Electronic Structure of a Transient Histidine Radical in Liquid Aqueous Solution:  EPR Continuous-Flow Studies and Density Functional Calculations

Abstract

Transient histidine radicals formed in aqueous solutions by oxidation of histidine with a Ti3+/H2O2 Fenton system at pH 2.0 have been studied by EPR using a fast continuous-flow setup and a dielectric ring resonator equipped with a mixing chamber. A histidine peroxy radical with a single EPR line at g = 2.0151 and a histidine cation radical with a complex hyperfine structure and g = 2.0023 have been detected. The hyperfine structure of the latter radical was analyzed by investigating two selectively deuterated histidines and using an EPR simulation and fit program for analysis of the spectra. Isotropic hyperfine coupling constants of two β-protons, three ring protons, and two nitrogen nuclei have been determined in this way and assigned to a histidine−OH adduct cation radical. Density functional theory (DFT) calculations at the B3LYP and PWP86 levels have been performed on protonated cation radicals of 4-ethylimidazole (as histidine models), yielding isotropic hyperfine coupling constants for three different positions of OH addition. The C5 position for OH addition (a 5-oxohistidine cation radical) is clearly supported by the calculated hyperfine coupling constants. The agreement between DFT and EPR is further improved when hydrogen-bonding interactions to the N1 and C2 protons are introduced in the calculations.

Thermal rearrangement of an N-hydroxy-imidazole thiocarbamoyl derivative as a simple entry into the 4-thio-imidazole motif

“…Experimental -General Procedures) at carbon-4 (and -5) is 0.35, while at carbon-2 this value increases to 0.50, giving a ratio of spin densities at C-4 (Im2)/C-2 (Im3) of 1.4:1. Inspection of earlier literature shows a similar trend, 10 and Solé's calculations of the spin density of a π-type imidazoyl radical at C-4 (and C-5) found a value of 0.0952, while at C-2 the value increased to 0.1607, 11 resulting in a similar ratio of 1.2:1, closer to the ratio of products found. Of course if radicals are indeed involved they would have to be associated within a tight radical pair, since no dimeric products were detected.…”

Characterization of Three mnp Genes of Fomitiporia mediterranea and Report of Additional Class II Peroxidases in the Order Hymenochaetales

“…This performance has been attributed to a unique surface-exposed tyrosine residue and received support from homology modeling (38,55). However, it has also been suggested that the imidazol side group of histidine could serve as a radical stabilizing group in a manner analogous to the indol group of tryptophan (32). This would suggest a new type of VP in the Hymenochaetales, an idea that is further supported by the conservation of this residue in partial sequences of other hymenochaetalean taxa (Phylloporia ribis, Hymenochaete corrugata, Phellinidium ferrugineofuscum) that form a clade with FmMnP2 and FmMnP3.…”

Histidinyl Radical Formation in the Self-peroxidation Reaction of Bovine Copper-Zinc Superoxide Dismutase

“…Formation of that radical adduct would require either deprotonation at C-2 after hydroxyl radical addition at that position (unlikely because it would require breaking a -bond on what is probably a -radical) or would result in forming a carbon atom with 5 bonds. It is also possible that the free radical trapped from free histidine (but not necessarily SOD) is the free radical formed by addition of hydroxyl radical to C-5 of the imidazole ring, which has been directly observed both at pH 2 and 7 (21,24). The resulting hydroxyl addition radical would have to dehydrate to form 2-oxohistidine.…”