Purification and characterization of a hyaluronan-binding protein from rat chondrosarcoma.
Swarm rat chondrosarcoma contains a hyaluronan-binding protein of molecular mass 102 kDa (HABP102). The protein is present in 4 M-guanidinium chloride extracts of the chondrosarcoma and can be incorporated into reconstituted proteoglycan aggregates, but it is not present in native proteoglycan aggregates or in 0.5 M-guanidinium chloride extracts. HABP102 is unlikely to be an integral membrane protein, as it does not require detergent for extraction, is not enriched in hydrophobic amino acids and does not bind avidly to octyl-Sepharose. The protein stains poorly with Coomassie Blue and is only visible on PAGE gels after staining with silver. Disulphide bonds are essential for the binding of HABP102 to hyaluronan, and bivalent cations are not required for this interaction. HABP102 can be purified from dissociative chondrosarcoma extracts by sequential density-gradient centrifugation, hyaluronan-Sepharose affinity chromatography and hydrophobic-interaction chromatography. The amino acid composition is similar to that of domains 1-4 of the chondrosarcoma proteoglycan core protein, but peptide analysis after digestion with Staphylococcus aureus V8 proteinase and chymotrypsin and different immunoreactivity suggest that HABP102 is not closely related to proteoglycan hyaluronan-binding region. HABP102 is a glycoprotein containing N-acetylgalactosamine, N-acetylglucosamine, mannose and galactose.
Crossman
M V
MV
Department of Biochemistry, Charing Cross and Westminster Medical School (University of London), U.K.
Mason
R M
RM
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D001667:Binding, Competitive; D002813:Chondrosarcoma / Q000032:analysis*; D002846:Chromatography, Affinity; D006023:Glycoproteins / Q000302:isolation & purification* / Q000378:metabolism; D006820:Hyaluronic Acid / Q000378:metabolism*; D008970:Molecular Weight; D009844:Oligosaccharides / Q000378:metabolism; D010449:Peptide Mapping; D011485:Protein Binding; D051381:Rats; D011919:Rats, Inbred Strains
D000596:Amino Acids; D006023:Glycoproteins; D009844:Oligosaccharides; D006820:Hyaluronic Acid