X-ray absorption spectroscopic characterization of a cytochrome P450 compound II derivative

Abstract

The cytochrome P450 enzyme CYP119, its compound II derivative, and its nitrosyl complex were studied by iron K-edge x-ray absorption spectroscopy. The compound II derivative was prepared by reaction of the resting enzyme with peroxynitrite and had a lifetime of Ϸ10 s at 23°C. The CYP119 nitrosyl complex was prepared by reaction of the enzyme with nitrogen monoxide gas or with a nitrosyl donor and was stable at 23°C for hours. Samples of CYP119 and its derivatives were studied by x-ray absorption spectroscopy at temperatures below 140 (K) at the Advanced Photon Source of Argonne National Laboratory. The x-ray absorption near-edge structure spectra displayed shifts in edge and pre-edge energies consistent with increasing effective positive charge on iron in the series native CYP119 < CYP119 nitrosyl complex < CYP119 compound II derivative. Extended x-ray absorption fine structure spectra were simulated with good fits for k ‫؍‬ 12 Å ؊1 for native CYP119 and k ‫؍‬ 13 Å ؊1 for both the nitrosyl complex and the compound II derivative. The important structural features for the compound II derivative were an iron-oxygen bond length of 1.82 Å and an iron-sulfur bond length of 2.24 Å, both of which indicate an iron-oxygen single bond in a ferryl-hydroxide, Fe IV OH, moiety.EXAFS ͉ ferryl-oxo ͉ XANES

Nature of the Ferryl Heme in Compounds I and II

“…30 for a recent discussion). X-ray crystallographic studies varied in their conclusions on bond length probably because these early structures were photoreduced.…”

On the Electronic Structure ofmer,trans‐[RuCl3(1H‐indazole)2(NO)], a Hypothetical Metabolite of the Antitumor Drug Candidate KP1019: An Experimental and DFT Study

“…There are no reported XANES measurements for similar systems at the Ru:K edge, however, there are a number of papers on Rubased oxides [50][51][52] as well as on Fe-based systems. [53,82] An energy shift per oxidation state of 1.47(12) eV can be found for Ru-based perovskites. [85] Similar values have been determined for Ru-based double perovskites, [84] Ru-based layered perovskite-like structures, [83] or ruthenium oxides.…”

Reaction Intermediates and Molecular Mechanism of Peroxynitrite Activation by NO Synthases

“…In this regard, the similarity between the spectroscopic fingerprints of I435 and oxoferryl complexes has led the first research groups to assign this reaction intermediate to a Cpd-II-like species (12,13). More recently, Newcomb and coworkers used x-ray absorption spectroscopy to identify the reaction intermediate observed upon interaction of PN with cytochrome P450 as a Cpd-II species (13). The photo-oxidation of this intermediate apparently gave rise to the buildup of a Cpd-I species (14,17,50,51), similar to what was reported for chloroperoxidase (16).…”

“…1, red arrows) with the concomitant buildup of a reaction intermediate identified as an oxoferryl species. This intermediate was later assigned to a compound II species (Cpd-II) by Newcomb and colleagues when analyzing the interaction of PN with various cytochromes P450s (13)(14)(15)(16). This putative Cpd-II generated by the reaction of various cytochromes P450 with PN was photoreduced to an intermediate that was somewhat competent for catalysis (15,17).…”