GENETIC CONTROL OF BIOSYNTHESIS IN BACILLUS SUBTILIS

枯草芽孢杆菌生物合成的遗传控制

• 批准号: 3282512
• 负责人: STANLEY A ZAHLER
• 金额: $ 8.55万
• 依托单位: CORNELL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1984
• 资助国家: 美国
• 起止时间: 1984-04-01 至 1989-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3282512
• 关键词: Bacillus subtilis; Escherichia coli; alleles; aminoacid biosynthesis; bacterial genetics; bacteriophage lambda; branched chain aminoacid; chromosome translocation; enzyme induction /repression; fungal genetics; gel electrophoresis; gene complementation; gene expression; genes; genetic manipulation; genetic mapping; genetic strain; genetic transcription; gram positive bacteria; hydro lyase; isoleucine; leucine; microorganism metabolism; molecular cloning; nucleic acid sequence; operon; valine

The genetic controls that operate in biosynthetic pathways in Gram-positive bacteria have not previously been amenable to analysis. To a considerable extent, the difficulties that have hindered such studies have now been overcome. We have methods for studying cis-trans relationships between alleles, for determining dominance and recessiveness, for inserting transposons into and near to genes, and for cloning genes on a variety of multicopy vectors. We now plan to use these new techniques to study the synthesis of the branched-chain amino acids isoleucine, valine and leucine by Bacillus subtilis, using a combination of genetics, biochemistry and molecular biology. In the early stages of the work we will study the controls involved in the expression of the ilvBC-leu operon, encoding the second and third enzymes of the isoleucine-valine pathway and the four cistrons responsible for the first three steps of the leucine pathway. This operon differs from known biosynthetic operons in Escherichia coli and other bacteria, since it combines genes for enzymes of two separate (but related) pathways. We will clone and sequence the region near the beginning of the ilvB gene to learn if the region contains recognizable sequences suitable for transcription controls, and if it contains binding sites for trans-acting control proteins that are involved in the expression of the operon. We will also examine the region between the ilvC and leuA cistrons, to learn if B. subtilis has a way to control the leucine pathway without affecting the isoleucine-valine pathway. Other genes of the pathways will also be studied. The ilvA gene (threonine deaminase) has already been examined by us. We have shown that the production of that enzyme is constitutive in B. subtilis strain 168, and under isoleucine control in other strains, e.g., strain W23. We will determine the differences between the genes from the two strains to learn how ilvA is controlled in strain W23. Later we will examine the ilvD gene (dihydroxyacid dehydratase) to learn if it is coordinately controlled with ilvA. It is our hope that we will uncover new methods of byosynthetic control used by Gram-positive bacteria and not by Gram-negative ones.

革兰氏阳性菌生物合成途径中的遗传控制 细菌以前不适于分析。 在相当 在某种程度上，阻碍这种研究的困难现在已经被 克服 我们有研究顺反关系的方法， 等位基因，决定显性和隐性，插入 转座子进入和接近基因，并用于克隆基因上的各种 多拷贝向量 我们现在计划使用这些新技术来研究 支链氨基酸异亮氨酸、缬氨酸和亮氨酸的合成 枯草芽孢杆菌，使用遗传学，生物化学和 分子生物学 在工作的早期阶段，我们将研究 参与ilvBC-leu操纵子表达的控制，其编码 异亮氨酸-缬氨酸途径的第二和第三种酶以及 顺反子负责亮氨酸途径的前三步。 该操纵子不同于大肠杆菌中已知的生物合成操纵子， 其他细菌，因为它结合了两种不同的酶的基因（但 相关的路径。 我们将克隆和测序的区域附近 ilvB基因的开始，以了解该区域是否含有可识别的 适合于转录控制的序列，并且如果它含有结合 参与表达的反式作用控制蛋白的位点 的操纵子。 我们还将研究ilvC和leuA之间的区域 顺反子，学习如果B。枯草杆菌有一种方法来控制亮氨酸途径 而不影响异亮氨酸-缬氨酸途径。 还将研究途径的其他基因。 ilvA基因（苏氨酸 脱氨酶）已经被我们检查过了。 我们已经证明， 该酶产生在B中是组成型的。枯草杆菌菌株168，和 在其它菌株中在异亮氨酸控制下，例如，菌株W23。 我们将 确定两种菌株基因之间的差异， ilvA在菌株W23中是如何控制的。 稍后我们将研究ilvD基因 （二羟酸脱氢酶），以了解它是否与 ilvA. 我们希望我们能发现新的生物合成方法， 革兰氏阳性菌使用对照，革兰氏阴性菌不使用对照。