Dynamics of carbon monoxide binding with cytochromes P-450.

Abstract

The dynamics of CO rebinding with cytochromes P-450cam, P-450scc, and P-450LM2 after laser flash photolysis have been investigated from 293 to 77 K, and the distribution functions of the rate parameters P(k) and of the activation enthalpy P(H) were determined using the maximum entropy method. In a fluid solvent, geminate rebinding is nonexponential, presumably because of a spectral shift induced by protein relaxation on the same time scale. Substrate binding increases the yield of the bimolecular process and decreases the bimolecular rate by 1 or 2 orders of magnitude. The amplitude of these effects seems to correlate with substrate specificity. In a rigid environment at low temperature, cytochromes P-450 exhibit a bimodal distribution of activation enthalpy; P(H) consists of two distinct bands which are in a thermal equilibrium even at 77 K. The results lead to a scheme in which a common structural perturbation splits the conformational substates of cytochromes P-450 into pairs of "doublet" substates with different dynamic properties. The hierarchy of conformational substates of cytochromes P-450 thus contrasts with that of oxygen-binding hemoproteins such as myoglobin.