Structure and biochemical mechanism of DNA replication initiation machines

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Proteins are essential components of cells that do all the jobs that ensure the continuity of life. Some of them assemble to form "nanomachines" that do work. This work often involves rearranging or remodelling small molecules, other proteins or the DNA that encodes the information of life. Some of these machines are made up from groups of proteins that work together but others are proteins that act alone. Understanding how individual biological machines function is crucial because if they fail the consequences can be catastrophic for living organisms. For example, some cancers and neurodegenerative disorders are caused by the failure of nanomachines that process the DNA in our cells. In addition, pathogens that invade the body, such as viruses, bring their own nanomachines that destroy healthy cells or hijack them to make the virus stronger. Identifying these machines and understanding how they function can therefore be a starting point for combating diseases. We are studying the DNA replication proteins from a group of pathogenic viruses, the papillomaviruses (PV), that cause warts and serious diseases such as cancer. Accurate replication of our own DNA is essential for genome stability and healthy ageing. These systems have many components and to understand the process we require knowledge of the underlying mechanisms. The viral proteins we are studying, known simply as E1 and E2, bind to DNA and change its structure so that it can be replicated. In our own cells a complex assembly of many different proteins is required to do the same jobs that the PV E1 protein does. The aim of our research is to determine how the viral proteins E1 and E2 function on DNA during replication, to understand processes that are essential for life where potential therapeutic strategies could emerge to improve health and well-being.To do this we have designed ways to characterize these exceptionally small protein machines. Just as complex man-made mechanical machines could never be understood without understanding how they are made up, we need to know the structure of protein nanomachines to know how they work. We use biochemistry, powerful electron microscopes and computer technology to generate images of protein complexes and then use this information to re-construct their detailed three-dimensional structures. In parallel we apply biochemical techniques to understand the reactions performed by these proteins and relate this information to the structures we observe. Ultimately, we expect that our studies will have a significant impact in the healthcare sector and wider economy. The potential applications of our work include designing drugs to target viral proteins required for their replication. This research is important since it is estimated that up to 5% of all cancers are caused by human papillomavirus (HPV) and up to 600 million people are infected with HPV at any one time but there are currently no drug treatments of proven efficacy. Our studies will also inform us of how the more complex cellular counterparts of the PV replication proteins work, where there are further therapeutic applications to treat cellular and microbial (e.g. fungal) diseases. There are also potential applications in nanobiotechnology. This branch of science aims to develop small mechanical devices based on protein machines that can be harnessed to drive or alter molecular processes for our benefit.