Shoji H, Takahashi N, Nomoto H, Ishikawa M, Shimada I, Arata Y, Hayashi K
Department of Molecular Biology, Gifu Pharmaceutical University, Mitahora, Japan.
Eur J Biochem. 1992 Jul 15;207(2):631-41. doi: 10.1111/j.1432-1033.1992.tb17090.x.
The structures of the major oligosaccharide moieties of the nicotinic acetylcholine receptor (AcChoR) protein from Torpedo californica have been reported [Nomoto, H., Takahashi, N., Nagaki, Y., Endo, S., Arata, Y. and Hayashi, K. (1986) Eur. J. Biochem. 157, 233-242] to be high-mannose types. Here we report detailed analyses of the structures of the remaining oligosaccharides in this receptor. The sialylated oligosaccharides released by glycopeptidase (almond) digestion were separated according to the number of sialic acid residues using high-performance anion-exchange chromatography with pulsed amperometric detection. After removal of sialic acid from each fraction, the resulting neutral oligosaccharides were separately pyridylaminated and were analyzed by a combination of sequential exoglycosidase digestion and HPLC, then identified on a two-dimensional sugar map. The structures of two desialylated pyridylamino-oligosaccharides were further analyzed by high-resolution proton NMR. Each oligosaccharide was composed of species containing varying numbers of sialic acids. The desialylated complex-type oligosaccharides of AcChoR consisted of ten, eight and one different biantennary, triantennary and tetraantennary oligosaccharide, respectively. The biantennary oligosaccharides were divided into two groups; oligosaccharides with fucose at the proximal N-acetylglucosamine (six varieties) and oligosaccharides without fucose (four varieties). Each group consisted of species differing in the number of terminal galactose residues. The major component of the biantennary oligosaccharides had two galactose residues at the non-reducing termini. The terminal alpha-galactose residue(s) linked to C3 of beta-galactose were found in the fucose-containing biantennary oligosaccharides (two varieties). The triantennary oligosaccharides were also divided into two groups; oligosaccharides with (four varieties) and without (four varieties) besecting N-acetylglucosamine. These groups were composed of species differing in the number of terminal galactose residues. The major component of the triantennary oligosaccharides was fully galactosylated with three galactose residues. An unusual group, Gal beta 1-3GlcNAc, was present in low levels in the triantennary oligosaccharides. In contrast, the tetraantennary oligosaccharide was composed of only one species, which is fully galactosylated with four galactose residues.
据报道，来自加州电鳐的烟碱型乙酰胆碱受体（AcChoR）蛋白的主要寡糖部分的结构为高甘露糖型[野本浩、高桥直、长木洋、远藤慎、新田洋、林克（1986年），《欧洲生物化学杂志》157卷，233 - 242页]。在此，我们报告了对该受体中其余寡糖结构的详细分析。通过使用带脉冲安培检测的高效阴离子交换色谱，根据唾液酸残基的数量对经糖肽酶（杏仁）消化释放的唾液酸化寡糖进行分离。从每个馏分中去除唾液酸后，将所得的中性寡糖分别进行吡啶基胺化，并通过顺序外切糖苷酶消化和高效液相色谱相结合的方法进行分析，然后在二维糖图上进行鉴定。通过高分辨率质子核磁共振对两种去唾液酸化的吡啶基氨基寡糖的结构进行了进一步分析。每个寡糖都由含有不同数量唾液酸的种类组成。AcChoR的去唾液酸化复合型寡糖分别由十种、八种和一种不同的双天线、三天线和四天线寡糖组成。双天线寡糖分为两组；在近端N - 乙酰葡糖胺处带有岩藻糖的寡糖（六种变体）和不带岩藻糖的寡糖（四种变体）。每组都由末端半乳糖残基数量不同的种类组成。双天线寡糖的主要成分在非还原末端有两个半乳糖残基。在含岩藻糖的双天线寡糖（两种变体）中发现了连接到β - 半乳糖C3位的末端α - 半乳糖残基。三天线寡糖也分为两组；带有（四种变体）和不带有（四种变体）平分N - 乙酰葡糖胺的寡糖。这些组由末端半乳糖残基数量不同的种类组成。三天线寡糖的主要成分完全被三个半乳糖残基半乳糖基化。一个不寻常的基团，Galβ1 - 3GlcNAc，在三天线寡糖中含量较低。相比之下，四天线寡糖仅由一种组成，其完全被四个半乳糖残基半乳糖基化。
