N‑Glycosylation and Gaucher Disease Mutation Allosterically Alter Active-Site Dynamics of Acid-β-Glucosidase

Allosteric regulations of the catalytic activity of enzymes in cellular processes remain poorly understood. Here, we advance the understanding of these critical processes by providing insights into the allosteric mechanism of acid-β-glucosidase, an enzyme associated with Gaucher disease. Glycosylation, a post-translational modification of proteins, is essential for in vivo catalytic activity and stability of acid-β-glucosidase. Asparagine 19, a glycosylation site that is located ∼30 Å from the active site, was observed to be glycosylated with either one or two N-acetyl-glucosamines by X-ray crystallography. In addition, N370S and L444P mutations, located ∼18 and 30 Å, respectively, away from the active site, cause type I Gaucher disease that is common among certain demographic groups. Using multiple microsecond-long molecular dynamics simulations, we evaluated the dynamics of the wild-type, glycosylated with one and two N-acetyl-glucosamines at N19 and the N370S and L444P variants of acid-β-glucosidase. Our results suggest that access to the substrate-binding pocket is controlled by the dynamics of loops surrounding the active site and allosterically regulated. Catalytic residues, E235 and E340, are less localized and sample more conformational states upon glycosylation of acid-β-glucosidase when compared to the wild-type and N370S variant. Our findings could help explain the allosteric effects of glycosylation on enzyme mechanisms, other disease-causing mutations in acid-β-glucosidase, and facilitate the design of drugs for the treatment of Gaucher disease.