Accessibility of F0 subunits from Escherichia coli ATP synthase. A study with subunit specific antisera.
Antisera have been raised against denatured and non-denatured subunits a, b and c of the F0 complex of the ATP synthase from Escherichia coli. The subunit specificity of the antibodies has been established with immunoblot analysis or enzyme-linked immunosorbent assay (ELISA). In inside-out oriented membrane vesicles the binding avidities of both sets of antisera, against denatured and non-denatured subunits of F0, were similar in the presence as well as in the absence of the F1 part. F1-depleted everted membrane vesicles always produced an efficient binding of the different antisera. In the presence of F1 no antibody recognition could be observed with the anti-a antisera, while anti-b and anti-c antisera showed strong binding. However, a higher membrane protein concentration was necessary for the same antibody binding as in F1-stripped vesicles. In membrane vesicles with right-side-out orientation the recognition of the three F0 subunits was dependent on the antisera set used. Antisera raised against denatured subunits showed no binding to the membrane vesicles, only in case of anti-(dodecylsulfate-denatured b) antiserum could a slight affinity be detected. An antigen-antibody recognition with all three F0 subunits occurred when the antisera against non-denatured subunits were incubated with membrane vesicles of right-side-out orientation. The membrane protein concentration which was necessary to produce a significant binding was 10-100-fold higher compared to that of F1-depleted everted membrane vesicles.
D000906:Antibodies / Q000032:analysis; D000915:Antibody Affinity; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli / Q000201:enzymology; D007106:Immune Sera / Q000032:analysis*; D007120:Immunochemistry; D011489:Protein Denaturation; D006180:Proton-Translocating ATPases / Q000276:immunology*; D013379:Substrate Specificity
D000906:Antibodies; D007106:Immune Sera; D006180:Proton-Translocating ATPases