Differential characterization of neutrophil cytochrome p30 and cytochrome b-558 by low-temperature absorption and resonance Raman spectroscopies.
Cytochrome p30, a novel hemoprotein isolated from rabbit peritoneal neutrophils [Escriou, V., Laporte, F., Garin, J., Brandolin, G. & Vignais, P. V. (1994) J. Biol. Chem. 269, 14007-14014] has been characterized by low-temperature (77 K) absorption and resonance Raman spectroscopies. The spectral data have been compared with those obtained with neutrophil cytochrome b-558. At room temperature, the absorption difference spectra (reduced minus oxidized) of cytochrome p30 and cytochrome b-558 could not been distinguished from each other. However, at 77 K, significant differences were observed. In particular, the alpha band of cytochrome p30 was split whereas that of cytochrome b-558 was symmetrical, but particularly broad. The resonance Raman spectra of cytochrome p30 provided evidence for the presence of two hemes both in the ferric and ferrous states. One of them was a six-coordinated low-spin heme either oxidized or reduced whereas the other one was a high-spin heme, five-coordinated in the reduced state and six-coordinated in the oxidized state. It is probable that two histidine residues constitute the axial ligands of the six-coordinated low-spin heme of cytochrome p30. The resonance Raman spectra of cytochrome b-558 allowed the detection of a six-coordinated low-spin heme, similar to that found in cytochrome p30. The component typical of the high-spin heme of cytochrome p30 was however absent in the spectra of oxidized and reduced cytochrome b-558.
Escriou
V
V
Département de Biologie Moléculaire et Structurale, CEA et URA CNRS 1130, Centre d'Etudes Nucléaires, Grenoble, France.
Laporte
F
F
Vignais
P V
PV
Desbois
A
A
D000818:Animals; D003573:Cytochrome b Group / Q000737:chemistry*; D004227:Dithionite / Q000378:metabolism; D004229:Dithiothreitol / Q000378:metabolism; D006418:Heme / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D019255:NADPH Oxidases; D009504:Neutrophils / Q000201:enzymology*; D010084:Oxidation-Reduction; D010537:Peritoneum / Q000166:cytology; D011817:Rabbits; D013054:Spectrophotometry, Atomic; D013059:Spectrum Analysis, Raman
D003573:Cytochrome b Group; C106367:cytochrome p30; D004227:Dithionite; D006418:Heme; C036888:cytochrome b558; D019255:NADPH Oxidases; D004229:Dithiothreitol