Structure/function relationships of acid beta-glucosidase, the enzyme deficient 
in Gaucher disease, were evaluated by characterizing the proteins expressed from 
cDNAs encoding normal and mutant enzymes. Twenty-two Gaucher disease mutations 
or created mutations were expressed in Spodoptera frugiperda (Sf9) cells and 
analyzed for catalytic properties, stability, inhibitor binding, and modifier 
interactions. Many Gaucher disease mutations encoded highly disruptive amino 
acid substitutions (e.g. P289L and D409V) and produced severely compromised 
proteins with very reduced activity (kcat < 1% of normal) and/or stability. Six 
mutant enzymes had sufficient catalytic activity (kcat approximately 5-30% of 
normal) for extensive studies. The highly conservative substitutions, i.e. F216Y 
or S364T and V394L, led to severe, but selective, abnormalities of enzyme 
stability or large decreases in catalytic activity, respectively. The T323I, 
N370S, and V394L enzymes interacted abnormally with active site-directed 
inhibitors and localized these residues to the glycon binding region. Selected 
mutant enzymes were poorly activated by phosphatidylserine (V394L, L444P, and 
R463C) or by saposin C (L444P and T323I), indicating that the enzyme sites for 
interaction with these activators were within the carboxyl one-third of the 
enzyme. Substitutions of Ser, Glu, and/or Gly at residues Asp-443 and/or Asp-445 
demonstrated important steric roles for these residues in the active site, but 
neither is the catalytic nucleophile. Together with previous studies, the 
present analyses provide an insight into the pathogenesis of Gaucher disease and 
the functional organization of acid beta-glucosidase.