skip to main content
OSTI.GOV title logo U.S. Department of Energy
Office of Scientific and Technical Information

Title: Homotropic cooperativity of monomeric cytochrome P450 3A4

Abstract

Mechanistic studies of mammalian cytochrome P450s are often obscured by the phase heterogeneity of solubilized preparations of membrane enzymes. The various protein-protein aggregation states of microsomes, detergent solubilized cytochrome or a family of aqueous multimeric complexes can effect measured substrate binding events as well as subsequent steps in the reaction cycle. In addition, these P450 monooxygenases are normally found in a membrane environment and the bilayer composition and dynamics can also effect these catalytic steps. Here, we describe the structural and functional characterization of a homogeneous monomeric population of cytochrome P450 3A4 (CYP 3A4) in a soluble nanoscale membrane bilayer, or Nanodisc [Nano Lett. 2 (2002) 853]. Cytochrome P450 3A4:Nanodisc assemblies were formed and purified to yield a 1:1 ratio of CYP 3A4 to Nanodisc. Solution small angle X-ray scattering was used to structurally characterize this monomeric CYP 3A4 in the membrane bilayer. The purified CYP 3A4:Nanodiscs showed a heretofore undescribed high level of homotropic cooperativity in the binding of testosterone. Soluble CYP 3A4:Nanodisc retains its known function and shows prototypic hydroxylation of testosterone when driven by hydrogen peroxide. This represents the first functional characterization of a true monomeric preparation of cytochrome P450 monooxygenase in a phospholipid bilayer and elucidatesmore » new properties of the monomeric form.« less

Authors:
; ;  [1]
  1. UIUC
Publication Date:
Research Org.:
Argonne National Lab. (ANL), Argonne, IL (United States). Advanced Photon Source (APS)
Sponsoring Org.:
USDOE
OSTI Identifier:
1008737
Resource Type:
Journal Article
Journal Name:
Arch. Biochem. Biophys.
Additional Journal Information:
Journal Volume: 430; Journal Issue: 2004; Journal ID: ISSN 0003-9861
Country of Publication:
United States
Language:
ENGLISH
Subject:
59 BASIC BIOLOGICAL SCIENCES; 60 APPLIED LIFE SCIENCES; CYTOCHROMES; ENZYMES; FUNCTIONALS; HYDROGEN PEROXIDE; HYDROXYLATION; MEMBRANES; MICROSOMES; PHOSPHOLIPIDS; SCATTERING; SUBSTRATES; TESTOSTERONE

Citation Formats

Baas, Bradley J, Denisov, Ilia G, and Sligar, Stephen G. Homotropic cooperativity of monomeric cytochrome P450 3A4. United States: N. p., 2010. Web.
Baas, Bradley J, Denisov, Ilia G, & Sligar, Stephen G. Homotropic cooperativity of monomeric cytochrome P450 3A4. United States.
Baas, Bradley J, Denisov, Ilia G, and Sligar, Stephen G. 2010. "Homotropic cooperativity of monomeric cytochrome P450 3A4". United States.
@article{osti_1008737,
title = {Homotropic cooperativity of monomeric cytochrome P450 3A4},
author = {Baas, Bradley J and Denisov, Ilia G and Sligar, Stephen G},
abstractNote = {Mechanistic studies of mammalian cytochrome P450s are often obscured by the phase heterogeneity of solubilized preparations of membrane enzymes. The various protein-protein aggregation states of microsomes, detergent solubilized cytochrome or a family of aqueous multimeric complexes can effect measured substrate binding events as well as subsequent steps in the reaction cycle. In addition, these P450 monooxygenases are normally found in a membrane environment and the bilayer composition and dynamics can also effect these catalytic steps. Here, we describe the structural and functional characterization of a homogeneous monomeric population of cytochrome P450 3A4 (CYP 3A4) in a soluble nanoscale membrane bilayer, or Nanodisc [Nano Lett. 2 (2002) 853]. Cytochrome P450 3A4:Nanodisc assemblies were formed and purified to yield a 1:1 ratio of CYP 3A4 to Nanodisc. Solution small angle X-ray scattering was used to structurally characterize this monomeric CYP 3A4 in the membrane bilayer. The purified CYP 3A4:Nanodiscs showed a heretofore undescribed high level of homotropic cooperativity in the binding of testosterone. Soluble CYP 3A4:Nanodisc retains its known function and shows prototypic hydroxylation of testosterone when driven by hydrogen peroxide. This represents the first functional characterization of a true monomeric preparation of cytochrome P450 monooxygenase in a phospholipid bilayer and elucidates new properties of the monomeric form.},
doi = {},
url = {https://www.osti.gov/biblio/1008737}, journal = {Arch. Biochem. Biophys.},
issn = {0003-9861},
number = 2004,
volume = 430,
place = {United States},
year = {2010},
month = {11}
}