title: Isolation of a protein activator of the clathrin-coated vesicle proton pump.
abstract: An activator of the clathrin-coated vesicle proton translocating ATPase has been purified 1600-fold from bovine brain. The activator, which requires detergent (polyoxyethylene 9-lauryl ether) for release from clathrin-coated vesicles, is heat-stable, trypsin-sensitive, and has an apparent molecular mass of about 6 kDa as determined by high performance liquid chromatography. The activator stimulates the purified H(+)-ATPase of coated vesicles over 50-fold under acidic conditions. Similarly, the activator stimulates proton pumping catalyzed by the reconstituted proton pump. Importantly, this stimulation of proton pumping is observed only when the activator is reconstituted into the interior of the proteoliposomes. Moreover, the activator protein is demonstrated to protect, and co-sediment with, purified proton pump during glycerol gradient centrifugation performed in the presence of ATP. These observations support the notion that this activator serves to determine the pH set point of acidic endomembranes through interactions with the transmembranous sectors of the proton pump.
authors.affiliation: Department of Internal Medicine, University of Texas Southwestern Medical Center, Dallas 75235-9121.
authors.forename: X S
authors.initials: XS
authors.lastname: Xie
authors.forename: B P
authors.initials: BP
authors.lastname: Crider
authors.forename: D K
authors.initials: DK
authors.lastname: Stone
mesh_terms: D000818:Animals; D001923:Brain Chemistry; D002417:Cattle; D002851:Chromatography, High Pressure Liquid; D002966:Clathrin / Q000378:metabolism*; D003034:Coated Pits, Cell-Membrane / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation; D006801:Humans; D009419:Nerve Tissue Proteins / Q000302:isolation & purification* / Q000378:metabolism*; D017494:Proton Pumps / Q000378:metabolism*; D006180:Proton-Translocating ATPases / Q000378:metabolism*
chemical_list: D002966:Clathrin; D009419:Nerve Tissue Proteins; D017494:Proton Pumps; D006180:Proton-Translocating ATPases