Human acid beta-glucosidase. Use of inhibitory and activating monoclonal antibodies to investigate the enzyme's catalytic mechanism and saposin A and C binding sites.

摘要: Abstract Of 14 identified epitopes on human GCase (acid beta-glucosidase), monoclonal antibodies (MCABs) recognizing 3 produced inhibition and 1 resulted in activation of GCase. MCABs F1 F2 completely, MCAB 61 partially (approximately 70%), inhibited activity. Substrates active site-directed inhibitors (specific sphingolipid 5-amino-5-deoxyglucose derivatives) protected the enzyme from by F2, but not that 61. Conduritol B epoxide did protect these when covalently bound to site. These results indicated highly specific binding requirements for residues a complex In comparison, kinetic analyses using transition state analogues, N-alkyl-glucosylamines, demonstrated this "freezes" conformation inhibits preventing formation conformer needed maximal catalytic rates. The activating 122 mimicked effects saposin C competed with natural activator enzyme. Interaction A or synergistic effect leading marked sensitization activators. No such synergism additivity was found rate since it could be achieved saturating amounts any one combinations presence 61, only 15 25% obtained 122, indicating major effectors derived an induction conformational change. demonstrate saposins mediate their distinct sites Furthermore, components mechanisms catalysis are due changes during state. findings have implications understanding perturbations function missense mutations which cause Gaucher disease.