Astrocytic glucose-6-phosphatase and the permeability of brain microsomes to glucose 6-phosphate.
Cells from primary rat astrocyte cultures express a 36.5 kDa protein that cross-reacts with polyclonal antibodies to the catalytic subunit of rat hepatic glucose-6-phosphatase on Western blotting. Glucose-6-phosphate-hydrolysing activity of the order of 10 nmol/min per mg of total cellular protein can be demonstrated in cell homogenates. This activity shows latency, and is localized to the microsomal fraction. Kinetic analysis shows a Km of 15 mM and a Vmax. of 30 nmol/min per mg of microsomal protein in disrupted microsomes. Approx. 40% of the total phosphohydrolase activity is specific glucose-6-phosphatase, as judged by sensitivity to exposure to pH 5 at 37 degrees C. Previous reports that the brain microsomal glucose-6-phosphatase system does not distinguish glucose 6-phosphate and mannose 6-phosphate are confirmed in astrocyte microsomes. However, we demonstrate significant phosphomannose isomerase activity in brain microsomes, allowing for ready interconversion between mannose 6-phosphate and glucose 6-phosphate (Vmax. 15 nmol/min per mg of microsomal protein; apparent Km < 1 mM; pH optimum 5-6 for the two-step conversion). This finding invalidates the past inference from the failure of brain microsomes to distinguish mannose 6-phosphate and glucose 6-phosphate that the cerebral glucose-6-phosphatase system lacks a 'glucose 6-phosphate translocase' [Fishman and Karnovsky (1986) J. Neurochem. 46, 371-378]. Furthermore, light-scattering experiments confirm that a proportion of whole brain microsomes is readily permeable to glucose 6-phosphate.
Department of Child Health, Medical School, University of Newcastle upon Tyne, U.K.
R J
RJ
Forsyth
K
K
Bartlett
A
A
Burchell
H M
HM
Scott
J A
JA
Eyre
D000818:Animals; D001253:Astrocytes / Q000201:enzymology*; D001692:Biological Transport; D001921:Brain / Q000378:metabolism*; D005952:Glucose-6-Phosphatase / Q000378:metabolism*; D019298:Glucose-6-Phosphate; D005956:Glucose-6-Phosphate Isomerase / Q000378:metabolism; D005958:Glucosephosphates / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008297:Male; D008359:Mannose-6-Phosphate Isomerase / Q000378:metabolism; D008861:Microsomes / Q000378:metabolism*; D009419:Nerve Tissue Proteins / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar
D005958:Glucosephosphates; D009419:Nerve Tissue Proteins; D019298:Glucose-6-Phosphate; D005952:Glucose-6-Phosphatase; D008359:Mannose-6-Phosphate Isomerase; D005956:Glucose-6-Phosphate Isomerase