DEMONSTRATION OF A METABOLICALLY ACTIVE GLUCOSE-B-PHOSPHATE POOL IN THE LUMEN OF LIVER MICROSOMAL VESICLES

Glucose-6-phosphate transport was investigated in rat or human liver m icrosomal vesicles using rapid filtration and light-scattering methods , Upon addition of glucose-6-phosphate, rat liver microsomes accumulat ed the radioactive tracer, reaching a steady-state level of uptake, In this phase, the majority of the accumulated tracer was glucose, but a significant intraluminal glucose-6-phosphate pool could also be obser ved, The extent of the intravesicular glucose pool was proportional wi th glucose-6-phosphatase activity. The relative size of the intravesic ular glucose-6-phosphate pool (irrespective of the concentration of th e extravesicular concentration of added glucose-6-phosphate) expressed as the apparent intravesicular space of the hexose phosphate was inve rsely dependent on glucose-6-phosphatase activity, The increase of hyd rolysis by elevating the extravesicular glucose-6-phosphate concentrat ion or temperature resulted in lower apparent intravesicular glucose-6 -phosphate spaces and, thus, in a higher transmembrane gradient of glu cose-6-phosphate concentrations, In contrast, inhibition of glucose-6- phosphate hydrolysis by vanadate, inactivation of glucose-6-phosphatas e by acidic pH, or genetically determined low or absent glucose-6-phos phatase activity in human hepatic microsomes of patients suffering fro m glycogen storage disease type 1a led to relatively high intravesicul ar glucose-6-phosphate levels, Glucose-6-phosphate transport investiga ted by light-scattering technique resulted in similar traces in contro l and vanadate-treated rat microsomes as well as in microsomes from hu man patients with glycogen storage disease type 1a, It is concluded th at liver microsomes take up glucose-6-phosphate, constituting a pool d irectly accessible to intraluminal glucose-6-phosphatase activity. In addition, normal glucose-6-phosphate uptake can take place in the abse nce of the glucose-6-phosphatase enzyme protein, confirming the existe nce of separate transport proteins.