High rates of substrate hydroxylation by human cytochrome P450 3A4 in reconstituted membranous vesicles: influence of membrane charge

Biochem Biophys Res Commun. 1996 Apr 16;221(2):318-22. doi: 10.1006/bbrc.1996.0593.


CYP3A4 represents the most important form of human cytochrome P450 active in drug metabolism. Reconstitution of this enzyme has in the past been a major problem. Using purified cDNA-expressed CYP3A4 incorporated into membranous vesicles made from microsomal phospholipids, rates of nifedipine and testosterone oxidation of about 60 nmol/nmol P450/min were achieved, whereas similar reconstitution into dilauroyl-phosphatidylcholine micelles was unsuccessful. A higher Vmax for nifedipine oxidation was obtained in negatively charged vesicles as compared to neutral membranes, whereas the membrane charge did not influence the Km. It is concluded that the native function of CYP3A4 requires a negatively charged microsomal membrane.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Cell Membrane / enzymology
  • Cytochrome P-450 CYP3A
  • Cytochrome P-450 Enzyme System / metabolism*
  • Humans
  • Hydroxylation
  • Membrane Lipids / metabolism
  • Membrane Potentials
  • Mixed Function Oxygenases / metabolism*
  • NADH, NADPH Oxidoreductases / metabolism
  • NADPH-Ferrihemoprotein Reductase
  • Nifedipine / metabolism
  • Rats
  • Substrate Specificity
  • Testosterone / metabolism


  • Membrane Lipids
  • Testosterone
  • Cytochrome P-450 Enzyme System
  • Mixed Function Oxygenases
  • CYP3A protein, human
  • Cytochrome P-450 CYP3A
  • CYP3A4 protein, human
  • NADH, NADPH Oxidoreductases
  • NADPH-Ferrihemoprotein Reductase
  • Nifedipine