FIGURE 4.
PPIase activity of P3H1·CRTAP·CypB complex and inhibition by cyclosporine A. The PPIase activity of the P3H1·CRTAP·CypB complex was measured in 35 mm Hepes/NaOH, pH 7.8, at 5 °C using Suc-Ala-Ala-Pro-Phe-MCA as the substrate. A, the activity was monitored by the fluorescence at 380 nm of free MCA that was cleaved by chymotrypsin. In the absence (blue) and presence of 0.0016 (green), 0.0032 (red), and 0.0064 μm (black) P3H1 complex, respectively. B, the inhibition of the PPIase activity of the P3H1·CRTAP·CypB complex by cyclosporine A (CsA) was determined by the percentage of activity remaining upon increasing concentrations of cyclosporine A. The activity ((ki − ku)/(kc − ku)) was calculated using the rate constant ki in the presence of inhibitor, uninhibited rate constant kc, and ku, the rate constant of the uncatalyzed reaction. The final P3H1·CRTAP·CypB complex concentration was 0.013 μm.