D-Glucose Uptake by Isolated Human Erythrocyte Membranes versus D-Glucose Transport by Human Erythrocytes COMPARISON OF THE EFFECTS OF PROTEOLYTIC AND PHOSPHOLIPASE A, DIGESTION*

The effects of proteolytic and phospholipase AZ digestion on the transport of D-glucose by intact human erythrocytes and on the uptake of n-glucose by isolated erythrocyte membranes have been studied. This stereospecific uptake of D-glucose was previously shown to be compatible with the over-all monosaccharide transport system in erythrocytes and evidence was presented that it represented the binding of D-glucose to a specific site on the erythrocyte membrane (KAHLENBERG, A., URMAN, B., AND DOLANSKY, D. (1971) Biochemistry 10,3154-3162). In the absence of hemolysis, no change is observed in the transport of D-glucose by intact erythrocytes exposed to the above enzymes or in the uptake of D-glucose by membranes isolated from enzyme-treated cells. In contrast, limited proteolytic digestion of isolated erythrocyte membranes, under conditions identical with those for the digestion of intact cells, results in a 3to 5-fold increase in n-glucose uptake and a 2-fold increase in the apparent dissociation constant for the D-glucose-membrane complex. This increase in the uptake of D-glucose is different from the effects which result from the digestion of isolated membranes with phospholipase Az (decreased uptake of D-glucose) or neuraminidase (no change in D-glucose uptake). More drastic tryptic digestion leads to a loss of the stimulated D-glucose uptake with no further change in the affinity of the membrane for the sugar. Pronase hydrolysis of 20 to 30% of the proteins of isolated membranes results in the destruction of 50 to 70% of the D-glucose uptake. These results indicate that the transport system in erythrocytes involves membrane components not normally accessible to proteolytic and phospholipase AZ digestion. Isolation of the erythrocyte membrane renders the D-glucose uptake activity of these membrane components susceptible to enzymatic modification. The evidence suggests that the