REGULATION OF S MUTANTS PHOSPHOTRANSFERASE SYSTEM

S突变体磷酸转移酶系统的调控

• 批准号: 2131840
• 负责人: ALLEN L HONEYMAN
• 金额: $ 9.99万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1994
• 资助国家: 美国
• 起止时间: 1994-09-01 至 1999-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2131840
• 关键词: Escherichia coli; Streptococcus mutans; bacterial genetics; bacterial proteins; carbohydrate metabolism; carbohydrate transport; chloramphenicol acetyltransferase; fusion gene; gene expression; genetic promoter element; genetic regulation; molecular cloning; mutant; northern blottings; nucleic acid sequence; operon; phosphoenolpyruvate; phosphotransferases; recombinant DNA

The long range goal of this proposal is to understand the mechanism of catabolite repression which effects and controls carbohydrate utilization in Streptococcus mu tans and how this process is involved in the pathogenesis of S. mutans-induced dental caries. The mechanism of catabolite repression is not understood in Gram positive bacteria and elucidation of this process will potentially allow better prevention of dental caries. However, this long range goal can not be achieved without first attaining a basic understanding of the local regulation of the phosphoenolpyruvate-dependent phosphotransferase system (PTS) Of carbohydrate transport in S. mutans. Therefore, the short term goal of this proposal is to initiate studies into the regulation of the S. mutans PTS. The individual specific aims of this proposal include: 1.) isolation and cloning in E. coli K12 of S. mutans gene clusters involved in the transport and utilization of various carbohydrates via the PTS; 2.) identification of the gene products of the isolated PTS gene clusters by in vitro gene expression systems; 3.) determination of the nucleotide sequences of the isolated PTS gene clusters to determine the overall operon structures, to identify regions for the in vitro generation of transcriptional and translational fusions, and to identify putative regulatory control regions; 4.) in vitro generation of transcriptional fusions to monitor regulation of the S. mutans PTS in response to various carbohydrates; 5.) analysis of the putative promoter regions of the isolated PTS gene clusters and identification and analysis of putative control regions involved with regulation of these operons using translational fusions; and 6.) assay of transcriptional activity of the S. mutans PTS catabolic enzyme genes under various environmental conditions. Accomplishment of these specific aims will provide the necessary information to initiate studies into the long range goal and will be paramount to understanding carbohydrate utilization and regulation of this process in Streptococcus mutans.

该提议的远距离目标是了解 分解代谢物抑制作用，影响并控制碳水化合物利用率 在链球菌Mu TAN中以及该过程如何参与 链球菌诱导的龋齿的发病机理。机制 在革兰氏阳性细菌和 阐明此过程将有可能更好地预防 龋齿。但是，如果没有这个远距离目标 首先对当地监管的基本了解 磷酸烯醇丙酮酸依赖性磷酸转移酶系统（PTS） S. mutans中的碳水化合物转运。 因此，该提议的短期目标是开始研究 进入S. mutans pts的调节。个体的特定目标 该提议包括：1。）在S.的大肠杆菌K12中进行隔离和克隆。 涉及各种运输和利用的Mutans基因簇 通过PTS碳水化合物； 2.）鉴定的基因产物 通过体外基因表达系统分离的PTS基因簇； 3.） 测定分离的PTS基因的核苷酸序列 确定整体操纵子结构的集群，以识别区域 对于体外产生的转录和翻译融合， 并确定推定的监管控制区； 4.）体外 产生转录融合以监测S的调节。 响应各种碳水化合物的突变PT； 5.）分析 分离的PTS基因簇的推定启动子区域和 识别和分析所涉及的推定控制区 使用翻译融合对这些操纵子进行调节；和6.）测定 链球菌的转录活性PTS PTS分解代谢酶基因 各种环境条件。 这些特定目标的完成将提供必要的 信息以启动远程目标的研究，将是 了解碳水化合物利用和调节的至高无上 链球菌突变的过程。