Isolation and characterization of proteoglycans synthesized by human colon and colon carcinoma.
The biosynthesis of proteoglycans in short term organ culture of human colon and colon carcinoma was studied. Proteoglycans, labeled with [35S]sulfate and [3H]serine, were extracted with either 4 M or 0.5 M guanidine HCl in the presence of protease inhibitors and sequentially purified by associative and dissociative CsCl density gradient ultracentrifugation. Normal colon synthesized two polydisperse classes of proteoglycans: a large heparan sulfate-containing monomer, with a Kav of 0.48 on Sepharose CL-2B and a small dermatan sulfate-containing monomer with a Kav of 0.65. A portion (25%) of the proteoglycans was found as aggregate when chromatographed under associative conditions, and the larger monomers interacted with hyaluronic acid to an extent greater than the smaller proteoglycans. Following papain or alkali treatment, the free glycosaminoglycan side chains of both monomers eluted as a single broad peak (Kav = 0.5) from Sepharose CL-6B, with an estimated Mr of 20 X 10(3). In contrast, colon carcinoma synthesized only one proteoglycan monomer, which aggregated to a limited extent (12%). This proteoglycan population, with a Kav of 0.7 on CL-2B, contained chondroitin sulfate as the major glycosaminoglycan (greater than 81%), with small amounts of dermatan sulfate. The glycosaminoglycans had an estimated Mr of 9 X 10(3), and the disaccharides released by chondroitinase ABC consisted of 32% 4-sulfate and 68% 6-sulfate. Electron microscopy of mixed proteoglycancytochrome c monolayers from the associative fractions of normal and neoplastic colon revealed aggregated complexes which were similar in over structure, although smaller than the proteoglycan aggregates from cartilage.
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002356:Cartilage / Q000378:metabolism; D002417:Cattle; D002499:Centrifugation, Density Gradient; D003106:Colon / Q000378:metabolism*; D003110:Colonic Neoplasms / Q000378:metabolism*; D006025:Glycosaminoglycans / Q000096:biosynthesis / Q000302:isolation & purification; D006595:Hexosamines / Q000032:analysis; D006801:Humans; D008854:Microscopy, Electron; D011509:Proteoglycans / Q000096:biosynthesis* / Q000302:isolation & purification
D000596:Amino Acids; D006025:Glycosaminoglycans; D006595:Hexosamines; D011509:Proteoglycans