FIGURE 6.

Relative role of Thr²⁶⁴, Ser⁴²⁰, and Ser⁴⁷⁸ phosphorylation in CYP3A4 ubiquitination. A, PhosphorImager analyses of time-dependent ubiquitination of CuOOH-inactivated CYP3A4wt and its triple mutant (TSSm) in the presence or absence of PKA/PKC in the reaction. B, corresponding time-dependent ubiquitination of CuOOH-inactivated CYP3A4 triple mutant in the presence or absence of PKA/PKC in the reaction. Note that in B, the reaction mixture aliquots used were half the size of those in A. C, PhosphorImager analyses of CYP3A4 immunoprecipitates from ubiquitination reactions and corresponding controls carried out at 90 min containing either CuOOH-inactivated CYP3A4wt or TSSm and in the presence or absence of PKA/PKC in the reaction. Data from a typical experiment are shown. Each experiment was conducted in its entirety at the least three separate times. The color wheel intensity code is as follows: black > dark blue > light blue > green > yellow > orange > red > magenta > white.