K_0.5 = 19.7 ± 1.9 μm; closed triangles, Ca_occ,max = 0.046 ± 0.001 nmol of ⁴⁵Ca (mg of protein)⁻¹, K_0.5 = 17.4 ± 1.4 μm. B, open circles, Ca_occ,max = 0.064 ± 0.001 nmol of ⁴⁵Ca (mg of protein)⁻¹, K_0.5 = 17.2 ± 0.7 μm; open triangles, Ca_occ,max = 0.063 ± 0.005 nmol of ⁴⁵Ca (mg of protein)⁻¹, K_0.5 = 19.1 ± 4 μm; closed squares, Ca_occ,max = 0.084 ± 0.003 nmol of ⁴⁵Ca (mg of protein)⁻¹, K_0.5 = 6.8 ± 0.9 μm.