Activation energy of the cardiac Na+/Ca2+ exchanger in sarcolemmal vesicles and reconstituted proteoliposomes.
Sarcolemmal membrane vesicles isolated from bovine ventricular tissue accumulate Ca2+ through the Na+/Ca2+ exchanger when exposed to an outwardly directed Na+ gradient. This Ca2+ is then released by the same mechanism if the vesicles are transferred to a Ca(2+)-depleted Na+ buffer. Using the Ca+ indicator, arsenazo III, and a stopped-flow spectrophotometer, we can directly follow the kinetics of Ca2+ extrusion. We can thus measure the activity of the Na+/Ca2+ exchanger by the initial rate of Ca2+ release. We found that it depends upon the external Na+ concentration in a cooperative way, with a Hill coefficient of 2. By studying the temperature dependence of Na+/Ca2+ exchange, we found that it can be described by a single activation energy: Ea = 8.3 +/- 0.4 Kcal/mol. When the Na+/Ca2+ exchanger is reconstituted into lipid vesicles of defined composition, we observe a higher activity if cholesterol is among the lipids. The activation energy becomes 6.1 +/- 0.1 Kcal/mol in this system, but the Arrhenius plot shows a decreased slope for temperatures above 33 degrees C.
Dalla Serra
M
M
Dipartimento di Fisica, Università di Trento, Povo, Italy.
Pederzolli
C
C
Antolini
R
R
Cusinato
F
F
Luciani
S
S
Menestrina
G
G
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002352:Carrier Proteins / Q000378:metabolism*; D002417:Cattle; D066298:In Vitro Techniques; D007700:Kinetics; D008081:Liposomes; D009206:Myocardium / Q000378:metabolism*; D011510:Proteolipids / Q000378:metabolism; D012508:Sarcolemma / Q000378:metabolism; D012964:Sodium / Q000378:metabolism; D019831:Sodium-Calcium Exchanger; D013816:Thermodynamics
D002352:Carrier Proteins; D008081:Liposomes; D011510:Proteolipids; D019831:Sodium-Calcium Exchanger; C020485:proteoliposomes; D012964:Sodium; D002118:Calcium