Decreased Ca²⁺-ATPase Activity After Glycosylation of Erythrocyte Membranes In Vivo and In Vitro

Erythrocyte membranes drawn from diabetic patients with poor metabolic control have increased protein glycosylation and decreased Ca²⁺-ATPase activity. A significant relationship was found between these two parameters. Similar results were obtained when protein glycosylation and Ca²⁺-ATPase activity were measured in membranes from normal erythrocytes preincubated with glucose. In this condition, both parameters showed a clear time and dose dependence. Incubation of erythrocyte membranes instead of intact erythrocytes with glucose and glucose-6-phosphate strongly suggests that only the glycosylation of the membrane inner-surface proteins can affect Ca²⁺-ATPase activity. The simultaneous presence of 10 mM glucose and 5 mM ATP in the incubation medium did not affect the degree of erythrocyte membrane protein glycosylation but significantly blocked the inhibitory effect of glucose on Ca²⁺-ATPase activity. However, 5 mM ATP only partially blocked the inhibitory effect of 100 mM glucose, suggesting a competitive mechanism of glucose and ATP for the enzyme active site. Our results show that glycosylation of erythrocyte membrane proteins significantly inhibits Ca²⁺-ATPase activity. This effect could contribute to the development of the capillary closure process observed in diabetic patients. Furthermore, it could represent an index of a general impairment of enzyme function arising in cells chronically exposed to high glucose levels.