The mechanism of leukocytolysis and hemolysis of the Staphylococcal leukocidin and gamma-hemolysin

葡萄球菌杀白细胞素和γ-溶血素的白细胞溶解和溶血机制

• 批准号: 09460042
• 负责人: KAMIO Yoshiyuki
• 金额: $ 8.58万
• 依托单位: Tohoku University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 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-09460042/
• 关键词: Staphylococcus aureus; Staphylococcal leukocidin; Staphylococcal gamma-hemolysine; Tow-component toxin; Pore-forming toxin; X-ray crystallography; 膜孔; 結晶解析; ブドウ球菌; プロテインキナーゼA

It is well known that the proteins which show gross similarities in their biological function(s) have primary structures which are similar to each other. However, recently even proteins which show dissimilarity in primary structures have been reported to have identical biological functions and the three-dimensional structures. The Staphylococcal channel-forming toxins are taken as an example. Staphylococcus aureus secretes leukocidin and gamma-hemolysin gamma -HL) into a culture medium as water soluble monomers. Leukocidin and r -HL contain two separate and synergistic protein components, LukF and LukS, and LukF and Hlg2 (or H gamma H), respectively. LukF and LukS of leukocidin, and LukF and Hlg2 of gamma -HL assemble into a ring-shaped complexes, in which the molar ratios of LukF to LukS or Hlg2, respectively are both 1 : 1, within human polymorphonuclear leukocyte membranes and erythrocyte membranes. In each case a transmembrane pore results with a functional diameter of 2.1-2.5 nm ; … More this leads to lyse the respective target cell. The cell specificities of leukocidin and gamma -HL are determined by LukS and Hlg2, respectively. Despite the 72% overall identity between the LukS and Hlg2 aminoacid sequences, the respective cell specificities are strictly defined, Within the year suported by Grant-in-Aid for Scientific Research (B), we clarified the following evidences.[I] The staphylococcal Panton-Valentine leukocidin (PVL) genes, [lukS-PV-lukF-PV] exist on the genome of a temperate bacteriophage phiPVL isolated from mitomycin C-induced Staphylococcus aureus V8 (ATCC 49775) [Kaneko et al., 1997. Biosci. Biotechnol. Biochem. 61, 1960-l962]. In this study, the complete nucleotide sequence of the phiPVL genome was analyzed, and the att sites (attL, attR, and attB) required for site-specific integration of phiPVL into the host chromosome were also determined.[II] Identification of the minimum segment essential for the Hlg2-specific function of staphylococcal gamma Hemolysin : The 5-residue K^<23>RLA^<127> of Hlg2 is the minimum segment essential for the Hlg2-specificfunction of staphylococcal gamma-hemolysin.[III] Identification of the minimum segment in which threonine 246 residue is a phosphorylation site by protein kinase A for the LukS-specific function of staphylococcal leukocidin : The minimum segment responsible for the LukS specific funcion of leukocidin was identified. We conclude that the 5-residue segment I^<242>K^<243>R^<244>S^<245>T^<246> is pivotal segment of LukS responsible for the LukS-function of the staphylococcal leukocidin.[IV] The Phosphorylation of LukS by Protein Kinase A is Crucial for the LukS-Specific Function of the Staphylococcal Leukocidin on Human Polymorphonuclear Leukocytes[V] Crystal structure of staphylococcal LukF delineates conformational changes accompaning formation of a transmembrane channel. Less

众所周知，在生物学功能上大体相似的蛋白质(S)具有彼此相似的一级结构。然而，最近，甚至在一级结构上表现出不同的蛋白质也被报道具有相同的生物学功能和三维结构。以金黄色葡萄球菌通道形成毒素为例。金黄色葡萄球菌将杀白素和γ-溶血素(γ-HL)作为水溶性单体分泌到培养基中。白血球蛋白和r-HL分别含有两个独立的、协同作用的蛋白质组分LukF和Luks，以及LukF和Hlg2(或H-Gamma H)。在人多形核白细胞膜和红细胞膜中，杀白素的LukF和LuKs，以及Gamma-HL的LukF和Hlg2组装成环状复合体，其中LukF与Luks或Hlg2的摩尔比均为1：1。在每种情况下，都会产生一个功能直径为2.1-2.5 nm的跨膜孔；…更重要的是，这会导致裂解相应的目标细胞。杀白素和γ-HL的细胞特异性分别由Luks和Hlg2决定。尽管Luks和Hlg2的氨基酸序列有72%的同源性，但各自的细胞特异性是严格定义的，在科学研究补助金(B)支持的一年内，我们澄清了以下证据：[i]葡萄球菌Panton-valine杀白素基因[Luks-PV-lukF-PV]存在于从丝裂霉素C诱导的金黄色葡萄球菌V8(Atcc 49775)中分离的温和噬菌体phiPVL的基因组上[Luks-PV-lukF-PV][Kaneko et al.，1997]。比奥西。生物技术。生物化学。61,1960-1962年]。在这项研究中，分析了phiPVL基因组的完整核苷酸序列，并确定了phiPVL定位整合到宿主染色体中所需的ATT位点(attL、AttR和attB)。[II]确定了葡萄球菌丙种溶血素Hlg2特异性功能所必需的最小片段：5-残基K^&lt；23&gt；Rla^&lt；127&gt；Hlg2是葡萄球菌γ-溶血素Hlg2特异性功能所必需的最小片段。[III]蛋白激酶A对LuKS特异性杀白素功能中苏氨酸246残基为磷酸化位点的最小片段的鉴定：确定了负责LuKS特异性杀白素功能的最小片段。我们的结论是，5个残基的I^&lt；242&gt；K^&lt；243&gt；R^&lt；244&gt；S^&lt；245&gt；T^&lt；246&gt；是LuKs的关键片段，负责葡萄球菌杀白素的LuKs功能。[IV]蛋白激酶A对LuKs的磷酸化对于葡萄球菌Leukosidin在人多形核白细胞上的LuKS专一性功能至关重要。较少

项目成果

Jun Kaneko: "Sequential binding of Staphylococcal γ-hemolysin to human erythrocytes and complex formmation of the hemolysin on the cell surface." Biosci.Biotech.Biochem.61(5). 846-851 (1997)

Jun Kaneko：“葡萄球菌 γ-溶血素与人红细胞的顺序结合以及细胞表面溶血素的复合物形成。”Biosci.Biotech.Biochem.61(5) (1997)。

Jun Kaneko: "An N-terminal region of LukF of Staphylococcal leukocidin/γ-hemolysin crucial for the biologica activity of the toxin" Biosci.Biotechnol.Biochem.62(7). 1465-1467 (1998)

Jun Kaneko：“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.Biotechnol.Biochem.62(7) (1998)。

J.Kaneko, A.L.Mascarenas, M.N.Huda, T.Tomita, and Y.Kamio: "An N-terminal region of LukF of Staphylococcal leukocidin/gamma-hemolysin crucial for the biologica activity of the toxin" Biosci.Biotechnol.Biochem.62 (7). 1465-1467 (1998)

J.Kaneko、A.L.Mascarenas、M.N.Huda、T.Tomita 和 Y.Kamio：“葡萄球菌杀白细胞素/γ-溶血素的 LukF N 末端区域对于毒素的生物活性至关重要”Biosci.Biotechnol.Biochem.62

H.Nariya et al: "Identification of the minimum segment in which threonine^<246> residue is phosphorylated by protein kinase A for the LukS-specific function of staphylococcal leukocidin." FEBS Lett. 415. 96-100 (1997)

H.Nariya 等人：“鉴定了苏氨酸 ^ 246 残基被蛋白激酶 A 磷酸化的最小片段，以实现葡萄球菌杀白细胞素的 LukS 特异性功能。”

Noriko Suigawara: "Assembly of Staphylococcus aureus γ-hemolysin into a pore-forming ring-shaped complex on the human erythrocytes" FEBS Lett.410. 333-337 (1997)

Noriko Suikawara：“金黄色葡萄球菌 γ-溶血素在人红细胞上组装成成孔环形复合物”FEBS Lett.410 (1997)。