Localization of an exchange inhibitory peptide (XIP) binding site on the cardiac sodium-calcium exchanger.
The exchange inhibitory peptide (XIP; RRLLFYKYVYKRYRAGKQRG) is a potent inhibitor of cardiac Na-Ca exchange activity. This study attempted to 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 subjected to amino acid sequence analysis. A negatively charged region of intracellular loop f of the Na-Ca exchange protein (IDDDIFEEDEN; aa 445-455) was identified. The affinity and specificity of XIP interaction with peptides IDDDIFEEDEN and GEDDDDDECGEE (another negatively charged region of the Na-Ca exchange protein) were examined. XIP cross-linked to peptide IDDDIFEEDEN but not GEDDDDDECGEE in a pH-dependent manner. Fluorescence titration binding studies indicated that binding of IDDDIFEEDEN with XIP was saturable (Kd=5 microM) while binding with GEDDDDDECGEE was not specific. These data suggest that amino acids 445-455 on Na-Ca exchange loop f are involved in XIP binding.
Hale
C C
CC
The Dalton Cardiovascular Research Center, Department of Veterinary Biomedical Sciences, University of Missouri, Columbia 65211, USA. calvin_hale@muccmail.missouri.edu
Bliler
S
S
Quinn
T P
TP
Peletskaya
E N
EN
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002352:Carrier Proteins / Q000378:metabolism*; D002417:Cattle; D008969:Molecular Sequence Data; D009206:Myocardium / Q000378:metabolism* / Q000648:ultrastructure; D010455:Peptides / Q000737:chemistry* / Q000378:metabolism; D012508:Sarcolemma / Q000378:metabolism*; D019831:Sodium-Calcium Exchanger
D002352:Carrier Proteins; D010455:Peptides; D019831:Sodium-Calcium Exchanger; C066745:exchanger inhibitory peptide; C074530:potassium-dependent sodium-calcium exchanger