Purification and reconstitution of the 32Pi-ATP exchange activity of bovine chromaffin granule membrane.
Ghosts derived from bovine chromaffin granules have a 32Pi-ATP exchange activity which is associated with the H+ pump of that membrane. This activity was low when compared to bacteria, chloroplasts or submitochondrial particles, but had similar properties (Km for ATP and Pi, ATP/Mg2+ ratio, pH profile, inhibition by dicyclohexylcarbodiimide and tributyltin) to the ATPase from above membranes. The 32Pi-ATP exchange activity was solubilized by cholate/octylglucoside mixtures. The soluble extract was lipid depleted by ammonium sulfate fractionation and partially purified by sucrose gradient centrifugation. The purified preparation was reconstituted with phospholipids by freeze-thawing. The reconstituted vesicles had a 32Pi-ATP exchange sensitive to dicyclohexylcarbodiimide and trybutyltin and an ATPase with a sensitivity to the inhibitors which varied with the reconstitution conditions. The alpha- and beta-subunits of F1-ATPase were major components of the preparation.
D000255:Adenosine Triphosphate / Q000494:pharmacology*; D000818:Animals; D002417:Cattle; D002837:Chromaffin Granules / Q000187:drug effects / Q000378:metabolism*; D002838:Chromaffin System / Q000378:metabolism*; D004024:Dicyclohexylcarbodiimide / Q000494:pharmacology; D007425:Intracellular Membranes / Q000187:drug effects / Q000378:metabolism*; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D010710:Phosphates / Q000378:metabolism*; D010761:Phosphorus Radioisotopes
D010710:Phosphates; D010761:Phosphorus Radioisotopes; D004024:Dicyclohexylcarbodiimide; D000255:Adenosine Triphosphate; D008274:Magnesium