title: Assaying P-Type ATPases Reconstituted in Liposomes.
abstract: Reconstitution of P-type ATPases in unilamellar liposomes is a useful technique to study functional properties of these active ion transporters. Experiments with such liposomes provide an easy access to substrate-binding affinities of the ion pumps as well as to the lipid and temperature dependence of the pump current. Here, we describe two reconstitution methods by dialysis and the use of potential-sensitive fluorescence dyes to study transport properties of two P-type ATPases, the Na,K-ATPase from rabbit kidney and the K(+)-transporting KdpFABC complex from E. coli. Several techniques are introduced how the measured fluorescence signals may be analyzed to gain information on properties of the ion pumps.
authors.affiliation: Department of Biology, University of Konstanz, 635, Universitätsstr. 10, Konstanz, 78464, Germany. h-j.apell@uni-konstanz.de.
authors.forename: Hans-Jürgen
authors.initials: HJ
authors.lastname: Apell
authors.affiliation: Department of Biology, University of Konstanz, 635, Universitätsstr. 10, Konstanz, 78464, Germany.
authors.forename: Bojana
authors.initials: B
authors.lastname: Damnjanovic
mesh_terms: D000251:Adenosine Triphosphatases / Q000737:chemistry* / Q000378:metabolism; D000818:Animals; D027682:Cation Transport Proteins / Q000737:chemistry* / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000378:metabolism; D017136:Ion Transport; D007668:Kidney / Q000201:enzymology; D007700:Kinetics; D008081:Liposomes / Q000737:chemistry* / Q000378:metabolism; D011817:Rabbits; D012964:Sodium / Q000378:metabolism; D000254:Sodium-Potassium-Exchanging ATPase / Q000737:chemistry* / Q000235:genetics
chemical_list: D027682:Cation Transport Proteins; D029968:Escherichia coli Proteins; D008081:Liposomes; D012964:Sodium; D000251:Adenosine Triphosphatases; C058166:potassium translocating Kdp-ATPase, E coli; D000254:Sodium-Potassium-Exchanging ATPase
keywords: BioBeads; Detergents; Dialysis; Electrogenic ion transport; Lipids; Membrane potential; Pump current; Reconstitution; Substrate dependence; Vesicles; Voltage-sensitive fluorescence dyes