Imino- and azasugar protonation inside human acid Beta-glucosidase, the enzyme defective in Gaucher disease

Gaucher disease is caused by mutations in human acid β‐glucosidase, the enzyme responsible for hydrolysis of glucosyl ceramide in the lysosomes. Imino‐ and azasugars such as 1‐deoxynojirimycin and isofagomine are strong inhibitors of the enzyme and of interest in pharmacological chaperone therapy of the disease. Despite several crystal structures of the enzyme with the imino‐ and azasugars bound in the active site having been resolved, the actual acid‐base chemistry of the binding is not known. In this study we show, using photoinduced electron transfer (PET), that 1‐deoxynojirimycin and isofagomine derivatives are protonated by human acid β‐glucosidase when bound even if completely unprotonated outside the enzyme. While isofagomine derivative protonation to some degree was foreshadowed by earlier crystal structures, 1‐deoxynojirimycin derivatives were not believed to act as basic amines in the enzyme.