Cellular Localization, qurification, and function of plasmud-determinedrirulence proteins of Salmonella

沙门氏菌质粒决定的毒性蛋白的细胞定位、纯化和功能

• 批准号: 05670264
• 负责人: DANBARA Hirofumi
• 金额: $ 1.34万
• 依托单位: KITASATO,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: 1993
• 资助国家: 日本
• 起止时间: 1993 至 1994
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-05670264/
• 关键词: salmonella; Virulence; Plasmid; Regulation of expression; Virulance protein; lmmano-Electron Microscopy; 機能; 精製; 細胞局在性; 発現; 免疫電顕

Our previous studies showed that S.Choleraesuis strain RF-1 isolated from septicemic pig harbors 50kb-virulence plasmid designated pKDSC50, and the 6.4kb SalI-EcoRI region of pKDSC50 encompassing four spv genes, spvR,spvA,spvB,and spvC,is essential to produce systemic disease in pigs and mice. The spvR gene was characterized as a positive activator gene for transcription of the following three genes, spvA,spvB,and spvC.The aim of the present study is to obtain the basic knowledges of Spv proteins which are requird for understanding the Salmonella systemic infection.1. Regulatory function of Spv proteinsSpvR is a positive regulatory protein for the expression of its own gene, spvR.SpvA and SpvBare negative regulatory proteins for the expression of spvR gene. spvR gene is expressed in RpoS-dependent manner at the stationary phase of bacterial growth.2. Isolation, purification, and characterization of Spv proteinsThe four Spv proteins, spvR,spvA,spvB,and spvC were partially purified from S.Choleraesuis strain RF-1. Molecular weight of the purified Spv proteins by SDS-PAGE was determinded as 31 kDa for SpvR,26 kDa for SpvA,73 kDa for SpvB,and 24 kDa for SpvC.It was demonstrated that SpvB is a hydrophobic protein, and SpvC is a basic protein.3. Immunoelectron microscopic localization of Spv proteinsA method for the localizatiom of Spv proteins by immunoelectron microscopy was established. By using cryo ultra-thin section of S.Choleraesuis strain RF-1 fixed with periodete lysin-paraformaldehyde, SpvC was revealed to be a cytoplasmic and outer menbrane protein. The localization of SpvR,SpvA,and SpvB proteins is under investigation.

我们先前的研究表明，从败血症猪中分离出的S.胆碱菌菌株RF-1拥有50KB - 病毒质粒指定为PKDSC50，而PKDSC50的6.4kb Sali-Ecori区域包括四个SPV基因，SPVR，SPVR，SPVR，SPVA，SPVA，SPVB，SPVB和SPVC，对猪的生产和MICE是本质的。 SPVR基因的特征是用于以下三个基因SPVA，SPVB和SPVC转录的阳性激活因子基因。本研究的目的是获得SPV蛋白的基本知识，这是理解沙门氏菌系统感染的必要条件。1。 SPV ProteInsSPVR的调节功能是表达其自身基因SPVR.SPVA和SPVBARE负调控蛋白的阳性调节蛋白，用于SPVR基因的表达。 SPVR基因在细菌生长的固定阶段以RPOS依赖性方式表达。2。 SPV蛋白的分离，纯化和表征四种SPV蛋白，SPVR，SPVA，SPVB和SPVC被部分从S.Choleraesuis菌株RF-1中部分纯化。通过SDS-PAGE纯化的SPV蛋白的分子量被确定为SPVR的31 kDa，SPVA的26 kDa，SPVB的73 kDa和SPVC的24 kDa，这证明了SPVB是疏水蛋白，SPVB是一种基本蛋白质。3。通过免疫电子显微镜对SPV蛋白的SPV蛋白质方法的免疫电子显微镜定位进行了定位。通过使用用周期溶质蛋白 - 丙甲甲醛固定的S.胆碱菌菌株RF-1的冷冻超薄截面，SPVC被揭示为细胞质和外神经蛋白。正在研究SPVR，SPVA和SPVB蛋白的定位。

项目成果

Masahiko Endoh et.al: "lsolation, Purification and Characterization of Salmonella plasmid virulence proteins of Salmonella Choleraesuis" Infection and Immunology. (in preparation).

Masahiko Endoh 等人：“猪霍乱沙门氏菌质粒毒力蛋白的分离、纯化和表征”感染和免疫学。

Kachiko Sekiya: "Immunoelectron microscopic localization of Salmonella Plasmid virulence proteins of Salmonella Choleraesuis" Infection and Immunology. (in preparation).

Kachiko Sekiya：“猪霍乱沙门氏菌质粒毒力蛋白的免疫电镜定位”感染与免疫学。

Masahiro Endoh et al: "Isolation,Purification,and Charavterization of Salmonella Plasmid virulence proteins of Salmonella Choleraesuis" Infection and Immunology. (in preparation).

Masahiro Endoh 等人：“猪霍乱沙门氏菌质粒毒力蛋白的分离、纯化和表征”感染和免疫学。

Masahiko Endoh: "Isolation,purification,and characterization of Salmonella Plasmid virulence proteins of Salmonella Choleraesuis" Infection and Immunology. (in preparation).

Masahiko Endoh：“猪霍乱沙门氏菌质粒毒力蛋白的分离、纯化和表征”感染和免疫学。

Akio Abe et al: "Regulation of Spv gene expression of Salmonella virulence plasmid pkDSC50 in Salmonella choleraesuis serovar Choleraesuis" Molecular Microbiology. 12. 779-787 (1994)

Akio Abe 等人：“猪霍乱沙门氏菌血清型猪霍乱中沙门氏菌毒力质粒 pkDSC50 的 Spv 基因表达的调节”分子微生物学。