The cytochrome c peroxidase.cytochrome c electron transfer complex. Experimental support of a hypothetical model.
A hypothetical model of the cytochrome c peroxidase.cytochrome c complex (Poulos, T. L., and Kraut, J. (1980) J. Biol. Chem. 255, 10322-10330) predicts charge interactions between aspartic acid residues of the peroxidase having a spatial distribution that is complementary to the distribution of essential and highly conserved residues of cytochrome c. In a first attempt to test this model, carboxyl groups of cytochrome c peroxidase have been modified with a water-soluble carbodiimide, either alone or in combination with a nucleophile. Modification led to the loss of up to 90% of the ferrocytochrome c peroxidase activity. At least 4-5 carboxyl groups out of a total of 48, but none of the heme carboxyls, were modified in a derivative with 14% residual activity. In the peroxidase.cytochrome c complex the rate of peroxidase inactivation is slowed and approximately 2 carboxyl groups are protected from chemical modification. In the presence of the carbodiimide, cytochrome c and peroxidase were cross-linked to form a covalent 1:1 complex and the linkage sites were preliminarily characterized. Cross-linking occurred to carboxyl groups of the NH2-terminal fragment 1-119 and of fragment 172-229. The four crucial aspartates of the hypothetical model are located in these same two sequence regions.
D000818:Animals; D003432:Cross-Linking Reagents / Q000494:pharmacology; D003574:Cytochrome c Group / Q000378:metabolism*; D003578:Cytochrome-c Peroxidase / Q000378:metabolism*; D004579:Electron Transport; D005022:Ethyldimethylaminopropyl Carbodiimide / Q000494:pharmacology*; D006736:Horses; D007700:Kinetics; D008954:Models, Biological; D009097:Multienzyme Complexes / Q000378:metabolism*; D009206:Myocardium; D010446:Peptide Fragments / Q000032:analysis; D010544:Peroxidases / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000201:enzymology
D003432:Cross-Linking Reagents; D003574:Cytochrome c Group; D009097:Multienzyme Complexes; D010446:Peptide Fragments; D010544:Peroxidases; D003578:Cytochrome-c Peroxidase; D005022:Ethyldimethylaminopropyl Carbodiimide