Genetic and Biochemical Analysis of a Peptide Export System

提出的实验的目的是阐明 细菌中肽和蛋白质输出的机制。 具体来说，所使用的系统是Microcin的导出 B17（MccB 17）是一种由E.杆菌 六 基因，mcbB到mcbG，是生产所必需的， 对MccB 17的免疫力，MccB 17本身是McbA的产物。 的 抗生素作用于敏感的肠道细菌细胞， DNA复制。 mcbE和mcbF基因产物似乎 介导MccB 17的输出; mcbE和mcbF中的突变， 其基因外抑制因子将描述其作用， 相互作用，以及这些组件的功能域 MccB 17出口机械。 细胞内和细胞外 MccB 17将通过生物化学和细胞生物学表征， 分馏研究和电子显微镜。 突变 阻止其输出的MccB 17分子中的 转运的功能域; 这些允许输出改变的肽的突变将 鉴定输出的肽与 出口所需的蛋白质。 斯克维斯基博士正在确立自己的独立地位 研究人员通过这一研究机会的妇女 研究启动奖。 这项工作将是 有趣的是，蛋白质出口到外部的 细胞是罕见的细菌大肠杆菌。 的信息 在这些研究中获得的将有许多可能的 商业化生产的实际应用 细菌中克隆基因的产物。*//

The objective of the experiments proposed is to elucidate mechanisms of peptide and protein export in bacteria. Specifically, the system to be used is the export of Microcin B17 (MccB17), a peptide antibiotic produced by E. coli. Six genes, mcbB through mcbG, are necessary for production of and immunity to MccB17, itself the product of mcbA. The antibiotic acts on sensitive enteric bacterial cells to block DNA replication. The mcbE and mcbF gene products appear to mediate export of MccB17; mutations in mcbE and mcbF and extragenic suppressors thereof will delineate the roles, interactions, and functional domains of these components of the MccB17 export machinery. Intracellular and extracellular MccB17 will be characterized by biochemical and cell fractionation studies and by electron microscopy. Mutations in the MccB17 molecule that prevent its export will identify functional domains for transport; extragenic suppressors of these mutations that allow export of the altered peptide will identify interactions between the exported peptide and the proteins necessary for its export.%%% Dr. Skvirsky is establishing herself as an independent investigator through this Research Opportunities for Women Research Initiation award. The work promises to be interesting because export of proteins to the exterior of the cell is rare in the bacterium E.coli. The information obtained in these studies will have numerous possible practical applications to the commercial production of products of cloned genes in bacteria.***//