Structural studies on the chondroitinase ABC-resistant sulfated tetrasaccharides isolated from various chondroitin sulfate isomers

• Published in: Carbohydrate research 1994-03, Vol.255, p.145-163
• Main Authors: Sugahara, Kazuyuki, Shigeno, Kaori, Masuda, Masao, Fujii, Nobutaka, Kurosaka, Akira, Takeda, Kyoto
• Format: Article
• Language: English
• Subjects: Carbohydrate Sequence; Chondroitin Lyases - metabolism; Chondroitin Sulfates - chemistry; Chondroitin Sulfates - metabolism; Chromatography, High Pressure Liquid; Disaccharides - chemistry; Isomerism; Magnetic Resonance Spectroscopy; Molecular Sequence Data; Oligosaccharides - chemistry; Oligosaccharides - metabolism; Substrate Specificity; Sulfatases - metabolism; Sulfuric Acid Esters - chemistry; Sulfuric Acid Esters - metabolism
• ISSN: 0008-6215; 1873-426X
• DOI: 10.1016/S0008-6215(00)90976-5

Various commercially available chondroitin sulfates, including an A isomer from whale cartilage, C and D isomers from shark cartilage, and an E isomer from squid cartilage, were exhaustively digested with a commercial highly purified Proteus vulgaris chondroitinase ABC. Gel chromatography of all digests yielded a disaccharide and an oligosaccharide fraction which was resistant to the enzyme digestion and which accounts for 20–31 mol% of the produced total oligosaccharides. Variably sulfated tetrasaccharides were isolated from the oligosaccharide fraction of each chondroitin sulfate isomer by HPLC, then characterized chemically and enzymatically. One disulfated and three trisulfated components were also characterized by 500-MHz one- and two-dimensional 1H NMR spectroscopy. The structures of one tetrasulfated, four trisulfated, and five disulfated tetrasaccharides with the common core structure, α- l-Δ 4,5Hex pA-(1 → 3)-β- d-Gal pNAc(1 → 4)-β- d-Glc pA-(1 → 3)- d-Gal pNAc, were determined. All isolated tetrasaccharides were resistant to the highly purified enzyme, but susceptible to the conventional, commercial chondroitinase ABC. The former was also inactive towards α- l-Δ 4,5Hex pA-(1 → 3)-β- d-Gal pNAc(1 → 4)-β- d-Glc pA-(1 → 3)- d-Gal pNAc isolated from chondroitin, β- d-Glc pA-(1 → 3)-β- d-Glc pNAc-(1 → 4)-β- d-Glc pA-(1 → 3)-β- d-Glc pNAc from hyaluronan, and α- l-Δ 4,5Hex pA-(1 → 3)-β- d-Gal pNAc4SO − 3-(1 → 4)-α- l-Ido pA-(1 → 3)- d-Gal pNAc4SO − 3 from dermatan sulfate. These results indicate that, unlike the conventional enzyme, highly purified chondroitinase ABC cannot degrade tetrasaccharides irrespective of their sulfation profiles. The enzymatic action is size-dependent.