Purification and amino-terminal sequence of the bovine cardiac sodium-calcium exchanger: evidence for the presence of a signal sequence.
The Na(+)-Ca2+ exchange carrier was purified from bovine cardiac tissue by a new procedure which relies principally upon anion-exchange chromatography. The purified protein exhibited two major bands on sodium dodecyl sulfate gels, at 120 and 160 kDa. The relative intensities of the two bands could be altered by variations in the procedures used for preparing the samples for electrophoresis, suggesting that they represent two different conformational states of the same protein. The NH2-terminal amino acid sequences of the 120- and 160-kDa bands were identical and agreed closely with a region of the deduced amino acid sequence of the recently cloned canine cardiac exchanger. The NH2-terminal sequence was preceded in the deduced sequence by a 32-residue segment that exhibited the characteristics of a signal sequence; the initial amino acid in the NH2-terminal sequence followed immediately after the predicted cleavage site for the signal sequence. The Na(+)-Ca2+ exchanger appears to be unique among membrane transport carriers in encoding a cleaved signal sequence. The characteristics of the sequences flanking the first putative transmembrane segment of the mature exchanger suggest that the signal sequence is necessary to ensure the correct topological orientation of the exchanger in the membrane.
Durkin
J T
JT
Roche Institute of Molecular Biology, Department of Biochemistry, Nutley, New Jersey.
Ahrens
D C
DC
Pan
Y C
YC
Reeves
J P
JP
D000595:Amino Acid Sequence; D000818:Animals; D002118:Calcium / Q000737:chemistry*; D002352:Carrier Proteins / Q000737:chemistry / Q000302:isolation & purification*; D002417:Cattle; D002852:Chromatography, Ion Exchange; D006352:Heart Ventricles / Q000737:chemistry; D008969:Molecular Sequence Data; D009206:Myocardium / Q000737:chemistry*; D010446:Peptide Fragments / Q000737:chemistry; D011498:Protein Precursors / Q000737:chemistry*; D021382:Protein Sorting Signals / Q000737:chemistry / Q000302:isolation & purification*; D012508:Sarcolemma / Q000737:chemistry; D019831:Sodium-Calcium Exchanger
D002352:Carrier Proteins; D010446:Peptide Fragments; D011498:Protein Precursors; D021382:Protein Sorting Signals; D019831:Sodium-Calcium Exchanger; D002118:Calcium