Imaging DNA Replication at the Single Molecule Level

The Florey Institute (http://www.floreyinstitute.com) and Imagine: Imaging Life (http://www.imagine-imaginglife.com) are long term strategic initiatives at the University of Sheffield that provide a platform for ambitious research growth in areas that the University of Sheffield is recognised for internationally. The Florey Institute aims to create a world-leading focus on antimicrobial resistance and host-pathogen interactions, from fundamental science to translation. Imagine: Imaging Life aims to combine and harness cutting-edge imaging technologies in super-resolution microscopy, electron microscopy and atomic force microscopy to address key biological questions.

This project forms part of an interdisciplinary collaboration between Prof Jamie Hobbs (an experimental physicist working on the development and application of atomic force microscopy for understanding living systems) and Prof Jon Sayers (professor of functional genomics with interests in protein:DNA interactions and microbial proteases, pathogenesis and the host response). We are looking for a talented scientist from either the physical or life sciences with a strong desire to develop and apply cross-disciplinary approaches to a challenging problem. Flap endonucleases (FENs) play a vital role in DNA replication, repair and recombination in all living cells.

FENs are widely used as tools in molecular biology and clinical diagnostics. Aspects of how FENs locate their branched DNA substrates and the conformational changes associated with DNA hydrolysis remain unclear and could impact on medicine as well as biotechnology. We will address these questions using a combination of direct atomic force microscopy (AFM) imaging with conventional biochemical and structural approaches. AFM is a high resolution imaging technique that enables molecular and sub-molecular resolution imaging in liquid environments, allowing biological systems to be visualized at the molecular scale under physiological conditions. We have recently developed imaging of DNA to major and minor groove resolution using approaches initially proven for membrane protein imaging[1]. This provides the potential to directly visualize local strain within a single molecule during an interaction with a bound protein. Sayers has shown DNA threading through a FEN by X ray crystallography [2&3] and developed several mutants that stall at different processing stages. Using AFM we will address the question of how FENs engage their substrate using imaging of DNA-FEN complexes to characterize their hydrolysis. We seek enthusiastic applicants with first class hons or equivalent degree qualifications in any relevant area (chemistry, life sciences, physics) who are willing to take on an exciting emerging field. Training in all relevant areas will be provided but experience of one or more areas such as microscopy, crystallography, molecular biology, protein biochemistry is desirable.

Please contact Prof. Jamie Hobbs ([Email Address Removed]) and Prof. Jon Sayers ([Email Address Removed]) for more information. Please apply online through the University of Sheffield website, putting the department of Physics and Astronomy as the home department and Jamie Hobbs as the principle supervisor. For information on applying, please see https://www.sheffield.ac.uk/postgraduate/research/apply/applying

Award Details

• Full tuition fees (no geographic restrictions)
• An annual, tax-free maintenance stipend at the standard RCUK rate
These scholarships fund up to 3.5 years full-time study, subject to satisfactory progress.

Eligibility

• You should have drive, dedication, excellent academic qualifications including (or expect to achieve), a first class UK honours degree or equivalent. This should be in a relevant area (e.g. Physics, Chemistry, Engineering, Biochemistry, Microbiology).