Deglycosylation of chondroitin sulfate proteoglycan and derived peptides

Journal Article · Mon Jan 29 23:00:00 EST 1990 · Biochemistry; (USA)

DOI:https://doi.org/10.1021/bi00456a009· OSTI ID:6719672

Campbell, S C; Krueger, R C; Schwartz, N B ^[1]

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1. Univ. of Chicago, IL (USA)

In order to define the domain structure of proteoglycans as well as identify primary amino acid sequences specific for attachment of the various carbohydrate substituents, reliable techniques for deglycosylating proteoglycans are required. In this study, deglycosylation of cartilage chondroitin sulfate proteoglycan (CSPG) with minimal core protein cleavage was accomplished by digestion with chondroitinase ABC and keratanase, followed by treatment with anhydrous HF in pyridine. Nearly complete deglycosylation of secreted proteoglycan was verified within 45 min of HF treatment by loss of incorporated ({sup 3}H)glucosamine label from the proteoglycan as a function of time of treatment, as well as by direct analysis of carbohydrate content and xylosyltransferase acceptor activity of unlabeled core protein preparations. The deglycosylated CSPG preparations were homogeneous and of high molecular weight. Comparison of the intact deglycosylated core protein preparations with newly synthesized unprocessed precursors suggested that extensive proteolytic cleavage of the core protein did not occur during normal intracellular processing. Furthermore, peptide patterns generated after clostripain digestion of core protein precursor and of deglycosylated secreted proteoglycan were comparable. With the use of the clostripain digestion procedure, peptides were produced from unlabeled proteoglycan, and two predominant peptides from the most highly glycosylated regions were isolated, characterized, and deglycosylated. These peptides were found to follow similar kinetics of deglycosylation and to acquire xylose activity comparable to the intact core protein.

• OSTI ID: 6719672
• Journal Information: Biochemistry; (USA), Journal Name: Biochemistry; (USA) Vol. 29:4; ISSN 0006-2960; ISSN BICHA
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
AMINES
AMINO ACID SEQUENCE
AMINO ACIDS
ANIMAL CELLS
ANIMAL TISSUES
ANIMALS
BETA DECAY RADIOISOTOPES
BETA-MINUS DECAY RADIOISOTOPES
BIODEGRADATION
BIRDS
BODY
CARBOHYDRATES
CARBOXYLIC ACIDS
CARTILAGE
CELL CULTURES
CHEMICAL REACTIONS
CHICKENS
CHONDROITIN
CONNECTIVE TISSUE
DAYS LIVING RADIOISOTOPES
DECOMPOSITION
DISEASES
DRUGS
EVEN-ODD NUCLEI
FOWL
GLUCOSAMINE
GLYCOPROTEINS
HEXOSAMINES
HEXOSES
HYDROGEN COMPOUNDS
HYDROXY ACIDS
ISOTOPES
LIGHT NUCLEI
LIPOTROPIC FACTORS
MAMMALS
METHIONINE
MOLECULAR STRUCTURE
MOLECULAR WEIGHT
MONOSACCHARIDES
MUCOPOLYSACCHARIDES
NEOPLASMS
NUCLEI
ORGANIC ACIDS
ORGANIC COMPOUNDS
ORGANIC SULFUR COMPOUNDS
PEPTIDES
POLYSACCHARIDES
PROTEINS
RADIOISOTOPES
RATS
RODENTS
SACCHARIDES
SARCOMAS
SECRETION
SERINE
SULFUR 35
SULFUR ISOTOPES
TISSUES
TRITIUM COMPOUNDS
TUMOR CELLS
VERTEBRATES