Human acid beta-glucosidase: isolation and amino acid sequence of a peptide containing the catalytic site.
- 1 March 1986
- journal article
- research article
- Published by Proceedings of the National Academy of Sciences in Proceedings of the National Academy of Sciences
- Vol. 83 (6) , 1660-1664
- https://doi.org/10.1073/pnas.83.6.1660
Abstract
Human acid .beta.-glucosidase (D-glucosyl-N-acylsphingosine glucohydrolase, EC 3.2.1.45) cleaves the glucosidic bonds of glucosylceramide and synthetic .beta.-glucosides. The deficient activity of this hydrolase is the enzymatic defect in the subtypes and variants of Gaucher disease, the most prevalent lysosomal storage disease. To isolate and characterize the catalytic site of the normal enzyme, brominated 3H-labeled conduritol B epoxide (3H-Br-CBE), which inhibits the enzyme by binding convalently to this site, was used as an affinity label. Under optimal conditions 1 mol of 3H-Br-CBE bound to 1 mol of pure enzyme protein, indicating the presence of a single catalytic site per enzyme subunit. After V8 proteases digestion of the 3H-Br-CBE-labeled homogeneous enzyme, three radiolabeled peptides, designated peptide A, B, or C, were resolved by reverse-phase HPLC. The partial amino acid sequence (37 residues) of peptide A (Mr, 5000) was determined. The sequence of this peptide, which contained the catalytic site, had exact homology to the sequence near the carboxyl terminus of the protein, as predicted from the nucleotide sequence of the full-length cDNA encoding acid .beta.-glucosidase.This publication has 30 references indexed in Scilit:
- Genetic heterogeneity in Gaucher disease: Physicokinetic and immunologic studies of the residual enzyme in cultured fibroblasts from non‐neuronopathic and neuronopathic patientsAmerican Journal of Medical Genetics, 1985
- A BASIC microcomputer program for plotting the secondary structure of proteinsComputer Programs in Biomedicine, 1982
- A simple method for displaying the hydropathic character of a proteinJournal of Molecular Biology, 1982
- Mechanism of activation of glucocerebrosidase by CO-β-glucosidase (glucosidase activator protein)Biochimica et Biophysica Acta (BBA) - Lipids and Lipid Metabolism, 1981
- Isolation and structure of a tryptic glycopeptide from the active site of β-glucosidase A3 from Aspergillus wentiiBiochimica et Biophysica Acta (BBA) - Protein Structure, 1980
- Amino acid sequence at the active site of β-glucosidase a from bitter almondsBiochimica et Biophysica Acta (BBA) - Enzymology, 1978
- Solubilization of glucocerebrosidase from human placenta and demonstration of a phospholipid requirement for its catalytic activityBiochemical and Biophysical Research Communications, 1976
- Isolation and Amino Acid Sequence of a Hexadecapeptide from the Active Site of ß-Glucosidase A3 from Aspergillus wentiiBiological Chemistry, 1974
- Cleavage of Structural Proteins during the Assembly of the Head of Bacteriophage T4Nature, 1970
- Isolation of β-galactosidase and β-glucosidase from brainBiochimica et Biophysica Acta (BBA) - Enzymology and Biological Oxidation, 1966