A heme c-peptide model system for the resonance Raman study of c-type cytochromes: characterization of the solvent-dependence of peptide-histidine-heme interactions.
The visible absorption and Soret-excited resonance Raman spectra of ferrous microperoxidase-8 [MP8(II)], an octapeptide containing a heme c, are reported. These spectroscopies indicate that MP8(II), dissolved in aqueous buffered solutions, forms low-spin six-coordinated complexes in the 7-14 pH range. Intermolecular bonding interactions of MP8(II) in water account for this behavior. On the contrary, when the hemopeptide is dispersed in aqueous solutions containing detergent or an alcohol, the spectroscopic data show that the iron atom of MP8(II) is essentially high-spin five-coordinated in accordance with a monomeric structure of MP8(II). In addition to a high-spin signature to the heme skeletal modes, the high-frequency regions of resonance Raman spectra characterize an electronic influence of the thioether bridges on the frequency of stretching modes of C beta-C beta bonds (nu 2, nu 11, and nu 29). On the other hand, the low-frequency Raman spectra of monomeric MP8(II) at pH 7.5 present significant differences in the 150-250-cm-1 regions depending upon the solvent composition (pH, presence or absence of detergent, alcohol). These effects are attributed to frequency variations of the Fe-N(His)-involving mode which indicate changes in the H-bonding interactions of the axial His and therefore solvent-dependent changes of the octapeptide conformation. Our resonance Raman data further show that the axial His of monomeric MP8(II) could be totally deprotonated in aqueous cetyltrimethylammonium bromide solution at very alkaline pH (pKa = 13.3). The vibrational data (100-1700 cm-1) obtained for the various monomeric forms of MP8(II) are expected to be useful for determining the heme structure and environment in reduced c'-type cytochromes. Comparisons of resonance Raman data with X-ray crystallographic data available for different hemoproteins allow us to evaluate the ionization and H-bonding states of the His bound to the high-spin five-coordinated hemes. These data are discussed in terms of proximal influence of protein-His-heme interactions on the determination and the regulation of a particular biological function.
Département de Biologie Cellulaire et Moléculaire, Centre d'Etudes de Saclay, Gif-sur-Yvette, France.
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Othman
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Le Lirzin
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Desbois
D000595:Amino Acid Sequence; D000818:Animals; D003574:Cytochrome c Group / Q000737:chemistry*; D006418:Heme / Q000737:chemistry*; D006639:Histidine / Q000737:chemistry; D006736:Horses; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry; D011166:Porphyrins; D017433:Protein Structure, Secondary; D012997:Solvents; D013057:Spectrum Analysis; D013059:Spectrum Analysis, Raman; D013329:Structure-Activity Relationship
D003574:Cytochrome c Group; D010455:Peptides; D011166:Porphyrins; D012997:Solvents; D006418:Heme; D006639:Histidine