Epsilon-binding regions of the gamma subunit of Escherichia coli ATP synthase.
The interaction between the gamma and epsilon subunits of the F1-ATPase sector of Escherichia coli ATP synthase has been investigated using monoclonal antibodies directed against the gamma subunit and ligand blotting using 125I-epsilon. Monoclonal antibody (MAb) gamma-1 was able to bind to epsilon-depleted F1-ATPase but not to epsilon-replete F1, implying that epsilon blocked access to the epitope. A ligand blot assay for the binding of 125I-epsilon to gamma was developed. Both MAb gamma-1 and a second antibody, MAb gamma II, inhibited binding of 125I-epsilon to gamma in this assay while two other anti-gamma monoclonal antibodies did not. The epitope recognized by MAb gamma-1 was mapped between residues R49 and R70, quite distant in sequence from that of MAb gamma II, which is located C-terminal to residue K199 of the 286-residue polypeptide. The competition of these antibodies with epsilon for binding to gamma implies that their epitopes, quite separate in sequence, are both located in parts of the subunit involved in binding epsilon.
Dunn
S D
SD
Department of Biochemistry, University of Western Ontario, London, Canada. sdunn@julian.uwo.ca
D000911:Antibodies, Monoclonal; D001665:Binding Sites; D000939:Epitopes / Q000032:analysis; D004926:Escherichia coli / Q000201:enzymology*; D015151:Immunoblotting; D007457:Iodine Radioisotopes; D046911:Macromolecular Substances; D008970:Molecular Weight; D010449:Peptide Mapping; D006180:Proton-Translocating ATPases / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*
D000911:Antibodies, Monoclonal; D000939:Epitopes; D007457:Iodine Radioisotopes; D046911:Macromolecular Substances; D006180:Proton-Translocating ATPases