Sucrose gradient analysis of phospholipid-activated beta-glucosidase in type 1 and type 2 Gaucher's disease.
Using sucrose density gradients, differences in delipidated lysosomal beta-glucosidase isolated from control spleen and spleen from patients with nonneurologic (type 1) and neurologic (type 2) Gaucher's disease have been examined. The three enzymes differ in sedimentation properties as well as in their responsiveness to activation by phosphatidylserine and heat-stable factor. The control beta-glucosidase sedimented as an apparent 45,000-Da species whose activity was dependent upon the inclusion of exogenous sodium taurodeoxycholate in the assay medium. Preincubation with a mixture of phosphatidylserine and heat-stable factor converted the control enzyme to a faster-sedimenting form which exhibited considerable activity in the absence of exogenous bile salt. Spleen beta-glucosidase from a patient with type 1 Gaucher's disease exhibited an apparent molecular weight of 154,000 on sucrose gradients. Like the control enzyme, the activity of this form was bile salt dependent. Upon preincubation with phosphatidylserine and heat-stable factor, beta-glucosidase from the type 1 case was also converted to a faster-sedimenting form which was more active in the absence of sodium taurodeoxycholate than in the presence of the bile salt. Spleen beta-glucosidase from the patient with type 2 Gaucher's disease sedimented as a broad peak of activity in the most dense regions of the sucrose gradients, appearing to be much larger than the beta-glucosidase from either the control or the type 1 Gaucher's disease patient. The activity of this large species was strongly dependent upon bile salt, and was not affected by preincubation of the enzyme with phosphatidylserine and heat-stable factor. Using the chaotropic salt, sodium thiocyanate (0.15 M), the spleen beta-glucosidase isolated from the type 1 Gaucher's disease case was converted to a slower-sedimenting species. The control enzyme sedimented slightly farther into the sucrose gradients upon treatment with the NaSCN. Thiocyanate treatment had no effect on the spleen beta-glucosidase isolated from the case of type 2 Gaucher's disease.
D000368:Aged; D002499:Centrifugation, Density Gradient; D004789:Enzyme Activation / Q000187:drug effects; D005260:Female; D005776:Gaucher Disease / Q000145:classification / Q000201:enzymology*; D005959:Glucosidases / Q000378:metabolism*; D005962:Glucosylceramidase / Q000378:metabolism; D006801:Humans; D007223:Infant; D008297:Male; D010743:Phospholipids / Q000494:pharmacology*; D013154:Spleen / Q000201:enzymology; D013861:Thiocyanates / Q000494:pharmacology; D001617:beta-Glucosidase / Q000378:metabolism*
D010743:Phospholipids; D013861:Thiocyanates; C024553:sodium thiocyanate; D005959:Glucosidases; D001617:beta-Glucosidase; D005962:Glucosylceramidase