REGULATION OF GLUTAMINE SYNTHETASE IN STREPTOMYCES

链霉菌中谷氨酰胺合成酶的调控

• 批准号: 3135064
• 负责人: SUSAN H. FISHER
• 金额: $ 17.42万
• 依托单位: BOSTON UNIVERSITY MEDICAL CAMPUS
• 依托单位国家: 美国
• 财政年份: 1987
• 资助国家: 美国
• 起止时间: 1987-12-01 至 1994-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3135064
• 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; protein sequence; sporogenesis; structural genes

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.

链霉菌合成了70多种药用物质 抗菌药物以及抗肿瘤和抗寄生虫剂。这些 化合物是在营养过程中由初级代谢物合成的 限制生长和启动产孢子。令人惊讶的是， 人们对初级新陈代谢或后遗症知之甚少 在这些细菌中引发抗生素的产生和产孢量。在……里面 为了增加我们对链霉菌基本生理学的了解， 氨化关键酶谷氨酰胺合成酶的调节 同化作用，正在天蓝色链霉菌中进行研究。对克隆人的分析 1.Coelicolor GS结构基因(Sm)显示SM的转录 启动子在营养生长过程中受氮素调节。两种方法 被用来确定调节这一规定的因素。 第一，调节和表达基因所需的核苷酸区域 通过构建glnA-XylE鉴定Lm原声 转录融合和分离改变表达的突变体 以及对glnA-XylE融合的调控。第二，glnA监管 因子正在通过分离突变株来进行遗传鉴定 GlnA启动子表达的改变。需要谷氨酰胺的类别 GlnR突变体不能转录氮调节的glnA 推动者。该突变体中的缺陷基因产物已被克隆 互补作用，并正在被鉴定为亚克隆以促进DNA 测序。由于氮素调节酶尿素酶的表达是 同样缺乏这些突变体，全球氮素调控系统可能 出现在.1中。天蓝色。克隆的W DNA将用于研究 &W启动子与基因鉴定的相互作用 GlnA调节蛋白、RNA聚合酶和含氮小分子 代谢物。5.对天蓝色鱼氮代谢的认识 应该通过遗传促进抗生素产量的增加 操纵初级代谢物池，或通过选择 适量生长铍。此外，由于初级代谢 对链霉菌的抗生素生产和产孢量的影响， 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。