title: Sodium and proton effects on inward proton transport through Na/K pumps.
abstract: The Na/K pump hydrolyzes ATP to export three intracellular Na (Nai) as it imports two extracellular K (Ko) across animal plasma membranes. Within the protein, two ion-binding sites (sites I and II) can reciprocally bind Na or K, but a third site (site III) exclusively binds Na in a voltage-dependent fashion. In the absence of Nao and Ko, the pump passively imports protons, generating an inward current (IH). To elucidate the mechanisms of IH, we used voltage-clamp techniques to investigate the [H]o, [Na]o, and voltage dependence of IH in Na/K pumps from ventricular myocytes and in ouabain-resistant pumps expressed in Xenopus oocytes. Lowering pHo revealed that Ho both activates IH (in a voltage-dependent manner) and inhibits it (in a voltage-independent manner) by binding to different sites. Nao effects depend on pHo; at pHo where no Ho inhibition is observed, Nao inhibits IH at all concentrations, but when applied at pHo that inhibits pump-mediated current, low [Na]o activates IH and high [Na]o inhibits it. Our results demonstrate that IH is a property inherent to Na/K pumps, not linked to the oocyte expression environment, explains differences in the characteristics of IH previously reported in the literature, and supports a model in which 1), protons leak through site III; 2), binding of two Na or two protons to sites I and II inhibits proton transport; and 3), pumps with mixed Na/proton occupancy of sites I and II remain permeable to protons.
authors.affiliation: Department of Cell and Molecular Physiology, Texas Tech University Health Sciences Center, Lubbock, Texas; School of Biological Sciences. Illinois State University, Normal, Illinois.
authors.forename: Travis J
authors.initials: TJ
authors.lastname: Mitchell
authors.affiliation: Department of Cell and Molecular Physiology, Texas Tech University Health Sciences Center, Lubbock, Texas.
authors.forename: Camila
authors.initials: C
authors.lastname: Zugarramurdi
authors.affiliation: Department of Cell and Molecular Physiology, Texas Tech University Health Sciences Center, Lubbock, Texas.
authors.forename: J Fernando
authors.initials: JF
authors.lastname: Olivera
authors.affiliation: School of Biological Sciences. Illinois State University, Normal, Illinois.
authors.forename: Craig
authors.initials: C
authors.lastname: Gatto
authors.affiliation: Department of Cell and Molecular Physiology, Texas Tech University Health Sciences Center, Lubbock, Texas. Electronic address: pablo.artigas@ttuhsc.edu.
authors.forename: Pablo
authors.initials: P
authors.lastname: Artigas
mesh_terms: D000818:Animals; D006168:Guinea Pigs; D006352:Heart Ventricles / Q000166:cytology; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D015640:Ion Channel Gating / Q000187:drug effects; D017136:Ion Transport / Q000187:drug effects; D032383:Myocytes, Cardiac / Q000378:metabolism; D010042:Ouabain / Q000494:pharmacology; D018408:Patch-Clamp Techniques; D011522:Protons; D012964:Sodium / Q000494:pharmacology*; D000254:Sodium-Potassium-Exchanging ATPase / Q000378:metabolism*; D014981:Xenopus
chemical_list: D011522:Protons; D010042:Ouabain; D012964:Sodium; D000254:Sodium-Potassium-Exchanging ATPase