Skip to main content
PMC home page
Plant Physiology logoLink to Plant Physiology
. 1995 Sep;109(1):285–292. doi: 10.1104/pp.109.1.285

The Tonoplast H+-ATPase of Acer pseudoplatanus Is a Vacuolar-Type ATPase That Operates with a Phosphoenzyme Intermediate.

T Magnin 1, A Fraichard 1, C Trossat 1, A Pugin 1
PMCID: PMC157587  PMID: 12228595

Abstract

The tonoplast H+-ATPase of Acer pseudoplatanus has been purified from isolated vacuoles. After solubilization, the purification procedure included size-exclusion and ion-exchange chromatography. The H+-ATPase consists of at least eight subunits, of 95, 66, 56, 54, 40, 38, 31, and 16 kD, that did not cross-react with polyclonal antibodies raised to the plasmalemma ATPase of Arabidopsis thaliana. The 66-kD polypeptide cross-reacted with monoclonal antibodies raised to the 70-kD subunit of the vacuolar H+-ATPase of oat roots. The functional molecular size of the tonoplast H+-ATPase, analyzed in situ by radiation inactivation, was found to be around 400 kD. The 66-kD subunit of the tonoplast H+-ATPase was rapidly phosphorylated by [[gamma]-32P]ATP in vitro. The complete loss of radio-activity in the 66-kD subunit after a short pulse-chase experiment with unlabeled ATP reflected a rapid turnover, which characterizes a phosphorylated intermediate. Phosphoenzyme formed from ATP is an acylphosphate-type compound as shown by its sensitivity to hydroxylamine and alkaline pH. These results lead us to suggest that the tonoplast H+-ATPase of A. pseudoplatanus is a vacuolar-type ATPase that could operate with a plasmalemma-type ATPase catalytic mechanism.

Full Text

The Full Text of this article is available as a PDF (1.8 MB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Amory A., Foury F., Goffeau A. The purified plasma membrane ATPase of the yeast Schizosaccharomyces pombe forms a phosphorylated intermediate. J Biol Chem. 1980 Oct 10;255(19):9353–9357. [PubMed] [Google Scholar]
  2. Arai H., Terres G., Pink S., Forgac M. Topography and subunit stoichiometry of the coated vesicle proton pump. J Biol Chem. 1988 Jun 25;263(18):8796–8802. [PubMed] [Google Scholar]
  3. Beltrán C., Nelson N. The membrane sector of vacuolar H(+)-ATPase by itself is impermeable to protons. Acta Physiol Scand Suppl. 1992;607:41–47. [PubMed] [Google Scholar]
  4. Bowman B. J., Dschida W. J., Harris T., Bowman E. J. The vacuolar ATPase of Neurospora crassa contains an F1-like structure. J Biol Chem. 1989 Sep 15;264(26):15606–15612. [PubMed] [Google Scholar]
  5. Chanson A., Pilet P. E. Target Molecular Size and Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis Analysis of the ATP-and Pyrophosphate-Dependent Proton Pumps from Maize Root Tonoplast. Plant Physiol. 1989 Jul;90(3):934–938. doi: 10.1104/pp.90.3.934. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Forgac M. V-type ATPases. Introduction. J Bioenerg Biomembr. 1992 Aug;24(4):339–340. doi: 10.1007/BF00762526. [DOI] [PubMed] [Google Scholar]
  7. Galloway C. J., Dean G. E., Marsh M., Rudnick G., Mellman I. Acidification of macrophage and fibroblast endocytic vesicles in vitro. Proc Natl Acad Sci U S A. 1983 Jun;80(11):3334–3338. doi: 10.1073/pnas.80.11.3334. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Gogarten J. P., Kibak H., Dittrich P., Taiz L., Bowman E. J., Bowman B. J., Manolson M. F., Poole R. J., Date T., Oshima T. Evolution of the vacuolar H+-ATPase: implications for the origin of eukaryotes. Proc Natl Acad Sci U S A. 1989 Sep;86(17):6661–6665. doi: 10.1073/pnas.86.17.6661. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Ishikawa T. The ATP-dependent glutathione S-conjugate export pump. Trends Biochem Sci. 1992 Nov;17(11):463–468. doi: 10.1016/0968-0004(92)90489-v. [DOI] [PubMed] [Google Scholar]
  10. Kaestner K. H., Randall S. K., Sze H. N,N'-dicyclohexylcarbodiimide-binding proteolipid of the vacuolar H+-ATPase from oat roots. J Biol Chem. 1988 Jan 25;263(3):1282–1287. [PubMed] [Google Scholar]
  11. Kepner G. R., Macey R. I. Membrane enzyme systems. Molecular size determinations by radiation inactivation. Biochim Biophys Acta. 1968 Sep 17;163(2):188–203. doi: 10.1016/0005-2736(68)90097-7. [DOI] [PubMed] [Google Scholar]
  12. Kobayashi K., Sogame Y., Hara H., Hayashi K. Mechanism of glutathione S-conjugate transport in canalicular and basolateral rat liver plasma membranes. J Biol Chem. 1990 May 15;265(14):7737–7741. [PubMed] [Google Scholar]
  13. LAMPORT D. T. CELL SUSPENSION CULTURES OF HIGHER PLANTS: ISOLATION AND GROWTH ENERGETICS. Exp Cell Res. 1964 Jan;33:195–206. doi: 10.1016/s0014-4827(64)81026-0. [DOI] [PubMed] [Google Scholar]
  14. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  15. Lübben M., Lünsdorf H., Schäfer G. The plasma membrane ATPase of the thermoacidophilic archaebacterium Sulfolobus acidocaldarius. Purification and immunological relationships to F1-ATPases. Eur J Biochem. 1987 Sep 1;167(2):211–219. doi: 10.1111/j.1432-1033.1987.tb13325.x. [DOI] [PubMed] [Google Scholar]
  16. Mandala S., Taiz L. Characterization of the subunit structure of the maize tonoplast ATPase. Immunological and inhibitor binding studies. J Biol Chem. 1986 Sep 25;261(27):12850–12855. [PubMed] [Google Scholar]
  17. Mandala S., Taiz L. Proton transport in isolated vacuoles from corn coleoptiles. Plant Physiol. 1985 May;78(1):104–109. doi: 10.1104/pp.78.1.104. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Manolson M. F., Rea P. A., Poole R. J. Identification of 3-O-(4-benzoyl)benzoyladenosine 5'-triphosphate- and N,N'-dicyclohexylcarbodiimide-binding subunits of a higher plant H+-translocating tonoplast ATPase. J Biol Chem. 1985 Oct 5;260(22):12273–12279. [PubMed] [Google Scholar]
  19. McIntyre J. O., Churchill P. Glucose-6-phosphate dehydrogenase from Leuconostoc mesenteroides is a reliable internal standard for radiation-inactivation studies of membranes in the frozen state. Anal Biochem. 1985 Jun;147(2):468–477. doi: 10.1016/0003-2697(85)90300-8. [DOI] [PubMed] [Google Scholar]
  20. Medda P., Hasselbach W. Dependence on membrane lipids of the effect of vanadate on calcium and ATP binding to sarcoplasmic reticulum ATPase. Z Naturforsch C. 1984 Nov-Dec;39(11-12):1137–1140. doi: 10.1515/znc-1984-11-1224. [DOI] [PubMed] [Google Scholar]
  21. Mellman I., Fuchs R., Helenius A. Acidification of the endocytic and exocytic pathways. Annu Rev Biochem. 1986;55:663–700. doi: 10.1146/annurev.bi.55.070186.003311. [DOI] [PubMed] [Google Scholar]
  22. Montrichard F., Pugin A., Gaudemer Y. Inhibition of the vacuolar ATPase of Acer pseudoplatanus cells by vanadate. Biochimie. 1989 Jul;71(7):813–817. doi: 10.1016/0300-9084(89)90044-8. [DOI] [PubMed] [Google Scholar]
  23. Moriyama Y., Nelson N. Cold inactivation of vacuolar proton-ATPases. J Biol Chem. 1989 Feb 25;264(6):3577–3582. [PubMed] [Google Scholar]
  24. Moriyama Y., Nelson N. H+-translocating ATPase in Golgi apparatus. Characterization as vacuolar H+-ATPase and its subunit structures. J Biol Chem. 1989 Nov 5;264(31):18445–18450. [PubMed] [Google Scholar]
  25. Moriyama Y., Nelson N. Purification and properties of a vanadate- and N-ethylmaleimide-sensitive ATPase from chromaffin granule membranes. J Biol Chem. 1988 Jun 15;263(17):8521–8527. [PubMed] [Google Scholar]
  26. Mukohata Y., Ihara K., Yoshida M., Konishi J., Sugiyama Y., Yoshida M. The halobacterial H+-translocating ATP synthase relates to the eukaryotic anion-sensitive H+-ATPase. Arch Biochem Biophys. 1987 Dec;259(2):650–653. doi: 10.1016/0003-9861(87)90532-7. [DOI] [PubMed] [Google Scholar]
  27. Nelson N., Taiz L. The evolution of H+-ATPases. Trends Biochem Sci. 1989 Mar;14(3):113–116. doi: 10.1016/0968-0004(89)90134-5. [DOI] [PubMed] [Google Scholar]
  28. Ohkuma S., Moriyama Y., Takano T. Identification and characterization of a proton pump on lysosomes by fluorescein-isothiocyanate-dextran fluorescence. Proc Natl Acad Sci U S A. 1982 May;79(9):2758–2762. doi: 10.1073/pnas.79.9.2758. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Parry R. V., Turner J. C., Rea P. A. High purity preparations of higher plant vacuolar H+-ATPase reveal additional subunits. Revised subunit composition. J Biol Chem. 1989 Nov 25;264(33):20025–20032. [PubMed] [Google Scholar]
  30. Sarafian V., Potier M., Poole R. J. Radiation-inactivation analysis of vacuolar H(+)-ATPase and H(+)-pyrophosphatase from Beta vulgaris L. Functional sizes for substrate hydrolysis and for H+ transport. Biochem J. 1992 Apr 15;283(Pt 2):493–497. doi: 10.1042/bj2830493. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Sherwood J. B., Shouval D. Continuous production of erythropoietin by an established human renal carcinoma cell line: development of the cell line. Proc Natl Acad Sci U S A. 1986 Jan;83(1):165–169. doi: 10.1073/pnas.83.1.165. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Uchida E., Ohsumi Y., Anraku Y. Characterization and function of catalytic subunit alpha of H+-translocating adenosine triphosphatase from vacuolar membranes of Saccharomyces cerevisiae. A study with 7-chloro-4-nitrobenzo-2-oxa-1,3-diazole. J Biol Chem. 1988 Jan 5;263(1):45–51. [PubMed] [Google Scholar]
  33. Wang M. Y., Lin Y. H., Chou W. M., Chung T. P., Pan R. L. Purification and characterization of tonoplast ATPase from etiolated mung bean seedlings. Plant Physiol. 1989 Jun;90(2):475–481. doi: 10.1104/pp.90.2.475. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Zimniak L., Dittrich P., Gogarten J. P., Kibak H., Taiz L. The cDNA sequence of the 69-kDa subunit of the carrot vacuolar H+-ATPase. Homology to the beta-chain of F0F1-ATPases. J Biol Chem. 1988 Jul 5;263(19):9102–9112. [PubMed] [Google Scholar]

Articles from Plant Physiology are provided here courtesy of Oxford University Press

RESOURCES