Exploring the structure characteristics and major channels of cytochrome P450 2A6, 2A13, and 2E1 with pilocarpine

Abstract

The majority of cytochromes P450 play a critical role in metabolism of endogenous and exogenous substrates, some of its products are carcinogens. Therefore, inhibition of P450 enzymes activity can promote the detoxification and elimination of chemical carcinogens. In this study, molecular dynamics (MD) simulations and adaptive steered molecular dynamics (ASMD) simulations were performed to explore the structure features and channel dynamics of three P450 isoforms 2A6, 2A13, and 2E1 bound with the common inhibitor pilocarpine. The binding free energy results combined with the PMF calculations give a reasonable ranking of binding affinity, which are consistent with the experimental data. Our results uncover how a sequence divergence of different CYP2 enzymes causes individual variations in major channel selections. On the basis of channel bottleneck and energy decomposition analysis, we propose a gating mechanism of their respective major channels in three enzymes, which may be attributed to a reversal of Phe209 in CYP2A6/2A13, as well as the rotation of Phe116 and Phe298 in CYP2E1. The hydrophobic residues not only make strong hydrophobic interactions with inhibitor, but also act as gatekeeper to regulate the opening of channel. The present study provides important insights into the structure-function relationships of three cytochrome P450s and the molecular basis for development of potent inhibitors.

Molecular Dynamics Simulation Framework to Probe the Binding Hypothesis of CYP3A4 Inhibitors

“…The availability of CYP crystallographic structures and the use of various in silico approaches including docking, molecular dynamic simulations and the MM(GB/PB)SA method for the investigation of ligand binding have been described extensively in literature to further supplement the understanding of the underlying mechanism of ligand recognition by the CYP isoforms [36,87,88,89,90,91,92]. The MM(GB/PB)SA method is commonly used for the calculation of binding free energies based on the MD simulation trajectories [76].…”

Tryptophan-75 Is a Low-Energy Channel-Gating Residue that Facilitates Substrate Egress/Access in Cytochrome P450 2D6

“…Like most CYPs, CYP2D6 has a buried active site to isolate the active site heme from bulk solvent and to prevent uncoupling reactions (Wade et al, 2004). Understanding substrate and inhibitor access to the CYP2D6 active site is of interest due to applications for rational drug design, artificial catalyst design and for better understanding of enzyme structure and function (Dubey et al, 2016;Yu et al, 2016;Fan et al, 2018).…”

“…Different roles have been proposed for amino acids that line channels including 'tunnel-gating' and 'guide rails' (Li et al, 2005;Fishelovitch et al, 2009;Zawaira et al, 2011;Shen et al, 2012;Urban et al, 2015;Dubey et al, 2016;Yu et al, 2016;Albertolle et al, 2018;Fan et al, 2018;Fischer and Smieško, 2019). Tunnel-gating refers to the flexible amino acid side chain barriers within the active site or substrate channels that control access to the active site, while amino acids described as guide rails function via electrostatic interactions to shuttle ligands along channels from the protein's surface to the buried active site.…”

“…DMD Fast Forward. Published on December 29, 2020 as DOI: 10.1124/dmd.120.000274 at ASPET Journals on January 3, 2021 dmd.aspetjournals.org Downloaded from 7 the role of gating residues in impeding ligand movement through channels (Ludemann et al, 2000a;Ludemann et al, 2000b;Fishelovitch et al, 2009;Berka et al, 2012;Dubey et al, 2016;Yu et al, 2016;Fan et al, 2018). Most, but not all, of the gating residues identified in other CYPs (e.g.…”

“…Most, but not all, of the gating residues identified in other CYPs (e.g. 3A4, 2E1, 2A6, 2A13, CYP51, others) have been phenylalanine (Li et al, 2005;Fishelovitch et al, 2009;Zawaira et al, 2011;Shen et al, 2012;Yu et al, 2016;Fan et al, 2018). There have been limited investigations into likely ligand bottlenecks in 2D6 and their roles in metabolism (de Waal et al, 2014;Fischer et al, 2018;Fischer and Smieško, 2019).…”