Comparison of the dynamics of substrate access channels in three cytochrome P450s reveals different opening mechanisms and a novel functional role for a buried arginine.
Understanding the mechanism and specificity of substrate binding in the cytochrome P450 (P450) superfamily is an important step toward explaining its key role in drug metabolism, toxicity, xenobiotic degradation, and several biosynthetic pathways. Here we investigate the ligand exit pathways and mechanisms of P450cam (CYP101), P450BM-3 (CYP102), and P450eryF (CYP107A1) by using random expulsion molecular dynamics and classical molecular dynamics simulations. Although several different pathways are found for each protein, one pathway is common to all three. The mechanism of ligand exit along this pathway is, however, quite different in the three different proteins. For P450cam, small backbone conformational changes, in combination with aromatic side chain rotation, allow for the passage of the rather rigid, compact, and hydrophobic substrate, camphor. In P450BM-3, larger transient backbone changes are observed on ligand exit. R47, situated at the entrance to the channel, appears important in guiding negatively charged fatty acid substrates in and out of the active site. In P450eryF, an isolated buried arginine, R185, stabilized by four hydrogen bonds to backbone carbonyl oxygen atoms, is located in the exit channel and is identified as having a particularly unusual functionality, dynamically gating channel opening. The results for these three P450s suggest that the channel opening mechanisms are adjusted to the physico-chemical properties of the substrate and can kinetically modulate protein-substrate specificity.
Winn
Peter J
PJ
European Molecular Biology Laboratory, Meyerhofstrasse 1, 69117 Heidelberg, Germany.
Lüdemann
Susanna K
SK
Gauges
Ralph
R
Lounnas
Valère
V
Wade
Rebecca C
RC
D001120:Arginine / Q000737:chemistry*; D001426:Bacterial Proteins; D001665:Binding Sites; D002164:Camphor / Q000737:chemistry; D019475:Camphor 5-Monooxygenase / Q000737:chemistry*; D002244:Carbon / Q000737:chemistry; D003577:Cytochrome P-450 Enzyme System / Q000737:chemistry*; D004917:Erythromycin / Q000031:analogs & derivatives* / Q000494:pharmacology; D006860:Hydrogen Bonding; D007700:Kinetics; D008024:Ligands; D006899:Mixed Function Oxygenases / Q000737:chemistry*; D008956:Models, Chemical; D008958:Models, Molecular; D009251:NADPH-Ferrihemoprotein Reductase; D010100:Oxygen / Q000737:chemistry; D011485:Protein Binding; D012984:Software; D013379:Substrate Specificity; D013816:Thermodynamics; D013997:Time Factors
D001426:Bacterial Proteins; D008024:Ligands; C034277:6-deoxyerythronolide B; D004917:Erythromycin; D002244:Carbon; D002164:Camphor; D003577:Cytochrome P-450 Enzyme System; D001120:Arginine; D006899:Mixed Function Oxygenases; C054444:eryF protein, Saccharopolyspora erythraea; D019475:Camphor 5-Monooxygenase; D009251:NADPH-Ferrihemoprotein Reductase; C052423:flavocytochrome P450 BM3 monoxygenases; D010100:Oxygen