Effects on ATPase activity of monoclonal antibodies raised against (Ca2+ + Mg2+)-ATPase from rabbit skeletal muscle sarcoplasmic reticulum and their correlation with epitope location.
A total of 28 monoclonal antibodies have been raised against the (Ca2+ + Mg2+)-ATPase of rabbit skeletal muscle sarcoplasmic reticulum. Epitope mapping, using protein fragments generated by proteolysis, indicates that these antibodies include examples binding to at least four distinct epitopes on the A1 and B tryptic fragments of the ATPase. Competition data also show that the 28 antibodies are directed against at least five spatially distinct regions. Altogether, nine inhibitory antibodies were produced: six of these inhibitory antibodies mapped to the same spatial region, although they appear to bind to two distinct epitopes located within the hinge region and the nucleotide-binding domains of current structural models; one antibody bound to an epitope located within the phosphorylation domain and the stalk-transmembranous region designated M4S4 by Brandl, Green, Korczak & MacLennan [(1986) Cell 44, 597-607]. Two of the inhibitory antibodies recognized assembled epitopes exclusively and could not be mapped. Binding to four of the five identified spatial regions was without effect on activity. These data show that the inhibition of catalytic activity by monoclonal antibodies is achieved only by binding to defined regions of the ATPase and they may therefore provide useful probes of structure-function relationships.
Colyer
J
J
Department of Biochemistry, University of Southampton, U.K.
Mata
A M
AM
Lee
A G
AG
East
J M
JM
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D017301:Ca(2+) Mg(2+)-ATPase / Q000037:antagonists & inhibitors* / Q000276:immunology; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors* / Q000276:immunology; D000939:Epitopes / Q000276:immunology*; D011817:Rabbits; D012519:Sarcoplasmic Reticulum / Q000201:enzymology*
D000911:Antibodies, Monoclonal; D000939:Epitopes; D017301:Ca(2+) Mg(2+)-ATPase; D000252:Calcium-Transporting ATPases