Single-molecule imaging of bacterial gene expression and antibiotic resistance
As Richard Feynman noted back in 1959, 'it is very easy to answer many of these fundamental biological questions…you just look at the thing!' Our research group is turning this bold vision into a reality by developing and applying cutting-edge fluorescence spectroscopy and imaging methods (single-molecule FRET, super-resolution imaging, and single-particle tracking) to monitor sub-nanometre motions of single molecular machines with sub-millisecond temporal resolution both inside and outside living cells.
Much of our single-molecule work focuses on the mechanisms of the protein RNA polymerase, a molecular machine that copies DNA into RNA during the process of gene transcription. This protein is a major target for antibiotics, making our work important for tackling antibiotic resistance, one of the most pressing health challenges the world faces today.
For examples of our recent work, see:
Kummerlin et al. Bleaching-resistant, Near-continuous Single-molecule Fluorescence and FRET Based on Fluorogenic and Transient DNA Binding. ChemPhysChem 2023, 24, e202300175.
El Sayyed H et al. Single-molecule tracking reveals the functional allocation, in vivo interactions and spatial organization of universal transcription factor NusG. Mol Cell 2024, 84 (5), 926-937. e4.
We are also developing advanced microscopy for single-cell bacterial imaging and sophisticated image analysis (based largely on AI and deep learning) to find whether an antibiotic is effective in stopping bacteria from growing; these tests will also identify the type of bacteria causing the infection. This is a collaboration with the John Radcliffe Hospital and the Big Data Institute. For a recent example of our work in this area, see Zagajewski A et al. Deep learning and single-cell phenotyping for rapid antimicrobial susceptibility detection in Escherichia coli. Communications Biology 2023.
We have available projects in the areas of in vitro and in vivo transcription, as well as in AI-based antibiotic resistance detection, and interested students are encouraged to contact Prof Kapanidis to discuss options.
Biological rotary molecular motors
Nature DID invent the wheel, at least 3 times! The bacterial flagellar motor is 50 nanometres across, spins at over 100,000 rp. driven by electric current, and propels swimming bacterial cells. ATP-synthase is even smaller, about 10 nm across, consists of two rotary motors coupled back-to-back, and generates most of the ATP – life's 'energy currency – in most living organisms. We are trying to discover how these living machines work. We develop and use a range of methods in light microscopy including ultra-fast particle tracking, magnetic and optical tweezers and single-molecule fluorescence microscopy. Current projects in the lab: measurement of stepping rotation and mechano-sensing in flagellar motors, synthetic biology using ATP-synthase,single-molecule fluorescence microscopy of protein dynamics in bacterial motility and chemotaxis, high-torque magnetic tweezers, templated assembly of flagellar rotors using DNA nanotechnology scaffolds (collaboration with Turberfield group). More information on Oxford Molecular Motors can be found here.
Ion channels and nanopores: from structure to function
Almost every single process in the human body is controlled at some level by electrical signals, from the way our hearts beat, the way our muscles move, to the way we think. These electrical signals are generated and controlled by a family of proteins called 'ion channels' which reside in the membrane of every living cell and which act as 'electrical switches' to control the selective movement of charged ions like potassium (K+) and Sodium (Na+) into and out of the cell.
Work in our lab uses a range of multidisciplinary approaches (molecular biology, electrophysiology, single-molecule fluorescence, molecular dynamics and crystallography) to study the structure and function of these channels. We have a range of projects available to people with physics, engineering, computing, biochemistry and physiology backgrounds.
In particular, we currently have an exciting new DPhil project available to investigate how the unusual behaviour of water within the nano-sized pore of an ion channel influences its behaviour. This project would suit someone with a computational background and/or programming skills as it involves development of a new software tool to annotate membrane protein structures.
In collaboration with other colleagues in Oxford, we also have PhD projects available to work on the structural determination of ion channel drug binding sites using serial femtosecond crystallography and X-ray free electron laser (XFEL).
Understanding the dynamics of DNA and chromatin replication
Nynke Dekker
During our lifetimes, we copy approximately a lightyear’s worth of DNA, and how the different components of the molecular machinery (the replisome) work together to achieve this successfully is an area of highly active research. Here, you will take on the exciting challenge of understanding the dynamics of DNA and chromatin replication by purifying, reconstituting, and studying the activity of eukaryotic replisome at the single-molecule level. You will examine replisome composition, replisome motion dynamics, and the interplay between these two quantities; and examine how these change in the context of chromatin or obstacles on the DNA. To do so, you will employ a combination of novel biophysical instrumentation (e.g. fluorescence imaging, single-molecule force spectroscopy, microfluidics, cryo-electron microscopy) and biochemical approaches (protein purification, biochemical assays, DNA sequencing). You will analyze the resulting datasets using biophysical modelling, interact with collaborators in molecular biology and biochemistry at the University of Oxford and at highly regarded institutions elsewhere in the United Kingdom. In doing so, you will publish high-quality scientific papers to advance this exciting field and ultimately, its impact on biomedical applications