Analysis of glycosaminoglycan chains from different proteoglycan populations in human embryonic skin fibroblasts.
1. The structure of chondroitin/dermatan and heparan-sulphate chains from various proteoglycan populations derived from cultured human skin fibroblasts have been examined. Confluent cell cultures were biosynthetically labelled with [3H]-glucosamine and 35SO4(2-), and proteoglycans were purified according to buoyant density, size and charge density [Schmidtchen, A., Carlstedt, I., Malmström, A. & Fransson, L.-A. (1990) Biochem. J. 265, 289-300]. Some proteoglycan fractions were further fractionated according to hydrophobicity on octyl-Sepharose in Triton X-100 gradients. The glycosaminoglycan chains, intact or degraded by chemical or enzymic methods were then analysed by gel chromatography on Sepharose CL-6B, Bio-Gel P-6, ion exchange HPLC and gel electrophoresis. 2. Three types of dermatan-sulphate chains were identified on the basis of disaccharide composition and chain length. They were derived from the large proteoglycan, two small proteoglycans and a cell-associated proteoglycan with core proteins of 90 kDa and 45 kDa. Intracellular, free dermatan-sulphate chains were very similar to those of the small proteoglycans. 3. Heparan-sulphate chains from different proteoglycans had, in spite of small but distinct differences in size, strikingly similar compositional features. They contained similar amounts of D-glucuronate, L-iduronate (with or without sulphate) and N-sulphate groups. They all displayed heparin-lyase-resistant domains with average molecular mass of 10-15 kDa. The heparan-sulphate chains from proteoglycans with 250-kDa and 350-kDa cores were the largest greater than 50 kDa), containing an average of four or five domains, in contrast to heparan-sulphate chains from the small heparan-sulphate proteoglycans which had average molecular mass of 45 kDa and consisted of three or four such domains. Free, cell-associated heparan-sulphate chains were heterogeneous in size (5-45 kDa). 4. These results suggest that the core protein may have important regulatory functions with regard to dermatan-sulphate synthesis. On the other hand, synthesis of heparan sulphate may be largely controlled by the cell that expresses a particular proteoglycan core protein.
Schmidtchen
A
A
Department of Physiological Chemistry, University of Lund, Sweden.
Fransson
L A
LA
D002478:Cells, Cultured; D005591:Chemical Fractionation; D002808:Chondroitin Lyases / Q000378:metabolism; D002845:Chromatography; D003871:Dermatan Sulfate / Q000032:analysis / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D004622:Embryo, Mammalian; D005347:Fibroblasts / Q000737:chemistry*; D005965:Glucuronates / Q000032:analysis; D020723:Glucuronic Acid; D006025:Glycosaminoglycans / Q000032:analysis* / Q000737:chemistry; D019763:Heparin Lyase; D006497:Heparan Sulfate / Q000032:analysis / Q000737:chemistry; D006801:Humans; D007067:Iduronic Acid / Q000032:analysis; D008970:Molecular Weight; D011133:Polysaccharide-Lyases / Q000378:metabolism; D011509:Proteoglycans / Q000032:analysis* / Q000737:chemistry; D013431:Sulfates / Q000032:analysis
D002808:Chondroitin Lyases; D003871:Dermatan Sulfate; D005965:Glucuronates; D020723:Glucuronic Acid; D006025:Glycosaminoglycans; D019763:Heparin Lyase; D006497:Heparan Sulfate; D007067:Iduronic Acid; D011133:Polysaccharide-Lyases; D011509:Proteoglycans; D013431:Sulfates