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REGULATION OF GLUTAMINE SYNTHETASE IN STREPTOMYCES

链霉菌中谷氨酰胺合成酶的调控

基本信息

批准号：
2062087
负责人：
SUSAN H. FISHER
金额：
$ 18.58万
依托单位：
BOSTON UNIVERSITY MEDICAL CAMPUS
依托单位国家：
美国
项目类别：
财政年份：
1987
资助国家：
美国
起止时间：
1987-12-01 至 1995-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/2062087
关键词：
Streptomyces antibiotics bacterial genetics biotechnology deficient growth media gene expression genetic promoter element genetic transcription glutamate ammonia ligase gram positive bacteria microorganism metabolism molecular cloning mutant nitrogen metabolism nucleic acid sequence protein sequence sporogenesis structural genes

项目摘要

Streptomyces species synthesize more than 70 medically useful antibioties as well as antitumor and antiparasitic agents. These compounds are synthesized from primary metabolites during nutritional limitation of growth and the initiation of sporulation. Surprisingly, very little is known about primary metabolism or the sechanisms that trigger antibiotic production and sporulation in these bacteria. In order to increase our understanding of Streptomyces basic physiology, the regulation of glutamine synthetase (GS), a key enzyme in ammoniin assimilation, is being studied in S. coelicolor. Analysis of the cloned I. coelicolor GS structural gene (sm) shows that transcription of the SM promoter is nitrogen regulated during vegetative growth. Two approaches are being used to identify tbe factors that mediate this regulation. First, nucleotide regions required for the regulation and expression of the LM pronoter are being identified by constructing glnA-xylE transcriptional fusions, and isolating nutations that alter expression and regulation of the glnA-xylE fusions. Secondly, glnA regulatory factors are being genetically identified by isolating mutants with altered expression of the glnA promoters. The glutamine-requiring ClassI GlnR mutants are unable to transcribe the nitrogen-regulated glnA promoter. The defective gene product in this nutant has been cloned by complementation and is being identified subcloned to facilitate DNA sequencing. Since expression of the nitrogen-regulated enzyme urease is also deficient in these mutants, a global nitrogen regulatory system may be present in .1. coelicolor. The cloned &W DNA will be used to study interactions between the &W promoters and the genetically identified glnA regulatory proteins, RNA polymerase and small nitrogencontaining metabolites. An understanding of nitrogen metabolism in 5. coelicolor should facilitate increased antibiotic production by genetic manipulation of primary uetabolite pools, or by choice of the appropriate growth nedium. Furthermore, since primary metabolism impinges on both antibiotic production and sporulation in Streptomyces, these studies may help define mechanisms that regulate these processes in Streptomyces.

链霉菌属物种合成70多种医学上有用的以及抗肿瘤和抗寄生虫剂。这些化合物是由营养过程中的初级代谢产物合成的。限制生长和孢子形成的起始。令人惊奇的是，我们对初级代谢或引发这些细菌产生抗生素和孢子。在为了增加我们对链霉菌基本生理学的了解，氨合成关键酶谷氨酰胺合成酶（GS）调控同化，正在研究在S。天蓝色。克隆的分析 I.腔棘鱼GS结构基因（sm）的转录表明，启动子在营养生长过程中受到氮的调控。两种方法被用来确定调节这种调节的因素。首先，调节和表达所需的核苷酸区域，通过构建glnA-xylE，转录融合和分离改变表达的章动和glnA-xylE融合的调节。其次，glnA调节通过分离突变体， glnA启动子的表达改变。需要谷氨酰胺的I类 GlnR突变体不能转录氮调节的glnA 启动子该突变体中的缺陷基因产物已被克隆，互补，并正在确定亚克隆，以促进DNA 测序由于氮调节酶尿素酶的表达，也缺乏这些突变体，一个全球性的氮调节系统，出现在.1中。天蓝色。克隆的&W DNA将用于研究 &W启动子和基因鉴定的 glnA调节蛋白，RNA聚合酶和小氮代谢物。了解氮代谢5. coelicolor 通过基因工程，操纵初级代谢物池，或通过选择适宜的生长介质。此外，由于初级代谢影响链霉菌的抗生素生产和孢子形成，这些研究可能有助于确定调节这些过程的机制，在链霉菌中。