Mechanistic analysis of DNA helicases using nanopore-based single molecule assays

Next generation DNA sequencing methods such as those pioneered by Oxford Nanopore Technologies (ONT) have
revolutionised biology by allowing rapid DNA sequencing at the whole organism level and impacting fields as diverse as
evolutionary biology, structural and molecular biology, agriculture, diagnostics and personalised medicine. The ONT
technology works by coupling an ATP-dependent DNA motor protein to a nanoscale pore in a biological membrane. As
the motor delivers DNA through the pore, the base composition is determined by changes in the current across the
membrane ultimately yielding accurate long read DNA sequencing data. In addition to their core uses for long-read DNA
sequencing and rapid diagnostics, nanopore-based assays have also found unexpected applications in basic science. For
example, because nanopore DNA sequencing is rate-limited by the activity of the motor, the sequencing traces also
contain rich information on the kinetics of DNA translocation providing unique insights into the mechanism(s) of
essential enzymes such as helicases.
In this project, you will integrate new DNA helicases into nanopore-based DNA sequence devices with two major goals.
Firstly, we aim to improve the speed, accuracy, simplicity and robustness of nanopore DNA sequencing by using DNA
helicases with novel and desirable biochemical properties. Secondly, we aim to study the mechanism of action of DNA
helicases of medical interest and how they are affected by small molecule drugs or genetic mutation. Importantly, we
anticipate that nanopore traces will yield unprecedented detail on the nature of the individual steps that are made along
DNA and their chemo-mechanical coupling to ATP hydrolysis. Your project will be based in the laboratory of Prof. Mark
Dillingham in the DNA:protein interactions Unit at the University of Bristol but will also involve close collaboration with
Oxford Nanopore Technologies (1) including internships spent at their Oxford headquarters. The Dillingham lab (2) is
studying helicases involved in the repair of broken DNA, including bacterial enzymes that are considered attractive
targets for antibiotics and human enzymes implicated in genetic diseases including cancer. For further details and
examples of our recent work see our website or contact the laboratory (2).
(1) https://nanoporetech.com/ (2) https://research-information.bris.ac.uk/en/persons/mark-s-dillingham