Cytochrome c peroxidase complexed with cytochrome c has an unperturbed heme moiety.
Transient resonance Raman, Raman difference, circular dichroism (CD), and optical absorption studies have been carried out on the electrostatic complexes formed by yeast cytochrome c peroxidase (CCP) with horse cytochrome c (Cytc) in low ionic strength solutions. In all the complexes examined [e.g., CCP(II)/Cytc(II), CCP(III)/Cytc(II), CCP(III)/Cytc(III)], the local heme environments of both proteins are largely unperturbed upon complexation. Specifically, CCP preserves a completely pentacoordinate high-spin heme in both its ferric and ferrous forms in CCP/Cytc complexes and uncomplexed mixtures. We found no evidence corroborating the previously reported increase in the low-spin fraction of CCP heme upon complexation with Cytc [Hildebrandt et al. (1992) Biochemistry 31, 2384-2392]. Instead, our Raman data strongly suggest that the H-bonding networks in the distal and proximal pockets of CCP are well maintained in the complexes. On the other hand, CD spectra of CCP(III)/Cytc(III) complexes showed substantial variations (relative to the uncomplexed mixtures) in the far-UV region, reflecting some protein conformational rearrangements. In addition, the spectral data suggest that complexation with Cytc affects the previously observed pH-dependent flexibility of the heme structure of CCP and thus influences the photodynamics of the CCP active site.
Wang
J
J
Department of Chemistry, University of New Mexico, Albuquerque 87131, USA.
Larsen
R W
RW
Moench
S J
SJ
Satterlee
J D
JD
Rousseau
D L
DL
Ondrias
M R
MR
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002942:Circular Dichroism; D003574:Cytochrome c Group / Q000737:chemistry* / Q000235:genetics; D003578:Cytochrome-c Peroxidase / Q000737:chemistry* / Q000235:genetics; D004563:Electrochemistry; D006418:Heme / Q000737:chemistry*; D006736:Horses; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D008969:Molecular Sequence Data; D015394:Molecular Structure; D009994:Osmolar Concentration; D010777:Photochemistry; D011487:Protein Conformation; D012441:Saccharomyces cerevisiae / Q000201:enzymology; D012996:Solutions; D013053:Spectrophotometry; D013059:Spectrum Analysis, Raman; D013816:Thermodynamics
D003574:Cytochrome c Group; D012996:Solutions; D006418:Heme; D003578:Cytochrome-c Peroxidase