Resonance Raman investigation of lysine and N-acetylmethionine complexes of ferric and ferrous microperoxidase

Abstract

In order to evaluate the steric and electronic effects of mixed axial ligations on the heme c structure, lysine (Lys) and N-acetylmethionine (AcMet) complexes of ferric and ferrous microperoxidase-8 (MP8(III) and MP8(II), respectively) are characterized by absorption and resonance Raman (RR) spectroscopies. Spectrophotometric titrations establish that MP8(III) binds one molecule of exogenous ligand while MP8(II) forms mono(ligated) and bis(ligated) compounds. The Soret-excited RR spectra of the six-coordinated low-spin MP8(III) complexes show that the macrocycle can adopt different structures between planar and ruffled conformations. The ferriheme c conformation is primarily determined by the ionization state of the His side chain of MP8(III) and, secondarily, by the bonding and nonbonding heme-ligand interactions. As far as the RR spectra of the MP8(II) complexes are concerned, they permit us to conclude that the mixed His/Lys and His/AcMet coordinations induce a nonplanar heme conformation, the extent of deformation again depending on the ionization state of the endogenous His ligand. In contrast, the RR spectra of the bis(Lys) and bis(AcMet) compounds are associated with a planar heme structure. When the His of MP8 is bound to heme c, the stabilization of distorted heme conformations is thus associated with constraints exerted by the Cys-Ala-Gln-Cys-His-peptide on the porphyrin macrocycle. More generally, the spectroscopic data obtained in this study can be used to predict both the axial coordination and the structure of heme in c-type cytochromes.