The molecular basis of glycogen storage disease type 1a

Glycogen storage disease type la is caused by a deficiency in glucose-6-phosphatase (G6Pase), a nine-helical endoplasmic reticulum transmembrane protein required for maintenance of glucose homeostasis. To date, 75 G6Pase mutations have been identified, including 48 mutations resulting in single-amino acid substitutions. However, only 19 missense mutations have been functionally characterized. Here, we report the results of structure and function studies of the 48 missense mutations and the DeltaF327 codon deletion mutation, grouped as active site, helical, and nonhelical mutations. The 5 active site mutations and 22 of the 31 helical mutations completely abolished G6Pase activity, but only 5 of the 13 nonhelical mutants were devoid of activity. Whereas the active site and nonhelical mutants supported the synthesis of G6Pase protein in a manner similar to that of the wild-type enzyme, immunoblot analysis showed that the majority (64.5%) of helical mutations destabilized G6Pase. Furthermore, we show that degradation of both wild-type and mutant G6Pase is inhibited by lactacystin, a potent proteasome inhibitor. Taken together, we have generated a data base of residual G6Pase activity retained by G6Pase mutants, established the critical roles of transmembrane helices in the stability and activity of this phosphatase, and shown that G6Pase is a substrate for proteasome-mediated degradation.