Structural studies on sulfated oligosaccharides derived from the carbohydrate-protein linkage region of chondroitin sulfate proteoglycans of whale cartilage

From the carbohydrate-protein linkage region of whale cartilage proteoglycans, which bear predominantly chondroitin 4-sulfate, one nonsulfated, two monosulfated and one disulfated hexasaccharide alditols were isolated after exhaustive digestions with Actinase E and chondroitinase ABC, and subsequent β-elimination. Their structures were analyzed by chondroitinase ACII digestion in conjunction with HPLC and by 500-MHz 1H-NMR spectroscopy. The nonsulfated compound (A) had the following conventional structure: ΔGlcA(β1 - 3)-GalNAc(β1 - 4)GlcA(β1 - 3)Gal(β1 - 3)Gal(β1 - 4)Xylol, where GlcA, ΔGlcA and GalNAc are glucuronic acid; 4,5-unsaturated glucuronic acid and 2-deoxy-2-N-acetylamino-D-galactose, respectively. The other compounds were sulfated derivatives of compound A. Two monosulfated compounds (B and C) had an ester sulfate on C4 or C6 of the GalNAc residue, respectively and the disulfated compound (D) had two ester sulfate groups, namely, one on C4 of the GalNAc and the other on C4 of the Gal residue substituted by GlcA. The molar ratio of A/B/C/D was 0.21:0.16:0.36:0.27. The compound containing Gal-4-O-sulfate was previously isolated by us in the form of a sulfated glycoserine [Δ GlcA(β1 - 3)GalNAc(4-O-sulfate)(β1 - 4)GlcA(β1 - 3)Gal(4-O-sulfate)(β1 - 3)-Gal(β1 - 4)Xylβ1-O-Ser] from the carbohydrate-protein linkage region of rat chondrosarcoma chondroitin-4-sulfate proteoglycans [Sugahara K., Yamashina, I., DeWaard P., Van Halbeek, H. and Viegenthart J.F.G. (1988) J. Biol. Chem. 263, 10168 - 10174]. The discovery of this structure in the carbohydrate-protein linkage region of chondroitin 4-sulfate proteoglycans from nontumorous cartilage indicates that it is not a tumor-associated product but rather a physiological biosynthetic product since it represents a significant proportion. The biological significance of this structure is discussed in relation to glycosaminoglycan biosynthesis.