Studies on protective function of glycoprotein in fish blood cell membrane(glycophorin) toward fish pathogen.

鱼类血细胞膜糖蛋白（糖蛋白）对鱼类病原体的保护作用研究。

• 批准号: 12660184
• 负责人: AOKI Takahiko
• 金额: $ 2.05万
• 依托单位: Mie University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2000
• 资助国家: 日本
• 起止时间: 2000 至 2002
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-12660184/
• 关键词: RED CELL MEMBRANE; GLYCOPROTEIN; GLYCOPHORIN; SIALIC ACID; OLIGOSACCHARIDE; FISH; 血液; O-グリコシド型糖鎖; NGNA

Glycophorin is a transmembrane sialoglycoprotein in the red cell membrane from mammalian and avian blood. Carp (Cyprinus carpio) blood was drawn, and prepared the carp red cell membrane. Membrane proteins and glycophorin were examined by SDS- polyacrylamide gel electrophoresis. The Coomassie blue stain pattern of carp red cell membrane was similar to that of rainbow trout and human. The prominent membrane protein, band 3 which designated for human red cell membrane, was detected as well as spectrin, ankyrin and actin. With PAS stain, glycoprotein was detected in the carp red cell membrane. Carp glycophorin was extracted from the membrane by LIS-phenol method, and this preparation showed weak N blood type activity. Sialic acid in carp glycophorin was determined NGNA by thin layer chromatography and other methods. From the result of amino acid analysis, composition was similar to human glycophorin. However, composition of oligosaccharide in carp glycophorin was differing from human oligo … More saccharide. The interaction of carp glycophorin and various lectins showed that the components of carp glycophorin's oligossacharides were O-linked (Gal・GalNAc・Ser/Thr), α Glc and Fuc. After β-elimination, released oligosaccharides from glycophorin were collected, and separated two fractions by using anion exchange column chromatography (HPLC). These fractions were hydrolyzed, following by analysis of high performance anion exchange chromatography with pulsed amperometric detector (HPAEC-PAD). The results from HPAEC-PAD also suggested that the components of oligosaccharides were Gal, Glc and Fuc. Although this results being strengthen by NMR spectra, the structure of these oligosaccharides were not determined without data from GC-MS analysis. We are going to preparation of mg-order desalted oligosaccharides for GC-MS analysis.From the observation by electron microscope, it seemed that carp glycophorin molecules attached to the flagellum of gram-negative bacterium and to the cell wall of gram-positive bacterium. These results suggested that carp glycophorin has a protective function toward fish pathogen. Less

血型糖蛋白是一种跨膜唾液酸糖蛋白，存在于哺乳动物和鸟类的红细胞膜上。取鲤鱼血，制备鲤鱼红细胞膜。SDS-聚丙烯酰胺凝胶电泳检测膜蛋白和血型糖蛋白。鲤鱼红细胞膜的考马斯亮蓝染色模式与虹鳟鱼和人的相似。主要的膜蛋白，带3，指定为人类红细胞膜，以及血影蛋白，锚蛋白和肌动蛋白被检测到。PAS染色可见鲤鱼红细胞膜糖蛋白。采用LIS-苯酚法从膜中提取鲤鱼血型糖蛋白，该蛋白具有弱的N血型活性。采用NGNA薄层色谱法等方法测定鲤鱼血型糖蛋白中唾液酸的含量。从氨基酸分析结果来看，其组成与人血型糖蛋白相似。但鲤鱼血型糖蛋白中的寡糖组成与人血型糖蛋白中的寡糖组成不同， ...更多信息 糖类鲤鱼血型糖蛋白与各种凝集素的相互作用表明，鲤鱼血型糖蛋白的低聚糖组成为O-连接的（Gal·GalNAc·Ser/Thr）、α Glc和Fuc。经β-消除后，收集从血型糖蛋白释放的寡糖，并通过阴离子交换柱色谱（HPLC）分离两个级分。将这些组分水解，然后通过高效阴离子交换色谱与脉冲安培检测器（HPAEC-PAD）进行分析。HPAEC-PAD的结果也表明，低聚糖的组成为Gal、Glc和Fuc。虽然这一结果得到了NMR光谱的加强，但没有GC-MS分析的数据，这些寡糖的结构无法确定。通过电镜观察发现，鲤鱼血型糖蛋白分子主要附着在革兰氏阴性菌的鞭毛上，革兰氏阳性菌的细胞壁上。说明鲤鱼血型糖蛋白对鱼类病原菌具有一定的保护作用。少