title: Structure and mechanism of Zn2+-transporting P-type ATPases.
abstract: Zinc is an essential micronutrient for all living organisms. It is required for signalling and proper functioning of a range of proteins involved in, for example, DNA binding and enzymatic catalysis. In prokaryotes and photosynthetic eukaryotes, Zn(2+)-transporting P-type ATPases of class IB (ZntA) are crucial for cellular redistribution and detoxification of Zn(2+) and related elements. Here we present crystal structures representing the phosphoenzyme ground state (E2P) and a dephosphorylation intermediate (E2·Pi) of ZntA from Shigella sonnei, determined at 3.2 Å and 2.7 Å resolution, respectively. The structures reveal a similar fold to Cu(+)-ATPases, with an amphipathic helix at the membrane interface. A conserved electronegative funnel connects this region to the intramembranous high-affinity ion-binding site and may promote specific uptake of cellular Zn(2+) ions by the transporter. The E2P structure displays a wide extracellular release pathway reaching the invariant residues at the high-affinity site, including C392, C394 and D714. The pathway closes in the E2·Pi state, in which D714 interacts with the conserved residue K693, which possibly stimulates Zn(2+) release as a built-in counter ion, as has been proposed for H(+)-ATPases. Indeed, transport studies in liposomes provide experimental support for ZntA activity without counter transport. These findings suggest a mechanistic link between PIB-type Zn(2+)-ATPases and PIII-type H(+)-ATPases and at the same time show structural features of the extracellular release pathway that resemble PII-type ATPases such as the sarcoplasmic/endoplasmic reticulum Ca(2+)-ATPase (SERCA) and Na(+), K(+)-ATPase. These findings considerably increase our understanding of zinc transport in cells and represent new possibilities for biotechnology and biomedicine.
authors.affiliation: 1] Centre for Membrane Pumps in Cells and Disease (PUMPkin), Danish National Research Foundation, Aarhus University, Department of Molecular Biology and Genetics, Gustav Wieds Vej 10C, DK-8000 Aarhus C, Denmark [2] Department of Biomedical Sciences, University of Copenhagen, Blegdamsvej 3B, DK-2200 Copenhagen, Denmark (K.W. and P.G.); Department of Experimental Medical Science, Lund University, Sölvegatan 19, SE-221 84 Lund, Sweden (P.G.). [3].
authors.forename: Kaituo
authors.initials: K
authors.lastname: Wang
authors.affiliation: 1] Centre for Membrane Pumps in Cells and Disease (PUMPkin), Danish National Research Foundation, Aarhus University, Department of Molecular Biology and Genetics, Gustav Wieds Vej 10C, DK-8000 Aarhus C, Denmark [2].
authors.forename: Oleg
authors.initials: O
authors.lastname: Sitsel
authors.affiliation: Centre for Membrane Pumps in Cells and Disease (PUMPkin), Danish National Research Foundation, Aarhus University, Department of Molecular Biology and Genetics, Gustav Wieds Vej 10C, DK-8000 Aarhus C, Denmark.
authors.forename: Gabriele
authors.initials: G
authors.lastname: Meloni
authors.affiliation: Centre for Membrane Pumps in Cells and Disease (PUMPkin), Danish National Research Foundation, Aarhus University, Department of Molecular Biology and Genetics, Gustav Wieds Vej 10C, DK-8000 Aarhus C, Denmark.
authors.forename: Henriette Elisabeth
authors.initials: HE
authors.lastname: Autzen
authors.affiliation: Science for Life Laboratory, Department of Theoretical Physics, Swedish e-Science Research Center, KTH Royal Institute of Technology, SE-171 21 Solna, Sweden.
authors.forename: Magnus
authors.initials: M
authors.lastname: Andersson
authors.affiliation: Centre for Membrane Pumps in Cells and Disease (PUMPkin), Danish National Research Foundation, Aarhus University, Department of Molecular Biology and Genetics, Gustav Wieds Vej 10C, DK-8000 Aarhus C, Denmark.
authors.forename: Tetyana
authors.initials: T
authors.lastname: Klymchuk
authors.affiliation: Centre for Membrane Pumps in Cells and Disease (PUMPkin), Danish National Research Foundation, Aarhus University, Department of Molecular Biology and Genetics, Gustav Wieds Vej 10C, DK-8000 Aarhus C, Denmark.
authors.forename: Anna Marie
authors.initials: AM
authors.lastname: Nielsen
authors.affiliation: Division of Chemistry and Chemical Engineering and Howard Hughes Medical Institute, California Institute of Technology, 1200 East California Boulevard, Pasadena, California 91125, USA.
authors.forename: Douglas C
authors.initials: DC
authors.lastname: Rees
authors.affiliation: Centre for Membrane Pumps in Cells and Disease (PUMPkin), Danish National Research Foundation, Aarhus University, Department of Molecular Biology and Genetics, Gustav Wieds Vej 10C, DK-8000 Aarhus C, Denmark.
authors.forename: Poul
authors.initials: P
authors.lastname: Nissen
authors.affiliation: 1] Centre for Membrane Pumps in Cells and Disease (PUMPkin), Danish National Research Foundation, Aarhus University, Department of Molecular Biology and Genetics, Gustav Wieds Vej 10C, DK-8000 Aarhus C, Denmark [2] Department of Biomedical Sciences, University of Copenhagen, Blegdamsvej 3B, DK-2200 Copenhagen, Denmark (K.W. and P.G.); Department of Experimental Medical Science, Lund University, Sölvegatan 19, SE-221 84 Lund, Sweden (P.G.).
authors.forename: Pontus
authors.initials: P
authors.lastname: Gourdon
mesh_terms: D000251:Adenosine Triphosphatases / Q000737:chemistry* / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism; D001665:Binding Sites; D002104:Cadmium / Q000378:metabolism; D000252:Calcium-Transporting ATPases / Q000737:chemistry; D017124:Conserved Sequence; D018360:Crystallography, X-Ray; D007854:Lead / Q000378:metabolism; D008958:Models, Molecular; D010766:Phosphorylation; D011510:Proteolipids / Q000737:chemistry / Q000378:metabolism; D006180:Proton-Translocating ATPases / Q000737:chemistry / Q000378:metabolism; D012760:Shigella / Q000201:enzymology*; D000254:Sodium-Potassium-Exchanging ATPase / Q000737:chemistry; D015032:Zinc / Q000378:metabolism
chemical_list: D011510:Proteolipids; C020485:proteoliposomes; D002104:Cadmium; D007854:Lead; D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases; C109898:Zn(II)-translocating P-type ATPase; D006180:Proton-Translocating ATPases; D000252:Calcium-Transporting ATPases; D000254:Sodium-Potassium-Exchanging ATPase; D015032:Zinc