title: Membrane pyrophosphatases from Thermotoga maritima and Vigna radiata suggest a conserved coupling mechanism.
abstract: Membrane-bound pyrophosphatases (M-PPases), which couple proton/sodium ion transport to pyrophosphate synthesis/hydrolysis, are important in abiotic stress resistance and in the infectivity of protozoan parasites. Here, three M-PPase structures in different catalytic states show that closure of the substrate-binding pocket by helices 5-6 affects helix 13 in the dimer interface and causes helix 12 to move down. This springs a 'molecular mousetrap', repositioning a conserved aspartate and activating the nucleophilic water. Corkscrew motion at helices 6 and 16 rearranges the key ionic gate residues and leads to ion pumping. The pumped ion is above the ion gate in one of the ion-bound structures, but below it in the other. Electrometric measurements show a single-turnover event with a non-hydrolysable inhibitor, supporting our model that ion pumping precedes hydrolysis. We propose a complete catalytic cycle for both proton and sodium-pumping M-PPases, and one that also explains the basis for ion specificity.
authors.affiliation: Department of Life Science and Institute of Bioinformatics and Structural Biology, College of Life Science, National Tsing Hua University, Hsinchu 30013, Taiwan.
authors.forename: Kun-Mou
authors.initials: KM
authors.lastname: Li
authors.affiliation: Astbury Centre for Structural Molecular Biology, School of Biomedical Sciences, University of Leeds, Leeds LS2 9JT, England.
authors.forename: Craig
authors.initials: C
authors.lastname: Wilkinson
authors.affiliation: Division of Biochemistry, Department of Biosciences, University of Helsinki, Helsinki, FIN-00014, Finland.
authors.forename: Juho
authors.initials: J
authors.lastname: Kellosalo
authors.affiliation: Department of Life Science and Institute of Bioinformatics and Structural Biology, College of Life Science, National Tsing Hua University, Hsinchu 30013, Taiwan.
authors.forename: Jia-Yin
authors.initials: JY
authors.lastname: Tsai
authors.affiliation: Institute of Biotechnology, University of Helsinki, PO Box 65, Helsinki, FIN-00014, Finland.
authors.forename: Tommi
authors.initials: T
authors.lastname: Kajander
authors.affiliation: Astbury Centre for Structural Molecular Biology, School of Biomedical Sciences, University of Leeds, Leeds LS2 9JT, England.
authors.forename: Lars J C
authors.initials: LJ
authors.lastname: Jeuken
authors.affiliation: Department of Life Science and Institute of Bioinformatics and Structural Biology, College of Life Science, National Tsing Hua University, Hsinchu 30013, Taiwan.
authors.forename: Yuh-Ju
authors.initials: YJ
authors.lastname: Sun
authors.affiliation: Astbury Centre for Structural Molecular Biology, School of Biomedical Sciences, University of Leeds, Leeds LS2 9JT, England.|Division of Biochemistry, Department of Biosciences, University of Helsinki, Helsinki, FIN-00014, Finland.
authors.forename: Adrian
authors.identifier: 0000-0001-8032-9700
authors.initials: A
authors.lastname: Goldman
mesh_terms: D055162:Biocatalysis; D002462:Cell Membrane / Q000201:enzymology*; D018360:Crystallography, X-Ray; D006860:Hydrogen Bonding; D006868:Hydrolysis; D007477:Ions; D007700:Kinetics; D008958:Models, Molecular; D017433:Protein Structure, Secondary; D017494:Proton Pumps / Q000378:metabolism; D011755:Pyrophosphatases / Q000737:chemistry / Q000378:metabolism*; D012964:Sodium / Q000378:metabolism; D020124:Thermotoga maritima / Q000201:enzymology*; D000070658:Vigna / Q000201:enzymology*
chemical_list: D007477:Ions; D017494:Proton Pumps; D012964:Sodium; D011755:Pyrophosphatases