Reduction kinetics of purified rat liver cytochrome P-450. Evidence for a sequential reaction mechanism dependent on the hemoprotein spin state

Publisher Website
Google Scholar
Abstract
The anaerobic reduction kinetics of purified at liver ferric cytochrome P-450 from phenobarbital-treated rat liver microsomes, reconstituted with saturating NADPH-cytochrome P-450 reductase, have been investigated and were shown not to be monophasic. From experiments correlating changes in the rate of fast-phase reduction with the spin state of the heme iron existing at preequilibrium, data were obtained consistent with a model for spin-state control of cytochrome P-450 reduction wherein the high-spin form of the hemoprotein is more rapidly reduced than the low-spin form. In addition, the temperature dependence of the reduction process in the presence of the substrate benzphetamine was studied. It is suggested that the endothermic nature of the low- to high-spin transition largely accounts for the apparent activation energy observed for the reduction process, with the actual rate constant for reduction of high-spin cytochrome P-450 being relatively temperature insensitive when compared to the rate constant for reduction of the membrane-bound form of the hemoprotein.