INTERACTION OF FACILITATIVE GLUCOSE-TRANSPORTER WITH GLUCOKINASE AND ITS MODULATION BY ADP AND GLUCOSE-6-PHOSPHATE

Bacterial glucokinase (GK) binds to purified, human erythrocyte glucos e transporter (GT) reconstituted in vesicles. The binding is largely a bolished if GT is predigested with trypsin, indicating that GK binds t o the cytoplasmic domain of GT. The binding is a saturable function of GK concentration showing two distinct affinities with apparent K(D) o f 0.33 and 5.1 muM. The binding is stimulated by an increasing concent ration of ADP with the 50% maximal effect at 5 mM. Glucose-6-phosphate (G6P) also stimulates the binding with a distinct optimum at 25 mM. T he binding is stimulated only slightly by ATP. D-glucose has no affect on the binding. KCl enhances the binding with the maximal effect at p hysiological intracellular concentrations. The binding is sensitive to changes in pH with an optimum at pH 4. The binding causes no detectab le functional change in GT. However, the enzymatic activity of GK meas ured at nanomolar concentrations of GK is significantly greater in the presence of GT vesicles than in its absence or in the presence of pro tein-free vesicles, indicating that GK interacts with GT at this low c oncentration range with an apparent K(D) of 10 mM. Although its physio logical significance is not known, the GK-GT interaction in vitro desc ribed here suggests that these two proteins may also interact in the c ell and regulate carbohydrate metabolism. (C) 1993 Wiley-Liss, Inc.