Thermodynamic Driving Forces of Redox-Dependent CPR Insertion into Biomimetic Endoplasmic Reticulum Membranes

J Phys Chem B. 2022 Mar 3;126(8):1691-1699. doi: 10.1021/acs.jpcb.1c09358. Epub 2022 Feb 16.

Abstract

Cytochrome P450 reductase (CPR) is a NADPH-dependent membrane-bound oxidoreductase found in the endoplasmic reticulum (ER) and is the main redox partner for most cytochrome P450 enzymes. Presented are the measured thermodynamic driving forces responsible for how strongly CPR partitions into a biomimetic ER with the same lipid composition of a natural ER. Using temperature-dependent fluorescence correlation spectroscopy and fluorescence single-protein tracking, the standard state free energies, enthalpies, and entropies of the CPR insertion process were all measured. The results of this study demonstrate that the thermodynamic driving forces are dependent on the redox states of CPR. In particular, the partitioning of CPRox into a biomimetic ER is an exothermic process with a small positive change in entropy, while CPRred partitioning is endothermic with a large positive change in entropy. Both resulted in negative free energies and strong association to the biomimetic ER, but the KP of CPRox insertion is measurably smaller than that of CPRred. Using this new information and known results from literature sources, we also present a phenomenological model that accounts for membrane-protein interactions, protein orientation relative to the membrane, and protein conformation as a function of the redox state.

Publication types

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

MeSH terms

  • Biomimetics
  • Cardiopulmonary Resuscitation*
  • Cytochrome P-450 Enzyme System / chemistry
  • Endoplasmic Reticulum / chemistry
  • Endoplasmic Reticulum / metabolism
  • NADPH-Ferrihemoprotein Reductase* / analysis
  • NADPH-Ferrihemoprotein Reductase* / chemistry
  • NADPH-Ferrihemoprotein Reductase* / metabolism
  • Oxidation-Reduction
  • Thermodynamics

Substances

  • Cytochrome P-450 Enzyme System
  • NADPH-Ferrihemoprotein Reductase