Figure 4.

Absence of positional isotope exchange with purified liposome-reconstituted P-gp. 31 nM P-gp was incubated with 2 mM γ-¹⁸O₄-ATP, 50 mM Tris·HCl, 15 mM NH₄Cl, 5 mM MgSO₄, 2.5 mM EGTA, 0.02 wt % NaN₃, 2 mM TCEP, pH 7.4 in the presence of 50 μM nicardipine. The specific activity for the Nicardipine-stimulated P-gp was 1660 nmol of P_i/min/mg protein, at 2 mM ATP concentration, 37 °C. The reactions were conducted as described in the text, and ³¹P NMR spectra were recorded at the times indicated. The incubation was also conducted in the presence of 50 μM loperamide or 50 μM verapamil, or 20 μM cyclosporin A (right-hand side inset).