Antigenic sites on cytochrome c2 from Rhodospirillum rubrum.
The antigenic determinants for three monoclonal antibodies against cytochrome c2 from Rhodospirillum rubrum were partially characterized by differential chemical modification of free and antibody-bound cytochrome c2 and by cross-reactivity analysis with different antigens. Circular dichroism spectroscopy was used to probe the effect of antibody binding on the conformation of cytochrome c2. The binding of two antibodies was strongly dependent on the native folding of the antigen. The first antibody bound to a determinant around the exposed heme edge on the 'front side' of the molecule which is not antigenic in mitochondrial cytochrome c2. Binding of this antibody to cytochrome c increased the induced CD of the ferric heme in a manner similar to that observed previously when mitochondrial cytochrome-c oxidase bound to the front side of cytochrome c. This observation points to a subtle conformational adaptation of the antigen induced by the antibody. The determinant for the second antibody, which also affected the heme CD spectrum of the antigen, was on a polypeptide loop where cytochrome c2 differs from mitochondrial cytochrome c by an eight-residue insertion. The third antibody, which did not induce a change in CD, bound to a sequential determinant near the amino end of cytochrome c2. Only this antibody cross-reacted with isolated cytochrome-c-derived peptides and with apo-cytochrome c2. A preliminary analysis of the polyclonal immune response of five rats against cytochrome c2 indicates that, unlike in eukaryotic cytochrome c, antigenic determinants are distributed over the whole polypeptide chain of the prokaryotic immunogen.
Saad
B
B
Biochemisches Institut, Universität Zürich, Switzerland.
Bosshard
H R
HR
D000107:Acetylation; D000911:Antibodies, Monoclonal / Q000276:immunology*; D001666:Binding Sites, Antibody; D001667:Binding, Competitive; D002942:Circular Dichroism; D003429:Cross Reactions; D003574:Cytochrome c Group / Q000276:immunology*; D045362:Cytochromes c2; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000032:analysis* / Q000235:genetics; D007106:Immune Sera / Q000276:immunology; D008239:Lysine / Q000032:analysis*; D008958:Models, Molecular; D011487:Protein Conformation; D012247:Rhodospirillum rubrum / Q000201:enzymology* / Q000235:genetics / Q000276:immunology
D000911:Antibodies, Monoclonal; D003574:Cytochrome c Group; D000939:Epitopes; D007106:Immune Sera; D045362:Cytochromes c2; D008239:Lysine