The insulin sensitive hexose transport system of the isolated rat adipocyte has been studied. 3-O-methyl-D-glucose, D-allose and D-xylose are all transported by the hexose transport system. By studying the inhibition of D-allose transport by a range of hexose analogues the hydrogen bonding and spatial requirements of the transporter have been investigated. The transporter accepts the hexose molecule in a pyranose ring form. The results indicate that the important hydrogen bonding positions are the ring oxygen, C-1 and C-3. There may be a weaker hydrogen bond to C-6. Sugar hydroxyls in non-glucose configurations may sterically hinder transport. The inhibition by alkyl substituted hexoses indicates that the hexose transporter requires a specific orientation of the hexose molecule as it approaches the external binding site. C-1 faces the transporter whilst C-4 faces the external solution. When the hexose molecule is bound to the hexose transporter there is little space around C-1 and C-2. There is more space around C-3, C-4 and C-6 with the possibility of a hydrophobic region adjacent to epsilon-6. 4,6-O-ethylidene-D-glucose and alkyl-beta-D-glucosides enter the adipocyte independently of the hexose transport system. The results indicate that 4,6-O-ethylidene-D-glucose is a good side specific analogue with a high affinity for the external site. In contrast alkyl-beta-D-glucosides are good side specific inhibitors on the inside of the hexose transport system. Both 4,6-O-ethylidene-D-glucose and n'-propyl-beta-D-glucoside are competitive inhibitors of the hexose transport system. Treatment of basal adipocytes with 10nM insulin can lead to a 50-fold stimulation of hexose transport. The hydrogen bonding and spatial requirements of the hexose transporter are unchanged by insulin. D-glucose and 2-deoxy-D-glucose showed reduced inhibition of D-allose transport when the intracellular ATP levels were depleted by cyanide poisoning. There was no effect of cyanide on the inhibition of transport by 3-O-methyl-D-glucose. This result suggests a possible effect of hexose metabolites on the hexose transporter. A range of purine and pyrimidine nucleosides, nucleotides and cyclic nucleotides were tested for possible effects on hexose transport when added exogenously to adipocyte suspensions. Adenosine (10muM) and inosine (0.1muM) were found to give small stimulations of hexose transport in basal adipocytes. Xanthine stimulated hexose transport in basal adipocytes and inhibited transport in insulin-stimulated cells. Exogenous ATP inhibited hexose transport in insulin-stimulated cells. Cyclic nucleotides did not affect the rate of hexose transport in adipocytes.
|Date of Award||1981|