Mapping epitopes on the (Ca(2+)-Mg2+)-ATPase of sarcoplasmic reticulum using fusion proteins

Epitopes for a number of monoclonal antibodies (mAbs) binding (Ca(2+)-Mg2+)-ATPase purified from skeletal muscle sarcoplasmic reticulum have been defined by studying binding to fusion proteins generated from cDNA fragment libraries. Comparison of these results with those of previous studies of binding of mAbs to proteolytic fragments of the ATPase have allowed the definition of the epitopes to within approx. 100 residues and for one (mAb 1/2H7) to within 45 residues. The experiments suggest considerable exposure of the nucleotide binding domain of the ATPase on the top surface of the protein. Those mAbs that were found to inhibit steady-state ATPase activity were found to bind to epitopes in the nucleotide binding domain of the ATPase.