Structural variations in the glycosaminoglycan-protein linkage region of recombinant decorin expressed in Chinese hamster ovary cells

Decorin is a small fibroblast proteoglycan consisting of a core protein and a single chondroitin/dermatan sulfate chain, The structure of the carbohydrate-protein linkage region of the recombinant decorin expressed in Chinese hamster ovary cells was investigated, The decorin was secreted in the culture medium and isolated by anion-exchange chromatography. The glycosaminoglycan chain was released from the decorin by beta-elimination using alkaline NaBH4, and then digested with chondroitinase ABC. These treatments resulted in a major and a few minor hexasaccharide alditols derived from the carbohydrate-protein linkage region, Their structures were analyzed by enzymatic digestion in conjunction with high-performance liquid chromatography, Two of these compounds have the conventional hexasaccharide core, Delta HexA alpha 1-3GalNAc beta 1-4GlcA beta 1-3GalNAc beta 1- -3Gal beta 1-4Xyl-ol. One is nonsulfated, and the other is monosulfated on C4 of the GalNAc residue, They represent 12% and 60% of the total linkage region, respectively, The other compound has the hexasaccharide alditol with an internal iduronic acid residue Delta HexA alpha 1-->3GalNAc(4-sulfate)beta 1-4IdoA alpha 1-3Gal beta 1-3Gal beta 1-4Xyl-ol, which was previously demonstrated in one of the five linkage hexasaccharide alditols isolated from dermatan sulfate proteoglycans of bovine aorta (Sugahara et al., J. Biol. Chem., 270, 7204-7212, 1995), The compound accounts for 11% of the total linkage region, These structural variations in the linkage hexasaccharide region of the decorin strikingly contrast to the uniformity demonstrated in the linkage hexasaccharide structure of human inter-alpha-trypsin inhibitor (Yamada ef al., Glycobiology, 5, 335-341, 1995) and urinary trypsin inhibitor (Yamada ef al., Eur, J. Biochem., 233, 687-693, 1995), both of which have a single chondroitin sulfate chain with a uniform linkage hexasaccharide structure, Delta HexA alpha 1-3GalNAc(4-sulfate)beta 1-4GlcA beta 1-3Gal(4-sulfate)beta 1-3Gal beta 1-4Xyl, containing a 4-O-sulfated Gal residue.