title: Electrogenicity of Na,K- and H,K-ATPase activity and presence of a positively charged amino acid in the fifth transmembrane segment.
abstract: The transport activity of the Na,K-ATPase (a 3 Na+ for 2 K+ ion exchange) is electrogenic, whereas the closely related gastric and non-gastric H,K-ATPases perform electroneutral cation exchange. We have studied the role of a highly conserved serine residue in the fifth transmembrane segment of the Na,K-ATPase, which is replaced with a lysine in all known H,K-ATPases. Ouabain-sensitive 86Rb uptake and K+-activated currents were measured in Xenopus oocytes expressing the Bufo bladder H,K-ATPase or the Bufo Na,K-ATPase in which these residues, Lys800 and Ser782, respectively, were mutated. Mutants K800A and K800E of the H,K-ATPase showed K+-stimulated and ouabain-sensitive electrogenic transport. In contrast, when the positive charge was conserved (K800R), no K+-induced outward current could be measured, even though rubidium transport activity was present. Conversely, the S782R mutant of the Na,K-ATPase had non-electrogenic transport activity, whereas the S782A mutant was electrogenic. The K800S mutant of the H,K-ATPase had a more complex behavior, with electrogenic transport only in the absence of extracellular Na+. Thus, a single positively charged residue in the fifth transmembrane segment of the alpha-subunit can determine the electrogenicity and therefore the stoichiometry of cation transport by these ATPases.
authors.affiliation: Institute of Pharmacology and Toxicology, University of Lausanne, CH-1005 Lausanne, Switzerland.
authors.forename: Muriel
authors.initials: M
authors.lastname: Burnay
authors.forename: Gilles
authors.initials: G
authors.lastname: Crambert
authors.forename: Solange
authors.initials: S
authors.lastname: Kharoubi-Hess
authors.forename: Käthi
authors.initials: K
authors.lastname: Geering
authors.forename: Jean-Daniel
authors.initials: JD
authors.lastname: Horisberger
mesh_terms: D000818:Animals; D002024:Bufo marinus / Q000235:genetics; D002462:Cell Membrane / Q000201:enzymology; D004553:Electric Conductivity; D004563:Electrochemistry; D005260:Female; D015870:Gene Expression; D008239:Lysine; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D009865:Oocytes / Q000201:enzymology; D010042:Ouabain / Q000494:pharmacology; D011188:Potassium / Q000494:pharmacology; D006180:Proton-Translocating ATPases / Q000737:chemistry* / Q000235:genetics; D012414:Rubidium Radioisotopes / Q000378:metabolism; D012694:Serine / Q000737:chemistry*; D000254:Sodium-Potassium-Exchanging ATPase / Q000737:chemistry* / Q000235:genetics; D014162:Transfection; D001743:Urinary Bladder / Q000201:enzymology; D014981:Xenopus
chemical_list: D012414:Rubidium Radioisotopes; D012694:Serine; D010042:Ouabain; D006180:Proton-Translocating ATPases; D000254:Sodium-Potassium-Exchanging ATPase; D008239:Lysine; D011188:Potassium