Structure and Relations of Proteins and Nucleic Acids

在本更新申请中，我们描述了继续并完成几项正在进行的研究的计划， E.大肠杆菌和噬菌体T4的复制复合物。两 代表了基因表达的等效“大分子机器”的最简单版本， 在高等生物中控制这些过程的大多数相同的成分。我们将特别关注 中心聚合酶如何在模板指导的合成中进行单核苷酸添加循环，以及 如何控制这一过程，并将其重定向到替代的和不太可能的动力学竞争反应中 途径（包括终止和编辑）在暂停网站，在终止网站，并作为一个结果， 残基错误掺入。我们还将关注这些“初级”反应途径是如何被进一步调节的 通过调节因子，如抗终止复合物。为支持这些总体目标，我们的具体 下一个报告所述期间的目标是进一步审查下列问题： 利用新的光谱方法研究局部RNA和DMA构象和动力学;（ii） 进一步研究转录和复制中单核苷酸添加循环的步骤;（iii） 继续研究转录过程中反应途径选择的控制;（iv）进一步了解 抗终止系统的基本控制机制;（v）完成我们的RNA和蛋白质模型研究 链循环和动力学;（vi）研究E.大肠杆菌转录 终止因子Rho与RNA聚合酶;（vii）继续我们的解旋酶机制的研究， （viii）进一步研究持续合成钳的机制 装载在噬菌体T4系统;和（ix）继续我们的蛋白质-核酸相互作用的理论研究 动力学这些研究应该有助于我们更好地了解这些复合物是如何组装的，以及它们是如何在体内形成的。 这些组分相互作用以构建稳定且易于调节的“大分子机器”。就其 生物医学研究的意义，这些研究将继续作为模型的功能， 控制类似的转录和复制系统的高等生物，并可能有助于揭示如何 这些复合物在癌症和其它不适当基因表达的疾病中会出错。

In this renewal application we describe plans to continue and complete several ongoing studies on the mechanisms and control of the transcription complex of E. coli and the replication complex of phage T4. Both represent the simplest versions of the equivalent 'macromolecular machines' of gene expression that use most of the same components to control these processes in higher organisms. We will focus, in particular, on how the central polymerases perform the single nucleotide addition cycle in template-directed synthesis, and how this process is controlled and redirected into alternative and less probable kinetically competing reaction pathways (including termination and editing) at pause sites, at termination sites, and as a consequence of residue misincorporation. We will also focus on how these 'primary' reaction pathways are further modulated by regulatory factors, such as antitermination complexes. In support of these overall objectives, our Specific Aims during the next reporting period will be to further examine the following issues: (i) to develop and exploit a new spectroscopic method for studying local RNA and DMAconformations and dynamics; (ii) to further examine the steps in the single nucleotide addition cycle in transcription and replication; (iii) to continue to study the control of reaction pathway selection in transcription; (iv) to further understand the basic control mechanisms of antitermination systems; (v) to complete our model studies of RNA and protein chain looping and dynamics; (vi) to examine the mechanistic details of the interactions of E. coli transcription termination factor Rho with RNA polymerase; (vii) to continue our studies of helicase mechanisms and coupling in transcription and replication; (viii) to further examine the mechanisms of processivity clamp loading in the phage T4 system; and (ix) to continue our theoretical studies of protein-nucleic acid interaction kinetics. These studies should help us to better understand how these complexes assemble, and how the components interact to build stable, and yet easily regulatable 'macromolecular machines'. In terms of their significance for biomedical research, these studies will continue to serve as models for the function and control of the analogous transcription and replication systems of higher organisms, and may help reveal how these complexes can go awry in cancer and other diseases of inappropriate gene expression.