Substrates modulate the rate-determining step for CO binding in cytochrome P450cam (CYP101) - A high-pressure stopped-flow study

Item Type:	Article
Title:	Substrates modulate the rate-determining step for CO binding in cytochrome P450cam (CYP101) - A high-pressure stopped-flow study
Creators Name:	Jung, C. and Bec, N. and Lange, R.
Abstract:	The high-pressure stopped-flow technique is applied to study the CO binding in cytochrome P450cam (P450cam) bound with homologous substrates (1R-camphor, camphane, norcamphor and norbornane) and in the substrate-free protein. The activation volume {delta}V# of the CO on-rate is positive for P450cam bound with substrates that do not contain methyl groups. The kon rate constant for these substrate complexes is in the order of 3 × 106 M-1·S-1. In contrast, P450cam complexed with substrates carrying methyl groups show a negative activation volume and a low kon rate constant of ≈ 3 × 104 M-1·S-1. By relating kon and {delta}V# with values for the compressibility and the influx rate of water for the heme pocket of the substrate complexes it is concluded that the positive activation volume is indicative for a loosely bound substrate that guarantees a high solvent accessibility for the heme pocket and a very compressible active site. In addition, subconformers have been found for the substrate-free and camphane-bound protein which show different CO binding kinetics.
Keywords:	High-Pressure Stopped-Flow, Cytochrome P450, CO Ligand Binding, Protein Dynamics
Source:	European Journal of Biochemistry
ISSN:	0014-2956
Publisher:	Blackwell Publishing
Volume:	269
Number:	12
Page Range:	2989-2996
Date:	June 2002
Official Publication:	https://doi.org/10.1046/j.1432-1033.2002.02980.x
PubMed:	View item in PubMed

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