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. 2010 Dec 13;107(52):22728–22733. doi: 10.1073/pnas.1007747108

Fig. 3.

Fig. 3.

Preparation of vacuolar membranes and vacuolar sequestration assay. (A) Protein gel-blot analysis of selected fractions probed with antibodies against V-PPase, Bip, and plasma membrane H+-ATPase. (B) Portions of HPLC traces showing recovered coniferyl alcohol 4-O-glucoside (coniferin) from the vacuolar vesicles in the sequestering assay in the presence (a) and absence (b) of ATP, the presence of sodium vanadate and ATP (c), and the presence of gramicidin D and ATP (d). (C) Effects of the membrane transporter inhibitors sodium vanadate (1 mM) and gramicidin D (5 μM) on the ATP-dependent uptake of coniferin by vacuolar vesicles. Data were from three replicates. *Significant changes at P < 0.01 under t test. (D and E) The kinetics of transport of coniferin into vacuolar vesicles for coniferin (D) and MgATP (E) concentrations. The data are from two or three replicates and are plotted by nonlinear regression analysis fit to the Michaelis–Menten equation. Insets show Lineweaver–Burk plots.