Structure-function studies of defense molecules found in horseshoe crab hemocytes

鲎血细胞中防御分子的结构功能研究

基本信息

批准号：
08458183
负责人：
KAWABATA Shun-ichiro
金额：
$ 5.38万
依托单位：
KYUSHU UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
1996
资助国家：
日本
起止时间：
1996 至 1997
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-08458183/
关键词：
Host defense Innate immunity Lipopolysaccharide Lectin リピドA

项目摘要

We identified a novel horseshoe crab hemocyte-derived lectin, which we named tachylectin-4. It has more potent hemagglutinating activity against human A-tupe erythrocytes than a previously identified hemocyte lectin with an affinity to N-acetylglucosamine, tachylectin-2. The purified tachylectin-4 is an oligomeric glycoprotein of 470 kDa, composed of subunits of 30 kDa and 31.5 kDa. Ca2+ at 10mM enhanced the hemagglutinatig activity 4-fold and the activity was inhibited by EDTA and o-phenanthroline. L-Fucose and N-acetylneuraminic acid at 100 mM completely inhibited the activity of tachylectin-4. The activity was also inhibited more strongly by bacterial S-type lipopolysaccharides (LPS) but not by R-type LPS lacking O-antigen. The most effective S-type LPS was from Escherichia coil O111 : B4 and the minimum concentration required for inhibiting agglutination against human A-type erythrocytes (0.1mg/ml) was 160-fold lower than those of S-type LPS from Salmonella minnesota. Therefore, colitose (3-deoxy-L-fucose), a unique sugar present in the O-antigen of Escherichia coli O111 : B4 with structural similarity to L-fucose, is the most probable candidate for a specific ligand of tachylectin-4. A cDNA coding for tachylectin-4 was isolated from a hemocyte cDNA library. The open reading frame of the 1344-base pair cDNA coded for the mature protein with 232 amino acids. There is no significant sequence similarity to any other known LPS-binding lectins, whereas tachylectin-4 is homologous to the NH2-terminal domain with unknown functions of Xenopus laevis pentraxin 1.

我们鉴定了一种新型鲎血细胞来源的凝集素，我们将其命名为 tachylectin-4。它对人 A 型红细胞的血凝活性比先前鉴定的与 N-乙酰葡糖胺（tachylectin-2）有亲和力的血细胞凝集素更有效。纯化的tachylectin-4是一种470 kDa的寡聚糖蛋白，由30 kDa和31.5 kDa的亚基组成。 10mM Ca2+ 使血凝活性增强 4 倍，但该活性被 EDTA 和邻菲咯啉抑制。 100 mM 的 L-岩藻糖和 N-乙酰神经氨酸完全抑制 tachylectin-4 的活性。该活性还受到细菌 S 型脂多糖 (LPS) 的更强抑制，但不受缺乏 O 抗原的 R 型脂多糖的抑制。最有效的S型LPS来自大肠杆菌O111:B4，抑制人A型红细胞凝集所需的最低浓度(0.1mg/ml)比来自明尼苏达沙门氏菌的S型LPS低160倍。因此，大肠糖（3-脱氧-L-岩藻糖）是一种存在于大肠杆菌 O111 : B4 的 O 抗原中的独特糖，其结构与 L-岩藻糖相似，是 tachylectin-4 特定配体最有可能的候选者。从血细胞 cDNA 文库中分离出编码 tachylectin-4 的 cDNA。 1344 个碱基对 cDNA 的开放阅读框，编码含 232 个氨基酸的成熟蛋白。与任何其他已知的 LPS 结合凝集素没有显着的序列相似性，而 tachylectin-4 与 Xenopus laevis pentraxin 1 的 NH2 末端结构域同源，功能未知。