Stoichiometry of the Cardiac Na+/Ca2+ exchanger NCX1.1 measured in transfected HEK cells.
The stoichiometry with which the Na+/Ca2+ exchanger, NCX1, binds and transports Na+ and Ca2+ has dramatic consequences for ionic homeostasis and cellular function of heart mycocytes and brain neurons, where the exchanger is highly expressed. Previous studies have examined this question using native NCX1 in its endogenous environment. We describe here whole-cell voltage clamp studies using recombinant rat heart NCX1.1 expressed heterologously in HEK-293 cells. This system provides the advantages of a high level of NCX1 protein expression, very low background ion transport levels, and excellent control over clamped voltage and ionic composition. Using ionic conditions that allowed bi-directional currents, voltage ramps were employed to determine the reversal potential for NCX1.1-mediated currents. Analysis of the relation between reversal potential and external [Na+] or [Ca2+], under a variety of intracellular conditions, yielded coupling ratios for Na+ of 1.9-2.3 ions per net charge and for Ca2+ of 0.45 +/- 0.03 ions per net charge. These data are consistent with a stoichiometry for the NCX1.1 protein of 4 Na+ to 1 Ca2+ to 2 charges moved per transport cycle.
Dong
Hui
H
Department of Biochemistry and Molecular Biology, University of Calgary, Calgary T2N 4N1, Canada.
Dunn
Jeremy
J
Lytton
Jonathan
J
D000818:Animals; D001692:Biological Transport; D002460:Cell Line; D018076:DNA, Complementary / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004594:Electrophysiology; D006801:Humans; D007477:Ions; D009206:Myocardium / Q000378:metabolism; D018408:Patch-Clamp Techniques; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry; D019831:Sodium-Calcium Exchanger / Q000737:chemistry*; D013816:Thermodynamics; D014162:Transfection
D018076:DNA, Complementary; D007477:Ions; D011994:Recombinant Proteins; D019831:Sodium-Calcium Exchanger; C110374:sodium-calcium exchanger 1