Opening of a double stranded DNA replication fork by a hexameric helicase

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The inner workings of mammalian cells involve a number of interrelated, small, "nanomachines". Individual machines are usually composed of a number of proteins that work together to perform one of the cell's functions in life, and determine its ability to survive and replicate. Understanding how individual machines function is crucial because when these machines breakdown the consequences can be catastrophic. For example, some cancers can be attributed directly to the failure of machines that process the genetic information (DNA) of a cell. In addition, invading microorganisms such as viruses bring their own machines that subvert or destroy a cell as it is hijacked for the pathogen's own advantage. The identification and understanding of how these machines function can therefore be a starting point for combating such disease states. The biosciences can inspire nanotechnology, which aims to develop small mechanical devices based on protein machines that can be harnessed to drive or modulate molecular processes from which we benefit. Significant examples are found in biotechnology, such as emerging nanopore DNA sequencing devices that can operate at the single molecule level. The precise understanding of cellular nanomachines at the atomic level is absolutely dependent on the availability of precise structural information. For large molecular assemblies this information comes from X-ray structural analysis and, for very large assemblies, it is vastly assisted by powerful electron microscopes that can image single particles. Alone, electron microscopy can guide our understanding, but for the most part lacks the resolving power to provide critical detail. E1, the protein that we will study, is from a large group of pathogens, the papillomaviruses that cause warts and cancer in animals and man. Papillomavirus-derived diseases are of significant health and economic importance. E1 is often referred to as a "motor protein" that forms a six membered (hexameric) ring-like assembly, a machine that unwinds the two strands of the DNA molecule during replication (reproduction) of viral DNA. It can be viewed as a small radial engine that can move along DNA and separate its two strands and as such is a prototypic nanomachine and important target for drugs that could inhibit viral replication and disease.Here we aim to understand how the E1 machine unwinds DNA. The objectives are to carry out detailed X-ray structural studies on E1 to determine its overall structure and how it interacts with DNA. We intend to verify these results and test models of function with complementary biochemical and biophysical studies. As a leading tractable model, it has great promise for being the first hexameric DNA-unwinding machine that we fully understand. The potential applications of our work include formulating strategies to target E1 and related helicases through structure based drug design. They will help understand replication processes in our own cells that are more complex and challenging to define. There are potential applications in nanobiotechnology and in medicine. Consequently, we envision that our studies will have a wide impact, with health benefits and improved living and economic standards nationally and internationally.