BRANCHED CHAIN AMINO ACID BIOSYNTHESIS IN E COLI

描述：该建议涉及转录的机制， 激活的非特异性（组蛋白样）DNA结合蛋白，称为 整合宿主因子（IHF），最初发现并命名基于其 对噬菌体λ位点特异性整合的影响 也是许多操纵子表达的一般调节因子。它是基于 在PI最近的证据表明，IHF激活了 通过一种新的用于支链氨基酸生物合成的ilvGMEDA操纵子 基因激活机制他已经证明了ilvG的激活 启动子需要IHF与上游激活位点（UAS 1）结合， 称为SIDD的单独上游位点和非锥形-10区域。他 还表明，IHF结合位点激活转录时， 在螺旋的任何一面，因此不太可能与IHF相互作用 与RNAP或不同的蛋白质因子。他进一步证实了这一点， IHF结合位点被一个或多个氨基酸的结合位点取代， 异源DNA结合和弯曲蛋白，淋巴增强剂- 结合因子LEF-1激活E. coliilvG启动子。结合和 在这些条件下的转录激活伴随着 在-10启动子区域中两个A-T碱基对的解旋。这些和 其他的实验使他提出了一个“DNA结构传递 IHF调控ilvG启动子的“模型”。其提出之新订的 实验的目的是测试一些预测， 模型

DESCRIPTION: This proposal concerns the mechanism of transcriptional activation by a non-specific (histone-like) DNA binding protein called integration host factor (IHF), originally found and named based on its effect on site-specific integration of phage lambda and since shown to act also as a general regulator of expression of numerous operons. It is based upon the P.I.'s recent evidence that IHF activated the ilvG promotor of the ilvGMEDA operon for branched chain amino acid biosynthesis by a novel gene activation mechanism. He already showed that activation of the ilvG promoter requires binding of IHF to an upstream activation site (UAS1), a separate upstream site called SIDD, and a non cononical -10 region. He also showed that the IHF-binding site activated transcription when located on either face of the helix, thus making it unlikely that IHF interacts with RNAP or a different protein factor. He further substantiated this by substitution of the IHF binding site with the binding site of a heterologous DNA binding and bending protein, the lymphoid enhancer- binding factor LEF-1 activates the E. coli ilvG promoter. Binding and transcriptional activation under these conditions is accompanied by an unwinding of two A-T base pairs in the -10 promoter region. These and other experiments has led him to propose a "DNA structural-transmission model" for regulation of the ilvG promoter by IHF. His proposed new experiments are aimed towards testing a number of predictions of this model.