Model and enzymatic studies with cytochrome P-450

Abstract

The cytochromes P-450 which have only been recognized for the past two decades are now the most widely studied of all protiens. As is often the case with enzymes much of the detailed information concerning thier mechanism of action comes princi- pally from the chemical studies that have been car- ried out. In thhe case of cytochrome P-450 the characteristic optical spectrum has been shown to arise from the axial coordination of a thiolate anion [1,2] and a theoretical interpretation of the resulting `split Soret band' in the electronic spectrum has been given [3]. The catalytic cycle for P-450 is shown in the scheme. After binding of substrate (RH) to the resting enzyme (1) the high spin ferric complex (11) is reduced to the give the ferrous complex (111) which is still coordinated by the axial thiolate ligand [4]. Oxygenation then gives IV which due to the thiolate ligation has properties quite different from other oxygenated hemeproteins such as oxymyoglobin [5]. The further steps, and subsequent intermediates, in the catalitic cycle have not yet been identified in the enzymic systems. However, model studies suggest strutures and mechanisms of action for the subsequent steps. Thus, the eletrochemical reduction of a simple oxygenated porphyrin, aimed at mimick- ing the second enzymatic reduction (IVar→V), gave an ɳ 2-peroxy complex (1) which may represent the penultimate step in the enzymatic cycle. Loss of oxygen at the oxidation level of water from V(what- ever its electronic configuration might be) would lead to a species which could be represented as a ferric oxene complex (2). While the chemistry that P-450 performs (alkene epoxidation and hydrocarbon hydroxylation) has been shown not to proceed via oxene-like reactions, another electronic configuration of (2), namely (3) appears as a promising candidate for the active oxidizing agent [6]. In addition, this oxo-iron (IV) porphyrin π-cation radical (3) has already been established as the key intermediate in the functioning opf the catalases and peroxidases [7] and may also be as the key intermediate in the cyto- chrome oxidase mediated for electron reduction of dioxygen to water [8]. Groves and his collaborators [9] have shown that a complex having the same electronic configuration as (3) can be prepared from ferric porphyrins and iodsylbenzene. Such systems cause both epoxidation and hydroxylation with selectivity similar to those of ▪ P-450 (Table I). We have extended this chemistry using ruthenium porphyrins [10] and find that a ruthenium (III) octaethylporphyrin and iodosylbenzene will catalyze epoxidation and hydroxylation in a similar fashion (Table 1). In addition, a relatively stable complex, characterized as (4) has been isolated which exhibits an ESR signal at g = 2.0 confirming its porphyrin radical nature. This isolated complex performs the same oxidations as in the catalytic regime described in Table I. where the formation of, ▪ inter alia, cyclohexyl bromide from cyclohexene confirms the radical nature of the oxidation processes.