Characterising DNA-protein interaction through the automated analysis of AFM images

Software for automatic image analysis has been widely applied to the characterisation of
biomolecules as it allows for faster data processing and reduces bias. However, many tools
developed for other bio-imaging fields are not directly compatible with atomic force
microscopy (AFM), which is a high-resolution characterisation technique that enables the
direct visualisation of biomolecules in a physiological environment. AFM presents unique
challenges to the implementation of automatic image analysis tools because it stores data in
special formats and is affected by artefacts that find no equivalence in other microscopy
techniques.
While automatic analysis methods tailored for AFM images have been developed, most
existing programs are project-specific and unsuitable for different sample types, while other
programs see limited usage due to their cost or unsuitability for batch processing.
To address these issues and contribute towards the establishment of a versatile and
accessible AFM bio-image analysis platform, this project improves existing automatic
analysis methods and develops more methods to characterise the conformation of DNA and
proteins. Since DNA is a flexible molecule that exhibits both intrinsic conformational
variation and conformational changes induced by protein interaction, these characterisation
methods will allow us to better understand how DNA functions and how its shape is altered
by proteins.
Specifically, this thesis analyses the conformation of the DNA-manipulating topoisomerase
TOP1, investigates the interaction between DNA and dumbbell-shaped autophagy receptor
NDP52, and probes the effects of the nucleoid-associated protein HU on DNA bending angle.
The work in this thesis is built on open-source software and forms part of a general AFM
image processing workflow. The methods developed can therefore be easily adapted for
other projects that study the conformation of different biomolecules.