Allosteric Effects of DNA on Bacteriophage 434 Repressor Function

与蛋白质结合的DNA序列可以深刻地影响蛋白质-DNA复合物参与基因调控过程的能力。因此，DNA不仅仅是蛋白质组装的被动平台，它可以作为蛋白质功能的变构效应物发挥积极的第二作用。DNA的变构效应是最近发现的一种现象。因此，理解DNA如何影响转录调节蛋白的形式和功能的目标代表了DNA结合蛋白功能研究的前沿。 噬菌体434的阻遏物似乎经历了两种构象变化，一种是在非特异性DNA存在下二聚化，另一种是阻遏物二聚体与特异性DNA结合位点结合。434阻遏物-DNA复合物的结构和功能可以随其结合的DNA序列的特性而变化。在其他变化中，这些不同的结构形式在它们形成更高级低聚复合物的能力方面不同。这些复合物之间的构象差异影响：1）阻遏物的序列识别; 2）DNA结合阻遏物之间的协同相互作用; 3）阻遏物与RecA的相互作用。 根据这些发现，将确定DNA对阻遏物功能的变构效应的基础。在确定了DNA变构的本质之后，这些信息将被用于探测DNA对阻遏物四聚体形成的变构效应，以提供对阻遏物的DNA依赖性合作结合机制的深入了解。此外，这些信息将指导实验，旨在提供一个了解如何分布的各种寡聚体形式的阻遏物可以通过与其他蛋白质和DNA在宿主细胞中的相互作用进行调节。从这些结果中获得的见解将进一步了解这些和其他序列特异性蛋白质-DNA复合物中功能的关键决定因素。 除了作为一个原型系统，其中的DNA序列对功能的影响可以进行调查，噬菌体提供了一个概念上和实际上可访问的系统，在其中研究蛋白质-蛋白质相互作用在基因调控中的作用。这些特点使各级学生，包括高中学生，能够为这一系统的观察和理解作出重大贡献。通过在这个系统中工作，研究生、本科生和高中生已经学会了如何提出重要的生物学问题，并利用他们的观察为理解基因调控机制做出了重要贡献。该系统的优雅简单性确保了其他人也会这样做。

The DNA sequence to which a protein binds can profoundly affect the ability of the protein-DNA complex to participate in gene regulatory processes. Hence, DNA is more than a passive platform onto which proteins assemble, it can play an active, second role as an allosteric effector of protein function. The allosteric effect of DNA is a recently discovered phenomenon. Thus, the goal of understanding how DNA influences the form and function of the transcriptional regulatory protein represents the leading edge of the work on the function of DNA binding proteins. The repressor of bacteriophage 434 appears to undergo two conformational changes, one as it dimerizes in the presence of nonspecific DNA and another as the repressor dimer binds to a specific DNA binding site. The structure and function of the 434 repressor-DNA complex may vary with the identity of the DNA sequence to which it binds. Among other changes these distinct structural forms vary in their abilities to form higher order oligomeric complexes. The conformational differences between these complexes influence 1) sequence recognition by the repressors; 2) cooperative interactions between DNA bound repressors and 3) the interaction of the repressor with RecA. In light of these findings, the basis for the allosteric effect of DNA on repressor function will be determined. Having established the nature of DNA allostery, this information will then be used to probe the allosteric effect of DNA on repressor tetramer formation to provide insight into the mechanism of DNA-dependent cooperative binding of repressor. In addition, this information will guide experiments aimed at providing an understanding of how the distribution of the various oligomeric forms of repressor can be regulated by interaction with other proteins and DNA in the host cell. Insights gained from these results will further the understanding of the critical determinants of function in these and other sequence-specific protein-DNA complexes. In addition to being an archetypal system in which the effect of DNA sequence on function can be investigated, the lambdoid bacteriophages provide a conceptually and practically accessible system in which to study the role of protein-protein interactions in gene regulation. These features have allowed students at all levels, including those at the high school level, to significantly contribute to the battery of observations and understanding in this system. By working in this system, students at the graduate, undergraduate and high school levels, have learned how to ask important biological questions, and used their observations to make important contributions to the understanding of gene regulatory mechanisms. The elegant simplicity of the system ensures that others will do the same.