Ferric His93Gly myoglobin cavity mutant and its complexes with thioether and selenolate as heme protein models

Abstract

The composition of ferric exogenous ligand-free His93Gly sperm whale myoglobin (H93G Mb) at neutral pH has been determined by examination of the spectral properties of the protein over the pH range from 3.0 to 10.5. An apparent pKa value of ~6.6 has been observed for the conversion of a postulated six-coordinate bis-water-bound coordination structure at pH 5.0 to a five-coordinate hydroxide-bound form at pH 10.5. Starting from the exogenous ligand-free ferric H93G protein, ferric mono- and bis-thioether (tetrahydrothiophene, THT)-ligated adducts have been prepared and characterized by UV-visible (UV-vis) absorption and magnetic circular dichroism (MCD) spectroscopy. The mon-THT ferric H93G Mb species has hydroxide as the sixth ligand. The bis-THT derivative is a model for the low-spin ferric heme binding site of native bis-Met-ligated bacterioferritin or streptococcal heme-associated protein (Shp). A novel THT-bound ferryl H93G Mb moiety has been partially formed. The high-spin five-coordinate ferric H93G(selenolate) Mb complex has been prepared using benzeneselenol and characterized by UV-vis and MCD spectroscopy as a model for Se-Cys-ligated ferric cytochrome P450. The results described herein further demonstrate the versatility of the H93G cavity mutant for modeling the coordination structures of novel heme iron protein active sites.