title: Na+ transport by the A1AO-ATP synthase purified from Thermococcus onnurineus and reconstituted into liposomes.
abstract: The ATP synthase of many archaea has the conserved sodium ion binding motif in its rotor subunit, implying that these A1AO-ATP synthases use Na(+) as coupling ion. However, this has never been experimentally verified with a purified system. To experimentally address the nature of the coupling ion, we have purified the A1AO-ATP synthase from T. onnurineus. It contains nine subunits that are functionally coupled. The enzyme hydrolyzed ATP, CTP, GTP, UTP, and ITP with nearly identical activities of around 40 units/mg of protein and was active over a wide pH range with maximal activity at pH 7. Noteworthy was the temperature profile. ATP hydrolysis was maximal at 80 °C and still retained an activity of 2.5 units/mg of protein at 45 °C. The high activity of the enzyme at 45 °C opened, for the first time, a way to directly measure ion transport in an A1AO-ATP synthase. Therefore, the enzyme was reconstituted into liposomes generated from Escherichia coli lipids. These proteoliposomes were still active at 45 °C and coupled ATP hydrolysis to primary and electrogenic Na(+) transport. This is the first proof of Na(+) transport by an A1AO-ATP synthase and these findings are discussed in light of the distribution of the sodium ion binding motif in archaea and the role of Na(+) in the bioenergetics of archaea.
authors.affiliation: From the Molecular Microbiology and Bioenergetics, Institute of Molecular Biosciences, Johann Wolfgang Goethe University Frankfurt/Main, Max-von-Laue-Str. 9, 60438 Frankfurt, Germany.
authors.forename: Florian
authors.initials: F
authors.lastname: Mayer
authors.affiliation: the Korea Institute of Ocean Science and Technology, 787 Haeanro, Ansan 426-744, South Korea, the Department of Marine Biotechnology, University of Science and Technology, 217 Gajeongro, Daejeon 350-333, South Korea, and.
authors.forename: Jae Kyu
authors.initials: JK
authors.lastname: Lim
authors.affiliation: the Department of Molecular Membrane Biology, Max Planck Institute of Biophysics, Max-von-Laue-Str. 3, 60438 Frankfurt/Main, Germany.
authors.forename: Julian D
authors.initials: JD
authors.lastname: Langer
authors.affiliation: the Korea Institute of Ocean Science and Technology, 787 Haeanro, Ansan 426-744, South Korea, the Department of Marine Biotechnology, University of Science and Technology, 217 Gajeongro, Daejeon 350-333, South Korea, and.
authors.forename: Sung Gyun
authors.initials: SG
authors.lastname: Kang
authors.affiliation: From the Molecular Microbiology and Bioenergetics, Institute of Molecular Biosciences, Johann Wolfgang Goethe University Frankfurt/Main, Max-von-Laue-Str. 9, 60438 Frankfurt, Germany, vmueller@bio.uni-frankfurt.de.
authors.forename: Volker
authors.initials: V
authors.lastname: Müller
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D006868:Hydrolysis; D008081:Liposomes / Q000378:metabolism; D021122:Protein Subunits / Q000302:isolation & purification / Q000378:metabolism; D006180:Proton-Translocating ATPases / Q000302:isolation & purification / Q000378:metabolism*; D012964:Sodium / Q000378:metabolism*; D019712:Thermococcus / Q000201:enzymology* / Q000378:metabolism
chemical_list: D008081:Liposomes; D021122:Protein Subunits; D000255:Adenosine Triphosphate; D012964:Sodium; D006180:Proton-Translocating ATPases
keywords: ATP Synthase; Archaea; Membrane Protein; Protein Purification; Proteoliposomes; Sodium Transport