LOCALIZATION OF AN EXCHANGE INHIBITORY PEPTIDE (XIP) BINDING-SITE ON THE CARDIAC SODIUM-CALCIUM EXCHANGER

The exchange inhibitory peptide (XIP; RRLLFYKYVYKRYRAGKQRG) is a poten t inhibitor of cardiac Na-Ca exchange activity, This study attempted t o identify the XIP binding site on the Na-Ca exchange protein. Bovine cardiac sarcolemmal vesicles were proteolyzed and fractionated by XIP- affinity column chromatography, A 24 kDa fragment was purified and sub jected to amino acid sequence analysis. A negatively charged region of intracellular loop f of the Na-Ca exchange protein (IDDDIFEEDEN; aa 4 45-455) was identified, The affinity and specificity of XIP interactio n with peptides IDDDIFEEDEN and GEDDDDDECGEE (another negatively charg ed region of the Na-Ca exchange protein) were examined. XIP cross-link ed to peptide IDDDIFEEDEN but not GEDDDDDECGEE in a pH-dependent manne r, Fluorescence titration binding studies indicated that binding of ID DDIFEEDEN with XIP was saturable (Kd=5 mu M) while binding with GEDDDD DECGEE was not specific, These data suggest that amino acids 445-455 o n Na-Ca exchange loop fare involved in XIP binding. (C) 1997 Academic Press.