Isolation and partial characterization of proteoglycans from sheep lung parenchyma.
Greater than 90% of the proteoglycans of sheep lung parenchyma, as measured by uronic acid, were solubilized employing a sequential procedure with guanidine hydrochloride, dithiothreitol and Triton X-100. The amounts solubilized were 68.7%, 16.2% and 5.9%, respectively. The guanidine hydrochloride extract was chromatographed using DEAE-cellulose in urea and eluted with increasing concentrations of NaCl. A major fraction (containing a 6.5-fold enrichment of uronic acid) was obtained with 0.5 M NaCl and further purified by Sepharose Cl-6B chromatography in guanidine hydrochloride. To demonstrate the presence of protein-linked glycosaminoglycans, the void volume peak containing protein and uronic acid was digested with papain and rechromatographed. Evidence for the presence of proteoglycans was obtained by observing an almost complete loss of uronic acid in the void volume and the appearance of a uronic acid peak in the included volume, migrating in the same area as single-chain glycosaminoglycans. Electrophoretic migration and disappearance of bands in electrophoresis after digestion with specific mucopolysaccharide lyases indicated that the small amount of uronic acid remaining in the void volume was hyaluronic acid whereas the included volume contained hyaluronic acid, heparan sulfate, chondroitin sulfates and/or dermatan sulfate.
D000818:Animals; D002847:Chromatography, Agarose; D002848:Chromatography, DEAE-Cellulose; D004229:Dithiothreitol; D004588:Electrophoresis, Cellulose Acetate; D006025:Glycosaminoglycans / Q000032:analysis; D006146:Guanidines; D008168:Lung / Q000032:analysis*; D008297:Male; D011092:Polyethylene Glycols; D011509:Proteoglycans / Q000302:isolation & purification*; D012756:Sheep; D014574:Uronic Acids / Q000032:analysis
D006025:Glycosaminoglycans; D006146:Guanidines; D011509:Proteoglycans; D014574:Uronic Acids; D011092:Polyethylene Glycols; D004229:Dithiothreitol