NUCLEOTIDE SEQUENCE AND CONTROL OF THE GLUCOSE DEHYDROGENASE OPERON

葡萄糖脱氢酶操纵子的核苷酸序列和控制

• 批准号: 3922582
• 负责人: E FREESE
• 金额: --
• 依托单位: NATIONAL INSTITUTE OF NEUROLOGICAL DISORDERS AND STROKE
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/3922582
• 关键词: Bacillus subtilis; Escherichia coli; bacterial genetics; bacterial proteins; cytogenetics; developmental genetics; gene expression; genetic manipulation; genetic mapping; genetic promoter element; genetic transcription; glucose dehydrogenases; nucleic acid sequence; operon; protein engineering; protein sequence; regulatory gene; site directed mutagenesis; sporogenesis; transposon /insertion element

During the development process leading to spore production in Bacillus subtilis, numerous proteins are synthesized. We have isolated a genetic region which codes for three protein that are synthesized only during differentiation and only in a particular forespore cell compartment. One of these genes codes for glucose dehydrogenase and is part of a bicistronic unit. We have determined the location of the transcription dehydrogenase and is part of a bicistronic unit. We have determined the location of the transcription initiation site as being 12 base pairs from the ribosomal binding site that precedes the first gene of the operon. We have attached beta-galactosidase to the promoter region as reporter gene and incorporated this whole DNA piece into the SP- beta region of the B. subtilis chromosome. Using specific deletion mutations we have then shown that the area from -10 to -35 suffices to control the expression of this operon during sporulation. Further mutations show that only the bases around -10 and -35 are important where those in the middle region could be all replaced by other bases without any effect. The promoter region looks like a typical receptor for a sigma factor and in fact a sigma G factor has been found whose deficiency leads to the absence of glucose dehydrogenase production. Other mutations were used to delete either the glucose dehydrogenase gene or one of the other two genes in order to determine their physiological importance. The gene included in the glucose dehydrogenase operon was thus shown to be essential for spore germination on the combination of glucose, fructose, asparagine, and potassium whereas germination on alanine was not affected. The significance of the other gene is still being studied.

在导致孢子产生的发育过程中， 枯草芽孢杆菌，合成许多蛋白质。 我们有 分离出了一个编码三种蛋白质的基因区域， 只在分化过程中合成， 前孢子细胞室 其中一个基因编码葡萄糖 脱氢酶，并且是双顺反子单元的一部分。 我们有 确定了转录脱氢酶的位置，并 双顺反子单元的一部分。 我们已经确定了 转录起始位点为12个碱基对， 位于操纵子第一个基因之前的核糖体结合位点。 我们将β-半乳糖苷酶连接到启动子区域， 报告基因，并将整个DNA片段纳入SP- B的β区。枯草染色体 使用特定删除 我们已经证明，从-10到-35的区域足以 来控制这种操纵子在孢子形成过程中的表达。 进一步的突变表明，只有-10和-35左右的碱基是 重要的是那些在中间区域的人可以全部被取代 其他基地没有任何影响。 启动子区域看起来像 一个典型的σ因子受体，事实上是σ G因子， 已经发现其缺乏导致葡萄糖的缺乏 脱氢酶生产。 其他突变用于删除 葡萄糖脱氢酶基因或其它两个基因之一 以确定它们的生理重要性。 基因 包括在葡萄糖脱氢酶操纵子中，因此显示 对孢子萌发至关重要的葡萄糖的结合， 果糖、天冬酰胺和钾，而在丙氨酸上萌发 没有受到影响。 另一个基因的意义仍然是 被研究。