Key result
The 70 kDa protein associated with Na/Ca exchange activity is truncated ~35% at its C-terminal.
Why the study?
The nature of the 70 kDa polypeptide associated with the Na/Ca exchanger in bovine cardiac sarcolemmal vesicles was poorly understood.
The 70 kDa polypeptide of the Na/Ca exchanger retains exchange activity and the N-terminal sequence but is truncated at the C-terminus compared to the wild-type protein.
No immediate clinical implications for cardiac ion handling; leaves open whether C-terminal truncation alters exchanger function in vivo.
The Na/Ca exchanger is associated with 160, 120 and 70 kDa polypeptides whose nature is poorly understood. We have purified and characterized the Na/Ca exchanger from bovine cardiac sarcolemmal vesicles (SLVs) by using ion-exchange and affinity chromatographies. The Na/Ca exchanger-enriched fraction was reconstituted into asolectin liposomes [lipid to protein ratio 10:1 (w/w)] that showed Na/Ca exchange activity. Under non-reducing conditions, SDS/PAGE showed a single 70 kDa polypeptide, which was further characterized by immunoblots with different antibodies: SWant, raised against the purified exchanger protein; NH2-terminus, residues 1-21; NCX1, residues 393-406; and Exon F, residues 622-644. Immunoblots under reducing conditions with SWant, NH2-terminus and NCX1 showed three bands migrating at 160, 120 and 70 kDa for SLV preparations, whereas Exon F reacted only with the 160 and 120 kDa bands. Under non-reducing conditions, immunoblots with purified reconstituted Na/Ca exchanger showed a single band at 70 kDa reacting with SWant, NH2-terminus and NCX1 but not with Exon F. We conclude that the 70 kDa protein is associated with Na/Ca exchange activity, has the same N-terminal sequence as the cloned bovine cardiac exchanger, and has its length decreased by at least 35% from its C-terminal portion as compared with that of the wild-type exchanger.
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Saba et al. (1999) studied this question. Na/Ca exchanger purification was evaluated on Polypeptide characterization. The 70 kDa protein is associated with Na/Ca exchange activity, shares the N-terminal sequence of the cloned bovine cardiac exchanger, and is truncated by at least 35% from its C-terminal portion.
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