Solubilization and reconstitution of the Na+/Ca2+ exchanger of cardiac sarcolemma. Properties of the reconstituted system and tentative identification of the protein(s) responsible for the exchange activity.
The cardiac sarcolemma Na+/Ca2+ exchanger has been solubilized in Triton X-100 and reconstituted into asolectin liposomes by removing the detergent with Amberlite XAD-2 beads. The specific Na+/Ca2+ exchange activity of the proteoliposomes was increased about 10 times with respect to the original sarcolemma. The exchange activity was strongly dependent on the amount of acidic phospholipids present in the liposomes. Phosphatidylserine and phosphatidylinositol increased the initial rate of the reconstituted exchanger. Fractionation of the solubilized sarcolemma by rate zonal centrifugation produced fractions in which the specific activity of the exchanger was enriched up to 130-fold with respect to the original membranes. The fractionation scheme used has shown that the exchange activity correlated best with a 33-kDa protein present in the fractionated extract. No correlation has been found between exchange activity and other proteins of higher molecular mass (70 or 82 kDa) which have previously been suggested to correspond to the Na+/Ca2+ exchanger. Thus, the 33-kDa protein may represent the Na+/Ca2+ exchanger of cardiac sarcolemma.
D000818:Animals; D002352:Carrier Proteins / Q000302:isolation & purification / Q000378:metabolism*; D002417:Cattle; D002458:Cell Fractionation; D002501:Centrifugation, Zonal; D007700:Kinetics; D008081:Liposomes / Q000378:metabolism; D009206:Myocardium / Q000032:analysis*; D017830:Octoxynol; D010716:Phosphatidylinositols / Q000502:physiology; D010718:Phosphatidylserines / Q000502:physiology; D011092:Polyethylene Glycols; D012508:Sarcolemma / Q000032:analysis*; D017923:Sodium-Hydrogen Exchangers; D012995:Solubility
D002352:Carrier Proteins; D008081:Liposomes; D010716:Phosphatidylinositols; D010718:Phosphatidylserines; D017923:Sodium-Hydrogen Exchangers; D011092:Polyethylene Glycols; D017830:Octoxynol