Single-molecule and single-cell studies of pro- and anti-mutagenic DNA repair in live Escherichia coli cells

All organisms incur damage to their DNA in the course of their daily lives and have developed sophisticated mechanisms for DNA repair. Many of the enzymes involved in these DNA repair pathways have been identified and their biochemical activities are well understood. However, there is a relatively poor understanding of how these enzymes are put into play in response to DNA damaging agents. As bacterial resistance to antibiotics has increasingly become a major world health problem, it is imperative to understand how DNA damage induced by antibiotics interacts with bacterial DNA repair systems. Detailed understanding of the molecular mechanisms of DNA repair could allow for the development of new antibiotics and, just as importantly, allow us to identify the sources of antibiotic-resistance mutations. The goal of this thesis is to better understand the anti- and pro-mutagenic DNA repair pathways in live Escherichia coli cells. The primary focus is on the interplay between the error-prone DNA polymerases and their binding partners. There is also significant focus on the resolution of toxic DNA repair intermediates. Utilising single-molecule fluorescence live-cell imaging, it has been possible to monitor the activities of proteins involved in these pathways in real time to gain new insights into their behaviour.