Effects on Ca2+ uptake of monoclonal antibodies raised against canine cardiac phospholamban and their correlation with epitopic location.

Abstract

Monoclonal antibodies raised against canine cardiac sarcoplasmic reticulum (SR) phospholamban were used to study the structure function relationships of phospholamban in terms of the regulation of the cardiac SR (Ca2+ - Mg2+)-ATPase. Five monoclonal antibodies were assessed for their ability to affect SR Ca2+ uptake, 3 of these antibodies had no effect on SR Ca2+ uptake, whereas the other 2 monoclonal were able to stimulate SR Ca2+ uptake to levels similar to those achieved by cAMP dependent protein kinase phosphorylation of phospholamban. With the use of synthetic peptides corresponding to various portions of phospholamban it was possible to map the epitope sites the monoclonal antibodies bound to. It was found that the monoclonal antibodies which do not stimulate SR Ca2+ uptake do not bind to the cytosolic domain of phospholamban, whereas the stimulatory antibodies have been shown to bind to phospholamban residues 7 to 16. These results may help to define the residues of phospholamban which interact with the (Ca2+ - Mg2+)-ATPase.