Structural analysis of the genetic switch controlling expression of the AhdI restriction-modification system

在所有生物体中，基因表达的调控都依赖于基因调控蛋白与其DNA靶点之间复杂的相互作用。对这种机制的充分理解需要在生物物理和生化水平上进行分析。在这里，我们研究了参与细菌限制性修饰系统调节的遗传开关的分子基础。细菌中的限制性修饰系统是一种原始的“免疫系统”；它们识别外来的DNA序列，并能够摧毁入侵的DNA。然而，破坏DNA的酶基因的表达必须延迟，以便有时间让宿主DNA被标记为“自我”（通过一个称为甲基化的过程），从而保护宿主DNA不被切割。一种控制蛋白负责这种时间延迟，我们已经通过x射线衍射解决了它的分子结构。我们还确定，当蛋白质与目标DNA序列相互作用时，它会形成许多复合物。该项目的目的是发现与DNA结合的各种调节复合物的三维结构，并了解它们如何相互作用以允许相关基因的调节。总之，这些实验方法应该提供一个详细的基因开关的分子图像，其基础是限制修饰系统的建立和维持，其中的许多特征应该普遍适用于广泛的基因调控系统。

The regulation of gene expression in all organisms is dependent on complex interactions between gene-regulatory proteins and their DNA targets. A full understanding of such mechanisms requires analysis at the biophysical and biochemical level. Here, we investigate the molecular basis of the genetic switch involved in the regulation of bacterial restriction-modification systems. Restriction-modification systems in bacteria act as a form of primitive 'immune system'; they recognise foreign DNA sequences and are able to destroy invading DNA. However, expression of the gene for the enzyme that destroys the DNA must be delayed, to allow time for the host DNA to become labelled as 'self' (by a process called methylation) and thus protected from cleavage. A controller protein is responsible for this timing delay, and we have solved its molecular structure by X-ray diffraction. We have also established that the protein forms a number of complexes when it interacts with its target DNA sequence. The aim of this project is to discover the 3-dimensional structure of the various regulatory complexes bound to DNA, and to understand how they interact to allow regulation of the relevant genes. Together, these experimental approaches should provide a detailed molecular picture of the genetic switch underlying the establishment and maintenance of restriction-modification systems, many features of which should be generally applicable to a wide range of gene regulatory systems.