Identification of the core proteins in proteoglycans synthesized by vascular endothelial cells.
Proteoglycans, metabolically labelled with [3H]leucine and 35SO4(2-), were isolated from the spent media and from guanidinium chloride extracts of cultured human umbilical-vein endothelial cells by using isopycnic density-gradient centrifugation, gel filtration and ion-exchange h.p.l.c. The major proteoglycan species were subjected to SDS/polyacrylamide-gel electrophoresis before and after enzymic degradation of the polysaccharide chains. The cell extract contained mainly a heparan sulphate proteoglycan that has a buoyant density of 1.31 g/ml and a protein core with apparent molecular mass 300 kDa. The latter was heterogeneous and migrated as one major and one minor band. After reduction, the apparent molecular mass of the major band increased to approx. 350 kDa, indicating the presence of intrachain disulphide bonds. The proteoglycan binds to octyl-Sepharose and its polysaccharide chains are extensively degraded by heparan sulphate lyase. The proteoglycans of the medium contained 90% of all the incorporated 35SO4(2-). Here the predominant heparan sulphate proteoglycan was similar to that of the cell extract, but was more heterogeneous and contained an additional core protein with apparent molecular mass 210 kDa. Furthermore, two different chondroitin sulphate proteoglycans were found: one 200 kDa species with a high buoyant density (approx. 1.45 g/ml) and one 100 kDa species with low buoyant density (approx. 1.3 g/ml). Both these proteoglycans have a core protein of molecular mass approx. 47 kDa.
Department of Physiological Chemistry 2, University of Lund, Sweden.
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Lindblom
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Carlstedt
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Fransson
D053674:Aggrecans; D002478:Cells, Cultured; D002500:Centrifugation, Isopycnic; D011508:Chondroitin Sulfate Proteoglycans / Q000302:isolation & purification; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D004730:Endothelium, Vascular / Q000032:analysis*; D016326:Extracellular Matrix Proteins; D006023:Glycoproteins / Q000302:isolation & purification*; D019812:Heparan Sulfate Proteoglycans; D006497:Heparan Sulfate / Q000302:isolation & purification; D006801:Humans; D037181:Lectins, C-Type; D008970:Molecular Weight; D011509:Proteoglycans
D053674:Aggrecans; D011508:Chondroitin Sulfate Proteoglycans; D016326:Extracellular Matrix Proteins; D006023:Glycoproteins; D019812:Heparan Sulfate Proteoglycans; D006497:Heparan Sulfate; D037181:Lectins, C-Type; D011509:Proteoglycans