Enzyme enhancement activity of N-octyl-beta-valienamine on beta-glucosidase mutants associated with Gaucher disease.

Gaucher disease (GD), caused by a defect of beta-glucosidase (beta-Glu), is the most common form of sphingolipidosis. We have previously shown that a carbohydrate mimic N-octyl-beta-valienamine (NOV), an inhibitor of beta-Glu, could increase the protein level and enzyme activity of F213I mutant beta-Glu in cultured GD fibroblasts, suggesting that NOV acted as a pharmacological chaperone to accelerate transport and maturation of this mutant enzyme. In the current study, NOV effects were evaluated in GD fibroblasts with various beta-Glu mutations and in COS cells transiently expressing recombinant mutant proteins. In addition to F213I, NOV was effective on N188S, G202R and N370S mutant forms of beta-Glu, whereas it was ineffective on G193W, D409H and L444P mutants. When expressed in COS cells, the mutant proteins as well as the wild-type protein were localized predominantly in the endoplasmic reticulum and were sensitive to Endo-H treatment. NOV did not alter this localization or Endo-H sensitivity, suggesting that it acted in the endoplasmic reticulum. Profiling of N-alkyl-beta-valienamines with various lengths of the acyl chain showed that N-dodecyl-beta-valienamine was as effective as NOV. These results suggest a potential therapeutic value of NOV and related compounds for GD with a broad range of beta-Glu mutations.

N-octyl-beta-valienamine up-regulates activity of F213I mutant beta-glucosidase in cultured cells: a potential chemical chaperone therapy for Gaucher disease.

Gaucher disease (GD) is the most common form of sphingolipidosis and is caused by a defect of beta-glucosidase (beta-Glu). A carbohydrate mimic N-octyl-beta-valienamine (NOV) is an inhibitor of beta-Glu. When applied to cultured GD fibroblasts with F213I beta-Glu mutation, NOV increased the protein level of the mutant enzyme and up-regulated cellular enzyme activity. The maximum effect of NOV was observed in F213I homozygous cells in which NOV treatment at 30 microM for 4 days caused a approximately 6-fold increase in the enzyme activity, up to approximately 80% of the activity in control cells. NOV was not effective in cells with other beta-Glu mutations, N370S, L444P, 84CG and RecNciI. Immunofluorescence and cell fractionation showed localization of the F213I mutant enzyme in the lysosomes of NOV-treated cells. Consistent with this, NOV restored clearance of 14C-labeled glucosylceramide in F213I homozygous cells. F213I mutant beta-Glu rapidly lost its activity at neutral pH in vitro and this pH-dependent loss of activity was attenuated by NOV. These results suggest that NOV works as a chemical chaperone to accelerate transport and maturation of F213I mutant beta-Glu and may suggest a therapeutic value of this compound for GD.