RECONSTITUTION OF THE HEXOSE PHOSPHATE TRANSLOCATOR FROM THE ENVELOPEMEMBRANES OF WHEAT ENDOSPERM AMYLOPLASTS

Amyloplasts were isolated and purified from wheat endosperm and the en velope membranes reconstituted into liposomes. Envelope membranes were solubilized in n-octyl beta-D-glucopyranoside and mixed with liposome s supplemented with 5.6 mol% cholesterol to produce proteoliposomes of defined size, which showed negligible leakage of internal substrates. Transport experiments with proteoliposomes revealed a counter-exchang e of glucose 1-phosphate (Glc1P), glucose 6-phosphate (Glc6P), inorgan ic phosphate (P-i), 3-phosphoglycerate and dihydroxyacetone phosphate. The Glc1P/P-i counter-exchange reaction exhibited an apparent K-m for Glc1P of 0.4 mM. Glc6P was a competitive inhibitor of Glc1P transport (K-i 0.8 mM), and the two hexose phosphates could exchange with each other, indicating the operation of a single carrier protein. Glc1P/P-i antiport in proteoliposomes showed an exchange stoichiometry at pH 8. 0 of 1 mol of phosphate transported per mol of sugar phosphate.