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Immunochemistry of cytochrome c. Identification of antigenic determinants through the study of hybrid molecules.

Authors
Type
Published Article
Journal
Journal of Biological Chemistry
Publisher
American Society for Biochemistry and Molecular Biology
Volume
255
Issue
13
Pages
6133–6137
Source
UCSC Bioinformatics biomedical-ucsc
License
Unknown

Abstract

Antibodies elicited in rabbits against horse cytochrome c cross-react in varying degree with the cytochromes c of other species. Radioimmunoassay experiments are described in which 23% of the immunoglobulin shown to bind to horse cytochrome c fails to react with beef cytochrome c, and 14% of that bound by beef cytochrome c does not react with rabbit cytochrome c. Studies of hybrid cytochromes, prepared by a reconstitution procedure involving the cleavage and re-formation of peptide bond 65-66, show that these differences result from single amino acid replacements, Beef cytochrome c differs in binding capacity from horse cytochrome c as a result of the substitution of a glycine for a lysine residue in sequence position 60, and the difference between rabbit cytochrome c and beef cytochrome c reflects the substitution of a valine for a proline residue in position 44. Reconstituted horse cytochrome c and reconstituted beef cytochrome c have binding capacities indistinguishable from those of the parent proteins. The presence of a homoserine rather than a methionine residue in position 65 results, in the case of the reconstituted horse molecule, in a slightly lower affinity for the antibody population directed against the lysine-60 region. A corresponding difference is not observed in the case of beef cytochrome c and its reconstitution product, which do not bind the population in question.

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