Conformational Landscape of Cytochrome P450 Reductase Interactions.

Manuel Sellner,Charleen G Don,Martin Smieško,André Fischer

doi:10.3390/ijms22031023

Manuel Sellner, Charleen G Don + Show 2 more

Open Access

https://doi.org/10.3390/ijms22031023

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Abstract

Oxidative reactions catalyzed by Cytochrome P450 enzymes (CYPs), which constitute the most relevant group of drug-metabolizing enzymes, are enabled by their redox partner Cytochrome P450 reductase (CPR). Both proteins are anchored to the membrane of the endoplasmic reticulum and the CPR undergoes a conformational change in order to interact with the respective CYP and transfer electrons. Here, we conducted over 22 microseconds of molecular dynamics (MD) simulations in combination with protein–protein docking to investigate the conformational changes necessary for the formation of the CPR–CYP complex. While some structural features of the CPR and the CPR–CYP2D6 complex that we highlighted confirmed previous observations, our simulations revealed additional mechanisms for the conformational transition of the CPR. Unbiased simulations exposed a movement of the whole protein relative to the membrane, potentially to facilitate interactions with its diverse set of redox partners. Further, we present a structural mechanism for the susceptibility of the CPR to different redox states based on the flip of a glycine residue disrupting the local interaction network that maintains inter-domain proximity. Simulations of the CPR–CYP2D6 complex pointed toward an additional interaction surface of the FAD domain and the proximal side of CYP2D6. Altogether, this study provides novel structural insight into the mechanism of CPR–CYP interactions and underlying conformational changes, improving our understanding of this complex machinery relevant for drug metabolism.

Highlights

Cytochrome P450 enzymes (CYPs) are the most relevant superfamily of drug-metabolizing enzymes occurring in humans, animals, plants, and microorganisms
Their interaction is based on the formation of an electron transfer (ET) complex, for which the Cytochrome P450 reductase (CPR) has to undergo a transition from a closed to an open conformation
We investigated this transition following protein–protein interactions with an atomistic model of the full-length membrane-anchored CPR, as well as the CPR–CYP2D6 complex with an integrated modeling protocol consisting of protein–protein docking, conventional molecular dynamics (MD), as well as metadynamics simulations

Summary

Introduction

Cytochrome P450 enzymes (CYPs) are the most relevant superfamily of drug-metabolizing enzymes occurring in humans, animals, plants, and microorganisms. In order to perform the wide spectrum of oxidative reactions, CYPs depend on a redox partner, the Cytochrome P450 reductase (CPR), transferring electrons from the cofactor nicotinamide adenine dinucleotide phosphate (NADPH) to the heme iron atom of the CYP. The CPR consists of the flavin mononucleotide (FMN) domain and the flavin adenine dinucleotide (FAD) domain, which bind the corresponding cofactors and transfer electrons from NADPH via FAD and FMN [5,6]. The interaction of the CPR and the CYP as well as the subsequent electron transfer (ET) require a conformational transition of the CPR from the closed to the open state, which remains incompletely understood in its details [6,8]. As the CPR is the main redox partner serving electrons to all microsomal CYPs [6], conformational adaptations to fine-tune these individual interactions are necessary

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