Interaction of lipopolysaccharide with anticoagulant protein tachyplesin from hemocytes of horseshoe crab

脂多糖与鲎血细胞抗凝蛋白鲎素的相互作用

• 批准号: 03808036
• 负责人: KAWANO Keiichi
• 金额: $ 1.09万
• 依托单位: Kyushu University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (C)
• 财政年份: 1991
• 资助国家: 日本
• 起止时间: 1991 至 1993
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-03808036/
• 关键词: tachyplesin; tachycitin; tachystatin; horseshoe crab; endotoxin; lipopolysaccharide; antimicrobial protein; limulus test; ペプチド抗生物質; NMR

In humans, lipopolysaccharide (LPS) released during infection by Gram-negative bacteria can cause the severe pathological changes associated with septic shock. In the USA,septic shock is responcible for about 100,000 deaths annually and no specific drugs are available. LPS is the principal component of the outer leaflet of the outer membrane of Gram-negative bacteria. Lipid A,the membrane anchor of LPS,consists of a central phosphodisaccharide unit that is attached to up to seven fatty acid chains and it prossesses most of the biological activities of LPS.The toxicity in humans arises from the interaction of LPS or Lipid A with membrane-bound receptors or serum proteins, leading to an increase in the pro-inflammatory mediators (e.g.tomor necrosis factor, interleukin-1 and interleukin-6).Horseshoe crab (Tachypleus tridentatus) are ancient arachnids that possess a primitive circulatory system, the hemolymph, containing only one kind of cell, the hemocyte. Exposure of hemocytes to bacterial endotoxins (LPS) results in the activation of an intracellular coagulation cascade, a defense against microbial invasion. The system consists of several proteins, including tachyplesin, tachycitin and tachystatin that may inhibit the cascade. These are small basic proteins, which bind and neutralize LPS and have strong anti-bacterial effects on the growth of Gram-negative bacteria.In this study, we studied the interaction of tachyplesin with synthetic lipid A and found that negative charges of phsphate groups of lippid A interact with positive charges of tachyplesin. The amphipathic loop was proposed as an important motif of LPS binding site and may be used in the design of molecules with therapeutic properties against septic shock.distance geometry calculation by using X-PLOR.In the structure of tachycitin, we found the chitinbinding motif like hevein. The main chain structure of tachystatin was similar to that of Ca-channel blocker omega-conotoxin.

在人类中，革兰氏阴性菌在感染过程中释放的脂多糖（LPS）可引起与感染性休克相关的严重病理变化。在美国，感染性休克每年造成约10万人死亡，并且没有特定的药物可用。LPS是革兰氏阴性菌外膜外小叶的主要成分。Lipid A是LPS的膜锚，由一个磷酸化二糖单元连接到多达7个脂肪酸链上，具有LPS的大部分生物学活性，其对人体的毒性是由于LPS或Lipid A与膜结合受体或血清蛋白相互作用，导致促炎介质的增加鲎（Tachypleus tridentatus）是古老的蜘蛛纲动物，其具有原始的循环系统，血淋巴，仅含有一种细胞，血细胞。血细胞暴露于细菌内毒素（LPS）导致细胞内凝血级联反应的激活，这是对微生物入侵的防御。该系统由几种蛋白质组成，包括tachyplesin，tachycitin和tachystatin，它们可以抑制级联反应。这些小的碱性蛋白质结合并中和LPS，并对革兰氏阴性菌的生长有很强的抗菌作用。在本研究中，我们研究了鲎素与合成脂质A的相互作用，发现脂质A的磷酸基团的负电荷与鲎素的正电荷相互作用。两亲环是LPS结合位点的一个重要模体，可用于设计抗感染性休克的分子。其主链结构与钙通道阻断剂ω-芋螺毒素相似。

项目成果

阿久津 秀雄: "他核のNMR" 廣川書店, (1992)

阿久津秀夫：“其他原子核的核磁共振”广川书店，（1992）

Iori Maeda: "Water-soluble chy motrypsin specific inhibitors containing arginine" Biochemical and Biophysical Research Communication. 193. 428-433 (1993)

Iori Maeda：“含有精氨酸的水溶性糜蛋白酶特异性抑制剂”生物化学和生物物理研究交流。

T.suetake, S.Tsuda, S.Kawabaata, K.Kawano, K.Miura, K.Hikichi and K.Nitta: "Three-dimensional structure of tachycitin in solution determined by ^1H NMR." R.P.P.P.J.(in press).

T.suetake、S.Tsuda、S.Kawabaata、K.Kawano、K.Miura、K.Hikichi 和 K.Nitta：“通过 ^1H NMR 测定溶液中速蛋白的三维结构。”

Hatsumi Nagadome: "Identification of the adsovbing site of lysozyme onto the hydroxy-apatite surface using hydrogen exchange and HNMR" FEBS Letters. 317. 128-130 (1993)

Hatsumi Nagadome：“利用氢交换和 HNMR 鉴定溶菌酶在羟基磷灰石表面上的吸附位点”FEBS Letters。

T. Suetake他: "Three-dimensional structure of tachycitin in solution determined by ^1H NMR" R.P.P.P.J.(印刷中).

T. Suetake 等人：“通过 1H NMR 测定溶液中速霉素的三维结构”R.P.P.P.J（出版中）。