The iminosugar isofagomine increases the activity of N370S mutant acid β-glucosidase in Gaucher fibroblasts by several mechanisms

TLDR

Gaucher disease results from deficient lysosomal acid β‑glucosidase, and the common N370S mutation reduces enzyme activity and impairs its exit from the endoplasmic reticulum. The study aimed to determine whether the iminosugar isofagomine (IFG) could enhance activity of the N370S mutant enzyme in fibroblasts. IFG promotes proper folding and ER export of newly synthesized GlcCerase, increasing its lysosomal pool. Treatment with IFG raised mutant GlcCerase activity by ~3‑fold, shifted its pH optimum from 6.4 to 5.2, altered SDS sensitivity, and allowed activity to recover within 24 h after washout, supporting active‑site inhibitors as a therapeutic strategy.

Abstract

Gaucher disease is a lysosomal storage disorder caused by deficiency in lysosomal acid beta-glucosidase (GlcCerase), the enzyme responsible for the catabolism of glucosylceramide. One of the most prevalent disease-causing mutations, N370S, results in an enzyme with lower catalytic activity and impaired exit from the endoplasmic reticulum. Here, we report that the iminosugar isofagomine (IFG), an active-site inhibitor, increases GlcCerase activity 3.0 +/- 0.6-fold in N370S fibroblasts by several mechanisms. A major effect of IFG is to facilitate the folding and transport of newly synthesized GlcCerase in the endoplasmic reticulum, thereby increasing the lysosomal pool of the enzyme. In addition, N370S GlcCerase synthesized in the presence of IFG exhibits a shift in pH optimum from 6.4 to 5.2 and altered sensitivity to SDS. Although IFG fully inhibits GlcCerase in the lysosome in an in situ assay, washout of the drug leads to partial recovery of GlcCerase activity within 4 h and full recovery by 24 h. These findings provide support for the possible use of active-site inhibitors in the treatment of some forms of Gaucher disease.