Oligosaccharide structure and amino acid sequence of the major glycopeptides of mature human. beta. -hexosaminidase

Journal Article · Tue Jul 12 04:00:00 EDT 1988 · Biochemistry; (United States)

DOI:https://doi.org/10.1021/bi00414a041· OSTI ID:7002496

O'Dowd, B F; Cumming, D A; Gravel, R A; Mahuran, D

Human ..beta..-hexosaminidase is a lysosomal enzyme that hydrolyzes terminal N-acetylhexosamines from GM/sub 2/ ganglioside, oligosaccharides, and other carbohydrate-containing macromolecules. There are two major forms of hexosaminidase: hexosaminidase A, with the structure ..cap alpha..(..beta../sub a/..beta../sub b/), and hexosaminidase B, 2(..beta../sub a/..beta../sub b/). Like other lysosomal proteins, hexosaminidase is targeted to its destination via glycosylation and processing in the rough endoplasmic reticulum and Golgi apparatus. Phosphorylation of specific mannose residues allows binding of the protein to the phosphomannosyl receptor and transfer to the lysosome. In order to define the structure and placement of the oligosaccharides in mature hexosaminidase and thus identify candidate mannose 6-phosphate recipient sites, the major tryptic/chymotryptic glycopeptides from each isozyme were purified by reverse-phase high-performance liquid chromatography. Two major concanavalin A binding glycopeptides, localized to the ..beta../sub b/f chain, and one non concanavalin A binding glycopeptide, localized to the ..beta../sub a/ chain, were found associated with the ..beta..-subunit in both hexosaminidase A and hexosaminidase B. The oligosaccharide structures were determined by nuclear magnetic resonance spectrometry. The unique glycopeptide associated with the ..beta../sub a/ chain contained a single GlcNAc residue. Thus all three mature polypeptides comprising the ..cap alpha.. and ..beta.. subunits of hexosaminidase contain carbohydrate, the structures of which have the appearance of being partially degraded in the lysosome. In the ..cap alpha.. chain they found only one possible site for in vivo phosphorylation. In the ..beta.. it is unclear if only one or all three of the sites could have contained phosphate. However, mature placental hexosaminidase A and B can be rephosphorylated in vitro. This requires the presence of an oligosaccharide containing an ..cap alpha..1,2-linked mannose residue.

• Research Organization: Research Institute, Toronto, Ontario (Canada)
• OSTI ID: 7002496
• Journal Information: Biochemistry; (United States), Journal Name: Biochemistry; (United States) Vol. 27:14; ISSN BICHA
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
AMINO ACID SEQUENCE
ANIMALS
BARYONS
BIOCHEMISTRY
CARBOHYDRATES
CELL CONSTITUENTS
CHEMICAL SHIFT
CHEMISTRY
CHROMATOGRAPHY
ELEMENTARY PARTICLES
ENZYMES
FERMIONS
GLUCOPROTEINS
GLYCOPROTEINS
GLYCOSYL HYDROLASES
HADRONS
HYDROLASES
LIQUID COLUMN CHROMATOGRAPHY
LYSOSOMES
MAGNETIC RESONANCE
MAMMALS
MAN
MOLECULAR STRUCTURE
NMR SPECTRA
NUCLEAR MAGNETIC RESONANCE
NUCLEONS
O-GLYCOSYL HYDROLASES
OLIGOSACCHARIDES
ORGANIC COMPOUNDS
ORGANOIDS
PRIMATES
PROTEINS
PROTONS
RESONANCE
SACCHARIDES
SEPARATION PROCESSES
SPECTRA
VERTEBRATES