Hexokinase and glucokinase binding in permeabilized guinea-pig hepatocytes.
The release of glucokinase (hexokinase IV) from digitonin-permeabilized hepatocytes from rat, guinea pig or mouse liver is inhibited by physiological concentrations of Mg2+ (> 0.25 mM). Preincubation of hepatocytes with fructose increases glucokinase release during permeabilization in the presence of Mg2+ but decreases glucokinase release in the absence of Mg2+, suggesting that fructose causes translocation of glucokinase from the Mg(2+)-dependent site. Glucose (25 mM) and sorbitol (1 mM) also induce translocation of glucokinase from the Mg(2+)-dependent site in guinea-pig, as in rat hepatocytes, but glucose is less effective than fructose or sorbitol, and the concentrations of fructose and sorbitol that cause half-maximal activation (A50) are 3-fold and 20-fold higher, respectively, in guinea-pig than in rat hepatocytes (170 microM and 257 microM, compared with 61 microM and 13 microM). Dihydroxyacetone and glycerol have no effect on fructose-induced or sorbitol-induced translocation in guinea-pig hepatocytes, in contrast with the potentiation and inhibition, respectively, by these substrates in rat hepatocytes. Some, but not all, of the differences between rat and guinea-pig hepatocytes could be due to the more reduced cytoplasmic NADH/NAD+ redox state in guinea-pig cells. The activity of low-Km hexokinases accounts for 30% of total hexokinase activity (low-Km hexokinases + glucokinase) in guinea-pig hepatocytes. Of the low-Km hexokinase activity, approx. 30% is released in the presence of Mg2+, 9% shows Mg(2+)-dependent binding and 60% shows Mg(2+)-independent binding. There was no substrate-induced translocation of low-Km hexokinase activity, indicating that translocation is specific for hexokinase IV.
Department of Medicine, University of Newcastle upon Tyne, U.K.
L
L
Agius
D000818:Animals; D001692:Biological Transport; D002463:Cell Membrane Permeability; D002478:Cells, Cultured; D004072:Digitonin / Q000494:pharmacology; D004098:Dihydroxyacetone / Q000494:pharmacology; D000431:Ethanol / Q000494:pharmacology; D005941:Glucokinase / Q000378:metabolism*; D005947:Glucose / Q000378:metabolism; D005990:Glycerol / Q000494:pharmacology; D006168:Guinea Pigs; D006593:Hexokinase / Q000378:metabolism*; D008099:Liver / Q000166:cytology / Q000187:drug effects / Q000201:enzymology*; D008274:Magnesium / Q000494:pharmacology; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D051381:Rats; D017208:Rats, Wistar; D013012:Sorbitol / Q000378:metabolism; D013379:Substrate Specificity
D000431:Ethanol; D013012:Sorbitol; D006593:Hexokinase; D005941:Glucokinase; D008274:Magnesium; D005947:Glucose; D004072:Digitonin; D004098:Dihydroxyacetone; D005990:Glycerol