MODE OF ACTION OF STAPHYLOCOCCAL LEUKOCIDIN AND gamma-HEMOLYSIN

金黄色葡萄球菌杀白细胞素和γ-溶血素的作用方式

• 批准号: 09670275
• 负责人: TOMITA Toshio
• 金额: $ 2.05万
• 依托单位: FACULTY OF AGRICULTURE,TOHOKU UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1997
• 资助国家: 日本
• 起止时间: 1997 至 1998
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-09670275/
• 关键词: Staphylococcus aureus; bi-component cytolysin; staphylococcal gamma-hemolysin; staphylococcal leukocidin; pore-forming toxin; complex formation; ring structure; erythrocyte; neutrophil; リング状構造

Staphylococcal gamma-hemolysin consists of Hlg1 (or Luk F) and Hlg2, which cooperatively lyze human erythrocytes. Since gamma-hemolysin caused swelling of human erythrocytes before lysis, we studied pore-forming nature of the toxin by use of polyethylene glycols as osmotic protectants and determined the functional diameter of the pore. To elucidate the molecular architecture of the membrane pore formed by gamma-hemolysin, we solubilized the pore complex with 2 % sodium dodecyl sulfate, purified it by sucrose gradient centrifugation, and observed the purified pore complex under an electron microscope. Our data showed that the Hlg1 and the Hlg2 components assemble into a ring-shaped 200 kDa complex in a molar ratio of 1 : 1, forming a membrane pore with a functional diameter of 2.1-2.4 nm.Staphylococcal leukocidin consists of LukS and LukF, which lyse human and rabbit polymorphonuclear leukocytes and rabbit erythrocytes. Here we studied the pore-forming properties of leukocidin and the molecular architecture of the leukocidin pore. (1) Leukocidin caused an efflux ofpotassium ions from rabbit erythrocytes and swelling of the cells before lysis. However, ultimate lysis of the swollen cells did not occur when polyethylene glycols with hydrodynamic diameters of <greater than or equal> 2.1 nm were present in the extracellular space. (2) Electron microscopy showed that the leukocidin formed a ring-shaped structure with outer and inner diameters of 9 and 3 nm, respectively, on human polymorphonuclear leukocytes and rabbit erythrocytes. (3) Ring-shaped structures of the same dimensions were isolated from the target cells, and they contained LukS and LukF in a molar ratio of 1 : 1. (4) A single ring-shaped toxin complex had a molecular size of 205 kDa. These results indicated that LukS and LukF assemble into a ring-shaped oligomer of approximately 200 kDa on the target cells, forming a membrane pore with a functional diameter of approximat

葡萄球菌γ溶血素由Hlg1（或Luk F）和Hlg2组成，它们协同溶解人红细胞。由于γ -溶血素在溶解前引起人红细胞肿胀，我们使用聚乙二醇作为渗透保护剂研究了毒素的成孔性质，并确定了孔的功能直径。为了阐明溶血素形成的膜孔的分子结构，我们用2%十二烷基硫酸钠将孔复合物溶解，用蔗糖梯度离心纯化，并在电镜下观察纯化后的孔复合物。我们的数据表明，Hlg1和Hlg2组分以1:1的摩尔比组装成一个200 kDa的环状配合物，形成一个功能直径为2.1-2.4 nm的膜孔。葡萄球菌的杀白细胞素由LukS和LukF组成，它们能裂解人和兔的多形核白细胞和兔的红细胞。本文研究了白细胞素的成孔特性和白细胞素孔的分子结构。(1)杀白细胞素引起兔红细胞钾离子外排，溶解前细胞肿胀。然而，当水动力直径<大于或等于bbb2.1 nm的聚乙二醇存在于细胞外空间时，肿胀细胞不会发生最终的裂解。(2)电镜观察发现，在人多形核白细胞和兔红细胞上，杀白细胞素形成外径为9 nm、内径为3 nm的环状结构。(3)从靶细胞中分离出相同尺寸的环状结构，其含有LukS和LukF，摩尔比为1:1。(4)单个环状毒素复合物的分子量为205 kDa。这些结果表明，LukS和LukF在靶细胞上组装成约200 kDa的环状低聚物，形成一个功能直径约为的膜孔

项目成果

Sugawara,N., Tomita,T., and Kamio,Y.: "Assembly of Staphylococcus aureus γ-hamolysin into a pore-forming,ring-shapged complex on the surface of human erythrocytes" FEBS Lett.410. 333-337 (1997)

Sukawara, N.、Tomita, T. 和 Kamio, Y.：“金黄色葡萄球菌 γ-溶血素在人红细胞表面组装成成孔环形复合物” FEBS Lett.410（ 1997）

Sugawara, N., Tomita, T., Sato, T., and kamio, Y.: "Assembly of staphylococcal leukocidin into a pore-forming ring-shaped complex on rabbit erythrocytes and human polymorphonuclear leukocytes" Biosci.Biotechnol.Biochem.63・5印刷中. (1999)

Sukawara, N.、Tomita, T.、Sato, T. 和 kamio, Y.：“将葡萄球菌杀白细胞素组装成兔红细胞和人多形核白细胞上的成孔环形复合物” Biosci.Biotechnol.Biochem.63・已出版 5 期（1999 年）。

Kaneko, J.et al.: "An N-terminal region of LukF of staphylococcal leuko-cidin/gamma-hamolysin crucial for the biological activity of the toxin" Biosci.Biotechnol.Biochem.62-7. 1465-1467 (1998)

Kaneko, J. 等人：“葡萄球菌白细胞杀素/γ-单溶素 LukF 的 N 末端区域对于毒素的生物活性至关重要”Biosci.Biotechnol.Biochem.62-7。

Kaneko, J., Mascamas, M.A., Huda, N., Tomita, T., and Kamio, Y.: "An N-terminal region of LukF of staphylococcal leukocidin/γ-hamolysin crucial for the biological activity of the toxin" Biosci.Biotech.Biochem.62・7. 1465-1467 (1998)

Kaneko, J.、Mascamas, M.A.、Huda, N.、Tomita, T. 和 Kamio, Y.：“葡萄球菌杀白细胞素/γ-溶血素的 LukF 的 N 末端区域对于毒素的生物活性至关重要”Biosci .生物技术.生物化学.62・7.

Tomita,T., Ishikawa, D., Noguchi,T., Katayama, E., and Hashimoto, Y.: "Assembly of flammutoxin, a cytolytic protein from the edible mushroom Flammulina velutipes, into a pore-forming ring-shaped oligomer on the target cell" Biochem.J.333・1. 129-137 (1998)

Tomita, T.、Ishikawa, D.、Noguchi, T.、Katayama, E. 和 Hashimoto, Y.：“金针菇毒素（一种来自食用蘑菇金针菇的溶细胞蛋白）组装成成孔环形低聚物靶细胞”Biochem.J.333・1.129-137(1998)