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The prime plasmalemma ATPase of the halophilic alga Dunaliella bioculata: purification and characterization

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Abstract

The prime plasmalemma ATPase of the halophilic green alga Dunaliella bioculata has been solubilized by Triton X-100 from a plasmalemma-rich membrane fraction and purified by anion-exchange chromatography. Vanadate-sensitive ATPase activity was totally enriched about 230-fold to a specific activity of approx. 250 nkat·mg protein−1. The presence of Mg2+ or Mn2+ is essential for ATP hydrolysis by the enzyme. In addition to an equimolar requirement (1∶1 Mg2+: ATP), there is further stimulation by Mg2+ (up to 20 mM) and by (100 mM) monovalent cations (K+ ∼ NH +4 >Rb+ ∼-Na+ >Cs+ >Li+∼-choline+). Most anions have no or little effect. With a molecular mass of about 105 kDa for the single subunit, sensitivity to vanadate and N,N′-dicyclohexylcarbodiimide (50% inhibition at about 1 μM and 0.3 mM, respectively), strict ATP-specificity, and an acidic pH optimum, this enzyme shows the typical characteristics of the common type of H+-ATPase in the plasmalemma of higher plants and fungi. These results undermine the hypothesis of a wider distribution of a special (high salt) type of plasmalemma ATPase as found in the marine alga Acetabularia.

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Abbreviations

BTP:

1,3-bis[tris(hydroxymethyl)-methylamino]propane

DCCD:

N,N′-dicyclohexylcarbodiimide

DES:

diethylstilbestrol

Mega-9:

nonanoyl-N-methyl-glucamide

Mes:

N-morpholinoethanesulfonic acid

Mops:

N-morpholinopropanesulfonic acid

PAGE:

polyacrylamide-gel electrophoresis

PM:

plasmalemma-enriched membrane fraction

SDS:

sodium dodecyl sulfate

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Authors and Affiliations

  1. Pflanzenphysiologisches Institut der Universität Göttingen, Untere Karspüle 2, D-3400, Göttingen, Federal Republic of Germany

    Martin Smahel, Antje Hamann & Dietrich Gradmann

Authors
  1. Martin Smahel
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  2. Antje Hamann
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  3. Dietrich Gradmann
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Additional information

This work was supported by the Deutsche Forschungsgemeinschaft; we thank Drs. M. Ikeda and D. Oesterhelt (MPI für Biochemie, Martinsried, FRG) for generous and valuable information about their work prior to publication.

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Smahel, M., Hamann, A. & Gradmann, D. The prime plasmalemma ATPase of the halophilic alga Dunaliella bioculata: purification and characterization. Planta 181, 496–504 (1990). https://doi.org/10.1007/BF00193002

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  • DOI: https://doi.org/10.1007/BF00193002

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