Pushing proteins off DNA - how do helicases unwind protein-coated DNA?

DNA编码了作为生命蓝图所需的所有遗传信息。然而，这一蓝图最终需要转换为细胞的构建块。然而，DNA存储的信息深埋在这种分子结构中，只有通过剥离组成DNA的两条链，即所谓的解离，才能获得这些信息。这一关键功能是由一种名为解旋酶的酶执行的，这种酶是一种微小的纳米马达，沿着DNA棘轮移动，在前进的过程中将两条DNA链分开。所有生物体都有各种不同的解旋酶，仅一种解旋酶的缺陷就会导致细胞的灾难，潜在地导致致命性或基因内的衰弱突变。解旋酶在维持所有细胞的活力和将基因蓝图从一代传给下一代的核心重要性几十年来一直是已知的。最近，我们也逐渐认识到，解旋酶在试图解开DNA时面临着一个特殊的问题。DNA被包裹在细胞内的各种不同的蛋白质中。这些蛋白质在包装DNA、阅读遗传蓝图和协调染色体在细胞内的运动方面发挥着重要作用。不幸的是，我们现在知道，这些结合的蛋白质也给解旋酶带来了问题，因为任何与DNA结合的蛋白质也必须被推到一边，以允许解旋酶分离两条DNA链。然而，我们对解旋酶如何取代与核酸结合的蛋白质知之甚少。我们一直在研究一种名为Rep的解旋酶，它通过取代与DNA结合的蛋白质在复制遗传蓝图中发挥重要作用。我们的初步工作发现，移除部分解旋酶可以激活DNA解旋，但同时也会抑制DNA中的蛋白质置换。这一发现很重要，因为它表明，蛋白质从DNA中置换出来，肯定不只是解旋酶沿着DNA棘轮移动。因此，这种解旋酶进化出了帮助蛋白质脱离DNA的特定功能，尽管目前我们还不清楚这些功能是什么。我们的目标是通过使用不同的分子工具的组合来研究Rep和这种解旋酶的版本的性质，从而研究这种解旋酶是如何将蛋白质从DNA中置换出来的，这些解旋酶的版本增加或降低了将蛋白质从DNA中推离的能力。这项工作将揭示这类重要的酶是如何处理包裹DNA的大量蛋白质的。这是所有有机体都必须面对的问题，因此我们的发现将帮助我们了解DNA是如何在细胞内有效保持的，同样重要的是，事情可能会出现什么问题。解旋酶的错误可能导致遗传密码内非常有害的重排，导致遗传病，因此我们拟议的工作将揭示遗传密码腐败的潜在来源。相反，这项工作也可能揭示故意抑制解旋酶的新方法。由于解旋酶对生存是如此重要，这些抑制剂具有潜在的抗病毒、抗菌和抗癌化合物的用途。

DNA encodes all the genetic information needed to act as a blueprint for life. This blueprint, though, needs to be converted ultimately into the building blocks of a cell. However, the information that DNA stores is buried deep within the structure of this molecule and can only be accessed by stripping apart the two strands that constitute DNA, so-called unwinding. This critical function is performed by enzymes called helicases that are tiny nanomotors that ratchet along DNA, separating the two DNA strands as they go. All organisms have a variety of different helicases and defects in just one type of helicase can result in catastrophe for a cell, potentially causing lethality or debilitating mutations within genes.The central importance of helicases in maintaining the viability of all cells and in passing the genetic blueprint from one generation to the next has been known for decades. Recently we have also come to appreciate that helicases face a particular problem when attempting to unwind DNA. DNA is coated in a wide variety of different proteins inside cells. These proteins play important roles in packaging DNA, reading the genetic blueprint and coordinating the movement of chromosomes inside cells. Unfortunately we now know that these bound proteins also present problems to helicases since any proteins bound to the DNA must also be pushed off to allow separation of the two DNA strands by a helicase. However, we know very little about how helicases displace proteins bound to the nucleic acid.We have been studying a helicase called Rep that plays an important role in copying of the genetic blueprint by displacing proteins that are bound to DNA. Our preliminary work has discovered that removal of part of this helicase activates DNA unwinding but at the same time inhibits displacement of proteins from the DNA. This discovery is important because it shows that displacement of proteins from DNA must involve something more than the helicase merely ratcheting along the DNA. This helicase has therefore evolved specific features to help push proteins off DNA although currently we do not understand what these features are. We aim to investigate how this helicase displaces proteins from DNA by using a combination of different molecular tools to investigate the properties of Rep and versions of this helicase that have increased or decreased abilities to push proteins off DNA. This work will cast light on how this important class of enzyme deals with the vast array of proteins that coat DNA. This problem is one that all organisms must face and so our findings will help us to understand how DNA is maintained effectively inside cells and, just as importantly, how things might go wrong. Mistakes made by helicases can result in very harmful rearrangements within the genetic code, contributing to genetic disease, and so our proposed work will shed light on potential sources of corruption of the genetic code. Conversely, this work may also reveal new ways of deliberately inhibiting helicases. Such inhibitors have potential uses as antiviral, antibacterial and anticancer compounds since helicases are so important for survival.