REGULATION OF ISOLEUCINE/VALINE BIOSYNTHESIS

我们感兴趣的是细胞协调 表达其生长所需的各种基因。人类的基因 异亮氨酸-缬氨酸生物合成分为五种转录 单位。多种分子信号作为表达的效应器 ILV基因。这些包括特定于ilv的调节器(例如 氨基酸最终产物和蛋白质)和一般或全球控制(如 作为ppGpp和蛋白质因子)。这些分子物种影响ilv基因。 通过不同的机制表达(即，转录启动与 衰减)。效应器的数量和光谱反映了这一点 异亮氨酸、亮氨酸和缬氨酸构成了高于平均百分比的 蛋白质中存在的氨基酸。因此，它们的合成代表了一个 细胞资源的主要承诺，细胞必须能够 以最佳方式调节这些基因的表达，以响应 另一种生理状态。 五个基因结合形成ilvGMEDA操纵子，它包含多个 以达到其基因所需的平衡 产品。调节的主要部位是启动子-衰减器。 IvG近端。操作子内的辅助站点包括：两个内部站点 发起人和内部终止站点。反式效应器(ppGpp和 蛋白质)似乎调节5‘启动子的表达。 为了研究操纵子调控中涉及的因素，我们 将使用我们构建的ilv-Galk融合。这些质粒 将使用各种技术进行研究。这将包括：(1) 寡核苷酸诱变；(2)蛋白质相互作用的体内分析； (3)RNAP相互作用的体外分析。此外，我们还定义了一个 启动子上游的位置，这是最大限度表达所必需的 并建议分离作用于ILV启动子的蛋白(S) 这个网站。 因为一组基因表达的协调是必不可少的 对于细胞功能，我们认为重要的是确定 实现这种协调的机制。人类的基因 异亮氨酸/缬氨酸的生物合成形成了这样一组。因为这些生物的DNA 基因已经被分离和鉴定，它们很容易被访问 详细分析了他们的监管。

We are interested in the mechanism by which a cell coordinates the expression of the various genes required for its growth. The genes for isoleucine valine biosynthesis are separated into five transcriptional units. A variety of molecular signals act as effectors of the expression of the ilv genes. These include both ilv specific regulators (such as the amino acid end products and proteins) and general or global controls (such as ppGpp and protein factors). These molecular species affect ilv gene expression by different mechanisms (ie, transcription initiation vs. attenuation). The number and spectrum of effectors reflects that isoleucine, leucine, and valine constitute an above average percentage of the amino acids present in protein. Thus, their synthesis represents a major commitment of cellular resources and it is essential the cell be able to optimally modulate the expression of these genes in response to alternative physiological conditions. Five the genes combine to form the ilvGMEDA operon, which contains multiple regulatory sites so as to achieve the requisite balance of its gene products. The primary site of regulation is the promoter-attenuator proximal to ilvG. Secondary sites within the operon include: two internal promoters and internal termination sites. The trans effectors (ppGpp and proteins) appear to regulate expression at the 5' promoter. In order to study the factors involved in the regulation of the operon, we will employ the ilv-galK fusions which we have constructed. These plasmids will be studied using a variety of techniques. This will include: (1) oligonucleotide mutagenesis; (2) in vivo analysis of proein interactions; and (3) in vitro analysis of RNAP interactions. Also, we have defined a site upstream of the promoter, which is required for maximal expression from the ilv promoter and propose to isolate the protein(s) that act at this site. Because the coordination of the expression of sets of genes is essential for cellular function, we believe it is important to determine the mechanisms by which this coordination is achieved. The genes for isoleucine/valine biosynthesis form such a set. Because the DNA of these genes has been isolated and characterized, they are readily accessible for detailed analysis of their regulation.