Isolation of H+ -translocating ATPase in tonoplast of Tradescantia virginiana L. leaf cells.
The tonoplast of Tradescantia virginiana L. was prepared from leaf cells and then solubilized with deoxycholate (DOC) and n-octyl-beta-D-glucoside (n-OG). Three major polypeptides (68, 60, 16 kDa) and several other minor components were isolated. These polypeptides were reconstituted in soybean phospholipids (asolectin). The H(+) pump activity was investigated with the reconstituted system as well as with the tonoplast. In both cases, the quinacrine-fluorescence quenching was observed in the presence of ATP-Mg(2+), indicating the H(+) pumping. The H(+) pump activity was inhibited by gramicidin D, a channel-forming ionophore, and by KNO(3), an inhibitor specific to tonoplast-type (V-type) H(+)-ATPase.
Saito
Mikako
M
Department of Biotechnology and Life Science, Tokyo University of Agriculture and Technology, 2-24-16 Nakamachi, Koganei, Tokyo 184-8588, Japan.
Matsuoka
Hideaki
H
Kasamo
Kunihiro
K
D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000737:chemistry; D006863:Hydrogen-Ion Concentration; D008081:Liposomes / Q000737:chemistry; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism*; D008970:Molecular Weight; D010455:Peptides / Q000737:chemistry; D018515:Plant Leaves / Q000737:chemistry / Q000378:metabolism*; D010940:Plant Proteins / Q000737:chemistry / Q000378:metabolism*; D006180:Proton-Translocating ATPases / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D034823:Tradescantia / Q000737:chemistry / Q000378:metabolism*
D004791:Enzyme Inhibitors; D008081:Liposomes; D008565:Membrane Proteins; D010455:Peptides; D010940:Plant Proteins; C077491:tonoplast intrinsic protein, plant; D006180:Proton-Translocating ATPases