Isolation and characterization of a link protein from bovine aorta proteoglycan aggregate.
Proteoglycan aggregates were isolated from bovine aorta by extraction with 0.5 M guanidine hydrochloride in the presence of proteinase inhibitors and purified by isopycnic CsCl centrifugation. The bottom two-fifths (A1) of the gradient contained 30% of proteoglycans in the aggregated form. The aggregate had 14.8% protein and 20.4% hexuronic acid with hyaluronic acid, dermatan sulfate and chondroitin sulfates in a proportion of 18:18:69. A link protein-containing fraction was isolated from the bottom two-fifths by dissociative CsCl isopycnic centrifugation. The link protein that floated to the top one-fifth of the gradient was purified by chromatography on Sephadex G-200 in the presence of 4 M guanidine hydrochloride. It moved as a single band in SDS-polyacrylamide gel electrophoresis with a molecular weight of 49 000. The amino acid composition of link protein resembled that of link protein from cartilage, but was strikingly different from that of the protein core of the proteoglycan monomer. The neutral sugar content of link protein was 3.5% of dry weight. Galactose, mannose and fucose constituted 21, 62 and 16%, respectively of the total neutral sugars. In aggregation studies the link protein was found to interact with both proteoglycan monomer and hyaluronic acid. Oligosaccharides derived from hyaluronic acid decreased the viscosity of link protein-free aggregates of proteoglycan and hyaluronic acid but not of link-stabilized aggregates, demonstrating that the link protein increases the stability of proteoglycan aggregates.
D000818:Animals; D001011:Aorta / Q000032:analysis*; D002417:Cattle; D016326:Extracellular Matrix Proteins; D006820:Hyaluronic Acid; D011487:Protein Conformation; D011506:Proteins / Q000302:isolation & purification*; D011509:Proteoglycans / Q000302:isolation & purification*; D012997:Solvents; D014783:Viscosity; D000099249:Proteoglycan Link Protein
D016326:Extracellular Matrix Proteins; D006820:Hyaluronic Acid; D011506:Proteins; D011509:Proteoglycans; D012997:Solvents; D000099249:Proteoglycan Link Protein