H+-ATPase and Auxin-stimulated ATPase in Membrane Fractions from Zucchini (Cucurbita pepo L.) and Pumpkin (Cucurbita maxima L.) hypocotyls

We separated two membrane fractions containing H⁺-ATPase. One fraction with H⁺-ATPase was enriched in plasma membranes while the other one contained no plasma membranes. Since mitochondrial or plastidial origin of H⁺-ATPase could be excluded, the latter one presumably contained H⁺-ATPase in tonoplast vesicles.

With a plasma membrane-enriched fraction, the effect of auxin on ATPase activity was investigated. Auxin decreased the apparent K_M of ATPase so that auxin stimulation of ATPase activity can be measured best at very low ATP concentrations (< 50 μM). The stimulation of ATPase activity is specific for auxin in that 2,3-dichlorophenoxyacetic acid, a weak auxin, had a much lower effects than 2,4-dichlorophenoxyacetic acid or indolyl-3-acetic acid.

These experiments support the hypothesis that auxin stimulates H⁺-extrusion by stimulation of a H⁺-ATPase in vivo and in vitro.