ISOLATION AND CHARACTERIZATION OF E COLI MUTANTS CONDITIONALLY DEFECTIVE

条件缺陷大肠杆菌突变体的分离和表征

• 批准号: 7609950
• 负责人: PUSHPA SAMKUTTY
• 金额: $ 2.29万
• 依托单位: LOUISIANA STATE UNIV A&M COL BATON ROUGE
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-05-01 至 2008-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7609950
• 关键词: Anabolism; Biogenesis; Computer Retrieval of Information on Scientific Projects Database; Condition; Cytoplasm; Data; Funding; Grant; Growth; Immunoprecipitation; Institution; Lipopolysaccharides; Membrane; Membrane Proteins; Molecular Chaperones; Numbers; Phospholipids; Proteins; Reporting; Research; Research Personnel; Resources; Source; Temperature; Transmembrane Transport; United States National Institutes of Health; Western Blotting; lipid transport; mutant; periplasm; temperature sensitive mutant; three dimensional structure

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The analysis of outer membrane protein biogenesis in an E.coli mutant defective in lipid transport was the focal point of this study. Following biosynthesis in the cytoplasm, proteins destined for the E.coli outer membrane (OM) are transported across the inner membrane (IM) by the Sec machinery. Once in the periplasm, they are folded into their correct three dimensional structures by a number of molecular chaperones. This folding has been reported to require lipopolysaccharide (LPS) for a number of OM proteins, including OmpA. Here, we present data on the folding of OmpA in a temperature-sensitive mutant of MsbA that is defective for export of LPS and phospholipids to the OM under nonpermissive conditions. W3110 (wildtype) and WD2 (MsbA mutant) were grown at both permissive (30¿C) and nonpermissive (44¿C) temperatures and analyzed for levels of unfolded OmpA using western blotting and immunoprecipitation. We found that there is no detectable increase in unfolded OmpA in MsbA mutant WD2 during growth under nonpermissive conditions.

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 本研究的重点是对一株脂质转运缺陷的大肠杆菌突变株的外膜蛋白生物发生进行分析。在细胞质中生物合成后，进入大肠杆菌外膜(OM)的蛋白质通过SEC机制穿过内膜(IM)。一旦进入周质，它们就被一些分子伴侣折叠成正确的三维结构。据报道，这种折叠需要许多OM蛋白的脂多糖(LPS)，包括OmpA。在这里，我们提供了关于MSBA的温度敏感突变体中OmpA折叠的数据，该突变体在不允许的条件下存在向OM出口内毒素和磷脂的缺陷。W3110(野生型)和WD2(MSBA突变体)在允许的(30℃)和不允许的(44℃)两种温度下生长，并用免疫印迹和免疫沉淀分析未折叠OmpA的水平。我们发现，在不允许的条件下，MSBA突变体WD2在生长过程中没有检测到未折叠OmpA的增加。