Although the majority of enzymes have buried active sites, very little is known about the energetics and mechanisms associated with substrate and product channeling in and out. Gaining direct information about these processes is a challenging task both for experimental and theoretical techniques. Here, we present a methodology that enables following of a ligand during its passage to the active site of cytochrome P450 (CYP) 3A4 and mapping of the free energy associated with this process. The technique is based on a combination of a bioinformatics tool for identifying access channels and bias-exchange metadynamics and provides converged free energies in good agreement with experimental data. In addition, it identifies the energetically preferred escape routes, limiting steps, and amino acids residues lining the channel. The approach was applied to mapping of a complex channel network in a complex environment, i.e., CYP3A4 attached to a lipid bilayer mimicking an endoplasmic reticulum membrane. The results provided direct information about the energetics and conformational changes associated with the ligand channeling. The methodology can easily be adapted to study channeling through other flexible biomacromolecular channels.

Regional Centre of Advanced Technologies and Materials Department of Physical Chemistry Faculty of Science Palacký University Olomouc tř 17 Listopadu 12 771 46 Olomouc Czech Republic

SISSA Scuola Internazionale Superiore di Studi Avanzati via Bonomea 265 34136 Trieste Italy

Citace poskytuje Crossref.org

MOLEonline: a web-based tool for analyzing channels, tunnels and pores (2018 update)

ChannelsDB: database of biomacromolecular tunnels and pores