ACETOIN PRODUCTION BY BACILLUS SUBTILIS

枯草芽孢杆菌生产乙偶姻

• 批准号: 3303126
• 负责人: STANLEY A ZAHLER
• 金额: $ 13.46万
• 依托单位: CORNELL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1990
• 资助国家: 美国
• 起止时间: 1990-04-01 至 1993-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3303126
• 关键词: Bacillus subtilis; DNA binding protein; bacterial genetics; bacterial proteins; carboxylation; cell growth regulation; gene expression; genetic transcription; gram positive bacteria; growth media; microorganism culture; microorganism growth; microorganism metabolism; molecular cloning; nucleic acid sequence; oxoacid lyase; polymerase chain reaction; protein biosynthesis; structural genes; transcription factor

We want to learn how Gram-positive bacteria control their synthesis of secondary metabolites -- substances that they produce only after vegetative growth and cell division have stopped. Such products include antibiotics, toxins, a number of enzymes, and a variety of small molecules. We will use as a model system the production of acetoin (2-oxo, 3-hydroxy-butane) by Bacillus subtilis. This compound is excreted by stationary-phase cells in large quantities. It comes from the decarboxylation of acetolactate, which the bacteria products by means of an enzyme, acetolactate synthase (AlsS), that is synthesized only after growth has stopped. We have cloned and sequenced the alsS gene that encodes AlsS, and an adjacent gene orfX that is transcribed and translated as a divergent operon. AlsA has sequence homology with IlvB, a biosynthetic enzyme of B. subtilis required for the synthesis of branched-chain amino acids. OrfX has sequence homology with a family of positive regulatory proteins found in other bacteria. We want to learn how the alsS gene's expression is controlled so that its product is not detectable during vegetative growth, but is produced once growth stops. We suspect that the orfX gene is involved in this control, but do not have direct evidence for this. A mutation, alsAl, unlinked to alsS, prevents the expression of alsS under all conditions. We have cloned alsA and will determine the role it plays in controlling the expression of alsS.

我们想知道革兰氏阳性菌是如何控制它们的 次级代谢物--它们只有在营养生长后才产生的物质 生长和细胞分裂已经停止。 这些产品包括抗生素， 毒素，一些酶和各种小分子。 我们将使用 作为模型系统，通过以下方法生产乙偶姻（2-氧代，3-羟基-丁烷）： 枯草芽孢杆菌。 该化合物由静止期细胞分泌， 是介于 它来自于乙酰乳酸的脱羧作用， 细菌产物通过酶，乙酰乳酸合酶（AlsS）， 只有在停止生长后才能合成。 我们已经克隆和 对编码AlsS的alsS基因和一个邻近的基因orfX进行测序， 被转录和翻译为一个分歧操纵子。 AlsA具有序列 与B的生物合成酶IlvB同源。所需的枯草芽孢杆菌 支链氨基酸的合成。 OrfX与一种 在其他细菌中发现的阳性调节蛋白家族。 我们想 了解alsS基因的表达是如何控制的， 在营养生长期间检测不到，但一旦生长停止就会产生。 我们怀疑orfX基因参与了这种控制，但没有证据表明， 直接证据。 与alsS无关的突变alsAl阻止了 所有条件下的alsS表达。 我们克隆了alsA和will 确定它在控制alsS表达中所起的作用。