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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.

我们鉴定了一种新的鲎血细胞凝集素，我们命名为tachylectin-4。它对人A型红细胞的血凝活性比先前鉴定的对N-乙酰葡糖胺具有亲和力的血细胞凝集素速凝素-2更强。纯化的速凝素-4是470 kDa的寡聚糖蛋白，由30 kDa和31.5 kDa的亚基组成。10 mM的Ca ~（2+）使其凝集活性增强4倍，EDTA和邻菲咯啉则抑制其凝集活性。100 mM的L-岩藻糖和N-乙酰神经氨酸完全抑制速凝素-4的活性。细菌S型脂多糖（LPS）也能更强烈地抑制该活性，但不受缺乏O抗原的R型LPS的抑制。大肠杆菌O 111：B4的S型LPS对人A型红细胞凝集抑制作用最强，其最低抑制浓度（0.1mg/ml）比沙门氏菌的低160倍。因此，大肠杆菌O 111：B4 O-抗原中存在的一种独特的糖--粘三糖（3-脱氧-L-岩藻糖）与L-岩藻糖结构相似，是最有可能成为速凝素-4特异性配体的候选物。从血细胞cDNA文库中分离到一个编码速凝素-4的cDNA。该cDNA的开放阅读框为1344个碱基，编码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，而速凝素-4与非洲爪蟾pentraxin 1的NH 2-末端结构域具有未知的功能。