skip to main content
Primo Search
Search in: Busca Geral

Solubilization and Functional Reconstitution of the Tonoplast H +-ATPase from Citrus in Liposomes

Bañuls, Josefina ; Ratajczak, Rafael ; Lüttge, Ulrich

Journal of Plant Physiology, 1994, Vol.144(1), pp.74-79 [Periódico revisado por pares]

Texto completo disponível

Citações Citado por
  • Título:
    Solubilization and Functional Reconstitution of the Tonoplast H +-ATPase from Citrus in Liposomes
  • Autor: Bañuls, Josefina ; Ratajczak, Rafael ; Lüttge, Ulrich
  • Assuntos: Citrus Sinensis (L) ; H + -Atpase ; Reconstitution ; Tonoplast ; ATP ; Ctp ; Dccd ; Dtt ; Gtp ; Idp ; Ppi ; SDS-Page ; Utp
  • É parte de: Journal of Plant Physiology, 1994, Vol.144(1), pp.74-79
  • Descrição: Proteoliposomes were formed after solubilization of tonoplast proteins from Citrus with octylglucoside in the presence of soybean phospholipids (phosphatidylcholine), with removal of detergent by passage through an Amberlite XAD-2 column. The specific ATP-hydrolysis activity of the proteoliposomes was increased 4-fold over the activity of the native tonoplast vesicles. Pumping of protons by the reconstituted ATPase was demonstrated by quinacrine-fluorescence quenching. Neither vanadate nor azide inhibited ATP-hydrolysis and transport activity. Both the ATP-hydrolysis and the H +-translocating activities were inhibited by nitrate, the first activity being the more sensitive one. Both activities were completely inhibited by bafilomycin A 1, an inhibitor of vacuolar type ATPase, at concentrations of 20 nM. A potent inhibitor of H +-transport, DCCD, completely inhibited both activities at 100 gM and with halfmaximal inhibition at concentrations of 40 pM and 60 μM for the H +-transport and the ATP-hydrolysis activities, respectively. The reconstituted H+-ATPase was stimulated by anions (Cl - > Br - > SO 4 2−), inhibited by NO 3 − and I − and largely insensitive to monovalent cations. The proteoliposomes showed no PP iase activity. SDS-PAGE showed that the reconstituted H +-ATPase from Citrus contained polypeptides of apparent molecular masses of 63, 52, 37, 33 and 16 kDa that cross-reacted with an antiserum against the tonoplast ATPase from Kalanchoë daigremontiana. Freeze-fracture electron microscopy of the proteoliposomes showed that proteins were incorporated into the bilayer membrane. The proteoliposomes from Citrus were mainly vesicles intramembraneous with particles whose diameter ranged from 6 to 7 nm. The results conclusively demonstrate a selective reconstitution of a functional H +-ATPase from Citrus.
  • Idioma: Inglês

Buscando em bases de dados remotas. Favor aguardar.