The 70 kD component of the heart sarcolemmal Na+/Ca(2+)-exchanger preparation is the C-terminal portion of the protein.

The cardiac sarcolemmal Na+/Ca(2+)-exchanger was expressed in COS-7 cells by the vaccinia virus system as a fusion protein with a poly-His tag at its C-terminus. Extracts of cells expressing the exchanger construct without the His-tag reacted with an antiserum against the C-terminal portion of the main intracellular loop of the exchanger: in agreement with the finding routinely made on heart sarcolemma and on plasma membrane of cells expressing the cardiac exchanger gene, three bands of about 160, 120, and 70 kD were detected in Western blots. All three bands shifted to higher molecular masses when the construct with the His-tag was expressed, indicating that the three proteins had the same C-terminus. Thus, the 70 kD protein, whose nature has always been obscure, appears to be a degradation product of one of the two larger proteins. N-terminal sequencing of partially purified exchanger preparations has identified the cleavage site(s) producing the 70 kD protein in the 257-269 residue region of the exchanger molecule.