Homotropic and heterotropic cooperativity of CYP3A4 and drug‐drug interactions

Abstract

The allosteric behaviour of monomeric human cytochrome P450 CYP3A4 solubilized in nanoscale lipid bilayers was systematically studied in order to understand the mechanisms of homotropic and heterotropic cooperativity. The ability of CYP3A4 to simultaneously bind multiple drug molecules alters their metabolism and gives rise to drug‐drug interactions. Using global analysis of several data sets obtained in different experiments under identical conditions, we are able to represent the overall cooperative properties of CYP3A4 and the functional contributions of each binding intermediate. We show that the homotropic cooperativity observed in testosterone (TS) hydroxylation is due to negligible product formation when only one substrate molecule is bound, and not to any significant conditional interaction free energies of substrate binding. A two‐dimensional binding isotherm is obtained for the heterotropic interactions between TS and alpha‐naphthoflavone, allowing complete deconvolution of homotropic and heterotropic cooperativity in this important system. . Supported by NIH grants GM 33775 and GM 31756 to S.G.Sligar.