A Highly Reactive P450 Model Compound I

Abstract

The detection and kinetic characterization of a cytochrome P450 model compound I, [OFe(IV)-4-TMPyP](+) (1), in aqueous solution shows extraordinary reaction rates for C-H hydroxylations. Stopped-flow spectrophotometric monitoring of the oxidation of Fe(III)-4-TMPyP with mCPBA revealed the intermediate 1, which displays a weak, blue-shifted Soret band at 402 nm and an absorbance at 673 nm, typical of a porphyrin pi-radical cation. This intermediate was subsequently transformed into the well-characterized OFe(IV)-4-TMPyP. Global analysis afforded a second-order rate constant k(1) = (1.59 +/- 0.06) x 10(7) M(-1) s(-1) for the formation of 1 followed by a first-order decay with k(2) = 8.8 +/- 0.1 s(-1). (1)H and (13)C NMR determined 9-xanthydrol to be the major product (approximately 90% yield) of xanthene oxidation by 1. Electrospray ionization mass spectrometry carried out in 47.5% (18)OH(2) indicated 21% (18)O incorporation, consistent with an oxygen-rebound reaction scenario. Xanthene/xanthene-d(2) revealed a modest kinetic isotope effect, k(H)/k(D) = 2.1. Xanthene hydroxylation by 1 occurred with a very large second-order rate constant k(3) = (3.6 +/- 0.3) x 10(6) M(-1) s(-1). Similar reactions of fluorene-4-carboxylic acid and 4-isopropyl- and 4-ethylbenzoic acid also gave high rates for C-H hydroxylation that correlated well with the scissile C-H bond energy, indicating a homolytic hydrogen abstraction transition state. Mapping the observed rate constants for C-H bond cleavage onto the Brønsted-Evans-Polanyi relationship for similar substrates determined the H-OFe(IV)-4-TMPyP bond dissociation energy to be approximately 100 kcal/mol. The high kinetic reactivity observed for 1 is suggested to result from a high porphyrin redox potential and spin-state-crossing phenomena. More generally, subtle charge modulation at the active site may result in high reactivity of a cytochrome P450 compound I.

Mechanism of 17α,20-Lyase and New Hydroxylation Reactions of Human Cytochrome P450 17A1

“…2, step 4) or the FeO 3ϩ species (Fig. 2, step 6), often referred to as compound I (4,10,11). With P450s 17A1 and 19A1, a variety of approaches has been applied, including theoretical calculations, biomimetic models, spectroscopy, substrate atom labeling, and kinetics .…”

Heme-thiolate ferryl of aromatic peroxygenase is basic and reactive

“…The central paradigm of CYP oxygen activation is recognized to involve the formation of ferryl intermediates, Fe IV =O. Oxoiron(IV) porphyrin cation radicals (compound I) and oxoiron(IV) porphyrins (compound II) have been observed in CYP (18) as well as classic peroxidases (19), chloroperoxidase (CPO) (20), and catalases (21) and in a variety of model porphyrin systems (22,23). Variations in the reactivities of these ferryl species and the roles of axial ligation and heme environment continue to be vigorously studied and debated.…”

Spectroscopic observation of iodosylarene metalloporphyrin adducts and manganese(V)-oxo porphyrin species in a cytochrome P450 analogue

“…Peroxyacids 26,[32][33][34][35] , hydroperoxides [36][37][38][39] or inorganic salts [40][41][42][43][44][45] were generally employed as terminal oxidants in mechanistic studies, and the reactions were always performed in water or mixed solvents (such as CH 3 OH/H 2 O or CH 3 CN/H 2 O). In fact, PhIO is often mechanistically cleaner than these alternatives 46 .…”