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PMID: 3456607 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.

Human acid beta-glucosidase: isolation and amino acid sequence of a peptide containing the catalytic site.

Proceedings of the National Academy of Sciences of the United States of America ·Vol. 83 ·No. 6 ·1986-03-00 ·Pages 1660-4

Dinur T, Osiecki KM, Legler G, Gatt S, Desnick RJ, Grabowski GA

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 covalently 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 protease 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.

MeSH Terms
Amino Acid Sequence Base Sequence Binding Sites DNA/analysis Gaucher Disease/enzymology Glucosidases/analysis Glucosylceramidase/analysis,antagonists & inhibitors Humans Inositol/analogs & derivatives,pharmacology Lysosomes/enzymology Peptides/isolation & purification Sequence Homology, Nucleic Acid
Chemicals
Peptides Inositol DNA Glucosidases Glucosylceramidase conduritol epoxide
Authors & Affiliations
6 authors, click to expand affiliations / ORCID
Dinur T
Osiecki K M
Legler G
Gatt S
Desnick R J
Grabowski G A
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Article Info
Journal
Proceedings of the National Academy of Sciences of the United States of America
Abbr.
Proc Natl Acad Sci U S A
ISSN
0027-8424
DOI
10.1073/pnas.83.6.1660
Published
1986-03-00
Pages
1660-4
Language
English
Region
United States
NLM ID
7505876
PMCID
PMC323143
Subset
IM
Grants
NIADDK NIH HHS · K04 AM01351 · United States
External Links
PubMed source DOI: 10.1073/pnas.83.6.1660 PMC Full Text View journal details
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