MOLECULAR BASIS AND ENZYMOLOGY OF MICROBIAL BIOSYNTHESIS

磷酸戊糖途径是一种重要的代谢途径 用于几乎所有生物的新陈代谢。增加了 两种独特的酶，核酮糖二磷酸羧基酶/加氧酶(Rubisco) 和磷酸核苷酸激酶(PRK)，使这一途径在 纯生物合成模式，使有机体获得利用 二氧化碳是碳的唯一来源。在这种情况下， 该途径的其他酶，包括在 原核生物和真核生物都是生物合成酶，在 相反的方向形成了它们在体内的通常作用。 我们实验室最近的研究表明，结构基因 上述酶，以及编码果糖二磷酸酶的基因 甘油醛磷酸脱氢酶(FBPase)和转酮醇酶 在兼性厌氧菌的两个不同的染色体操纵子中发现 紫色非硫细菌球形红杆菌。都是重组的 在大肠杆菌中的蛋白，以及许多重组蛋白 纯净到同质。因此，这个项目的主要目标之一是 研究PrK、FBPase和FBPase的结构-功能关系 转酮醇酶，应用蛋白质化学和分子技术 生物学上，这种双重方法第一次变得可行 这些重要的蛋白质。 这个项目的第二个主要推动力将是研究分子 对这两个操纵子进行不同调控的基础。控制是 在转录和转录后水平上都有表现。 由于暴露在碳和氧水平的增加或减少中 对基因表达有重要影响，我们将探讨其机制。 控制及其与已知的细胞内调节系统的关系 调控机制及其与细胞内调控系统的关系 已知对外界刺激有反应。这项研究也关注于 关于多顺反子转录本的核溶解处理的作用和 它与差异基因表达的关系，一个重要的关键因素 调控原核和真核基因的表达。 因此，这些研究提供了一个极好的机会，可以将控制与 在酶和分子水平上，提高我们的总体水平 对中间代谢的理解。

The pentose phosphate pathway is an important metabolic scheme required for the metabolism of virtually all living organisms. The addition of two unique enzymes, ribulose bisphosphate carboxylase/oxygenase (RubisCO) and phosphoribulokinase (PRK), allows this pathway to function in a purely biosynthetic mode, such that organisms gain the capacity to use carbon dioxide as the sole source of carbon. Under these conditions, other enzymes of the pathway, including ubiquitous catalysts found in both prokaryotes and eukaryotes, function as biosynthetic enzymes, in the opposite direction form their usual role in vivo. Recent work from our laboratory has shown that the structural genes for the above enzymes, along with genes encoding fructose bisphosphatase (FBPase), glyceraldehyde phosphate dehydrogenase, and transketolase, are found in two distinct chromosomal operons in the facultatively anaerobic purple nonsulfur bacterium Rhodobacter sphaeroides. Both recombinant proteins in Escherichia coli, and many of the recombinant proteins purified to homogeneity. Thus, one of the major goals of this project is to study structure-function relationships of PRK, FBPase and transketolase, using the techniques of protein chemistry and molecular biology, the first time that this dual approach has become feasible for these important proteins. The second major thrust of this project will be to study the molecular basis for differential regulation of the two operons. Control is manifested at both the transcriptional and posttranscriptional levels. Since exposure to increasing or decreasing levels of carbon and oxygen has a major effect on gene expression, we will probe the mechanism of control and its relation to intracellular regulatory systems known to be mechanism of control and its relation to intracellular regulatory systems known to be responsive to external stimuli. This research also focuses on the role of nucleolytic processing of polycistronic transcripts and its relation to differential gene expression, a key factor important for regulating expression of both prokaryotic and eukaryotic genes. These studies thus present an excellent opportunity to relate control at both the enzymic and molecular levels, to increase our general understanding of intermediary metabolism.