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. Author manuscript; available in PMC: 2014 May 22.
Published in final edited form as: Mol Cell Endocrinol. 2012 Oct 23;371(0):100–106. doi: 10.1016/j.mce.2012.10.013

Fig. 3.

Fig. 3

Spectral analysis of CYP11A interaction with clobenpropit. Numbers above or below the spectra indicate the wavelengths of absorption maxima or minima. A, absolute spectra; black and gray lines represent the spectra of cholesterol-free (black) and cholesterol-bound (gray) CYP11A1 in the absence (solid line) and presence (dashed line) of clobenpropit. The concentration of CYP11A1 was 0.4 μM; the concentrations of cholesterol and clobenpropit were 4 μM and 150 μM, respectively. These ligand concentrations are equal to 20 Kd of cholesterol (0.2 μM) and 8 Kd of clobenpropit (18 μM) for cholesterol-free CYP11A1. B–E, difference spectra of 0.4 μM CYP11A1 titrated under different assay conditions. B, titration with clobenpropit when no cholesterol or DLPC was added to the buffer (40 mM KPi, pH, 7.2, containing 1 mM EDTA). C, titration with cholesterol when no DLPC was added to the buffer. D, titration with clobenpropit in the presence of cholesterol when no DLPC was added to the buffer. E, titration with clobenpropit in the presence of cholesterol and DLPC. Under the concentrations used, the absorption of clobenpropit in the visible region is up to 0.003 absorbance units.