Elucidating How OMJ‐4 Restores Mutant Alpha‐Mannosidase Function Using Molecular Simulation Approaches

ABSTRACT To alleviate the implications caused by mutations in the alpha‐mannosidase (AFM) enzyme, a pharmacological chaperone therapy, namely N ‐[9‐( tert ‐butoxycarbonyl)amino]nonyl derivative 4 (OMJ‐4), was previously proposed to demonstrate significant therapeutic benefits. Although OMJ‐4 interacts with mutant alpha‐mannosidase, its precise restorative mechanism remains unclear. In this study, we investigated the site and mechanism of action of OMJ‐4 through in silico analyses. Binding analysis revealed a delayed dissociation of OMJ‐4 from the non‐competitive site, indicating a stronger interaction at that location. Moreover, molecular dynamics (MD) analysis revealed that binding of OMJ‐4 induced significant changes in the compactness and active‐site flexibility of the mutant enzyme, enabling better accommodation of the substrate molecule. Furthermore, the restorative effect of OMJ‐4 on mutant alpha‐mannosidase was studied using molecular orbital analysis and enzyme reaction pathway analysis based on semi‐empirical quantum mechanics. The molecular orbital assessment of mannose disaccharide dissociated from OMJ‐4‐treated mutant alpha‐mannosidase showed high reactivity with an energy gap of 4 eV. The enzyme reaction pathway analysis of OMJ‐4‐treated mutant alpha‐mannosidase exhibited an activation energy of 92 kcal/mol, notably lower than the 98.5 kcal/mol observed for the native enzyme. These findings demonstrate the therapeutic effect of the OMJ‐4 chaperone, emphasizing its ameliorative action at the non‐competitive site of the dysfunctional alpha‐mannosidase.