Sugahara K, Tanaka Y, Yamada S, Seno N, Kitagawa H, Haslam S M, Morris H R, Dell A
Department of Biochemistry, Kobe Pharmaceutical University, Higashinada-ku, Kobe 658, Japan.
J Biol Chem. 1996 Oct 25;271(43):26745-54. doi: 10.1074/jbc.271.43.26745.
We prepared a series of oligosaccharides from king crab cartilage chondroitin sulfate K after exhaustive digestion with testicular hyaluronidase, and determined the structures of four tetrasaccharides and a pentasaccharide by fast atom bombardment mass spectrometry, high performance liquid chromatography analysis of chondroitinase AC-II digests, and 500-MHz 1H NMR spectroscopy. The tetrasaccharides shared the common core structure GlcAbeta1-3GalNAcbeta1-4GlcAbeta1-3GalNAc with various sulfation profiles. One structure was GlcAbeta1-3GalNAc(4S)beta1-4GlcAbeta1-3GalNAc(4S), whereas three of them have the following hitherto unreported structures including a novel glucuronate 3-O-sulfate: GlcA(3S)beta1-3GalNAc(4S)beta1-4GlcAbeta1-3GalNAc(4S), GlcAbeta1-3GalNAc(4S)beta1-4GlcA(3S)beta1-3GalNAc(4S), and GlcA(3S)beta1-3GalNAc(4S)beta1-4GlcA(3S)beta1-3GalNAc(4S), where 3S or 4S represents 3-O- or 4-O-sulfate, respectively. The structure of the pentasaccharide was determined as GlcA(3S)beta1-3GalNAc(4S)beta1-4GlcA(3S)beta1- 3GalNAc(4S)beta1-4GlcA. Chondroitinase ABC digestion of the tetrasaccharides with GlcA(3S) at the internal position destroyed the disaccharide unit containing GlcA(3S) derived from the reducing side and resulted in only the disaccharide unit from the non-reducing side. In contrast, these tetrasaccharides remained totally resistant to chondroitinase AC-II. The results indicated that it is necessary to reevaluate the disaccharide composition of chondroitin sulfate poly- or oligosaccharides purified from various biological sources, since they were usually determined after chondroitinase ABC digestion. It is probable that the structures containing GlcA(3S) would not have been detected.
我们用睾丸透明质酸酶彻底消化帝王蟹软骨硫酸软骨素K后制备了一系列寡糖，并通过快原子轰击质谱、软骨素酶AC-II消化产物的高效液相色谱分析以及500兆赫的1H核磁共振光谱确定了四种四糖和一种五糖的结构。这些四糖具有共同的核心结构GlcAbeta1-3GalNAcbeta1-4GlcAbeta1-3GalNAc，但硫酸化模式各不相同。一种结构是GlcAbeta1-3GalNAc(4S)beta1-4GlcAbeta1-3GalNAc(4S)，而其中三种具有以下迄今未报道的结构，包括一种新型的葡萄糖醛酸3-O-硫酸盐：GlcA(3S)beta1-3GalNAc(4S)beta1-4GlcAbeta1-3GalNAc(4S)、GlcAbeta1-3GalNAc(4S)beta1-4GlcA(3S)beta1-3GalNAc(4S)和GlcA(3S)beta1-3GalNAc(4S)beta1-4GlcA(3S)beta1-3GalNAc(4S)，其中3S或4S分别代表3-O-或4-O-硫酸盐。五糖的结构确定为GlcA(3S)beta1-3GalNAc(4S)beta1-4GlcA(3S)beta1- 3GalNAc(4S)beta1-4GlcA。对内部位置含有GlcA(3S)的四糖进行软骨素酶ABC消化会破坏来自还原端的含有GlcA(3S)的二糖单元，仅产生来自非还原端的二糖单元。相比之下，这些四糖对软骨素酶AC-II完全具有抗性。结果表明，有必要重新评估从各种生物来源纯化的硫酸软骨素多糖或寡糖的二糖组成，因为它们通常是在软骨素酶ABC消化后确定的。含有GlcA(3S)的结构很可能未被检测到。
