Subunit composition of vacuolar membrane H(+)-ATPase from mung bean.
The vacuolar H(+)-ATPase from mung bean hypocotyls was solubilized from the membrane with lysophosphatidycholine and purified by QAE-Toyopearl column chromatography. The purified ATPase was active only in the presence of exogenous phospholipid and was inhibited by nitrate, dicyclohexyl carbodiimide and Triton X-100, but not by vanadate or azide. Dodecyl sulfate/polyacrylamide gel electrophoresis of the purified ATPase yielded ten polypeptides of molecular masses of 68 kDa, 57 kDa, 44 kDa, 43 kDa, 38 kDa, 37 kDa 32 kDa, 16 kDa, 13 kDa and 12 kDa. All polypeptides remained in the peak activity fraction after glycerol density gradient centrifugation. Nine of them, excluding the 43-kDa polypeptide, comigrated in a polyacrylamide gradient gel in the presence of 0.1% Triton X-100. The 16-kDa polypeptide could be labeled with [14C]dicyclohexylcarbodiimide. The amino-terminal amino acid sequence of the isolated 68-kDa polypeptide generally agreed with that deduced from the cDNA for the carrot 69-kDa subunit [Zimniak, L., Dittrich, P., Gogarten, J. P., Kibak, H. & Taiz, L. (1988) J. Biol. Chem. 263, 9102-9112]. Thus, mung bean vacuolar H(+)-ATPase seems to consist of nine distinct subunits.
Matsuura-Endo
C
C
Institute of Low Temperature Science, Hokkaido University, Sapporo, Japan.
Maeshima
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M
Yoshida
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D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D004024:Dicyclohexylcarbodiimide / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation / Q000187:drug effects; D007887:Fabaceae / Q000201:enzymology*; D008969:Molecular Sequence Data; D009566:Nitrates / Q000494:pharmacology; D010743:Phospholipids / Q000494:pharmacology; D010946:Plants, Medicinal; D011487:Protein Conformation; D006180:Proton-Translocating ATPases / Q000037:antagonists & inhibitors / Q000302:isolation & purification*; D013379:Substrate Specificity; D014617:Vacuoles / Q000201:enzymology*
D000596:Amino Acids; D009566:Nitrates; D010743:Phospholipids; D004024:Dicyclohexylcarbodiimide; D006180:Proton-Translocating ATPases