PhD in Biophysics, University of East Anglia
BSc Hons in Zoology, University of Liverpool
We aim to use biophysics in combination with molecular biology to understand dynamic aspects of life at the molecular level. Nevertheless we try to avoid the "The Perils of Reductionism" described by Albert Szent-Gyorgi (see footnote) by remembering the biological relevance in our work. This approach means that we work on a large range of proteins but our core research interests centre on the structure function relationships of bacterial pore-forming and membrane proteins. This includes both those that can be classed as integral membrane proteins and those soluble toxins that exist as both soluble and integral membrane proteins. In addition to their important role in bacterial pathogenesis and physiology, we see such proteins as important model systems to aid our understanding of pore function, protein folding and ligand/receptor interactions.
Colicin function: Colicins are a plasmid-borne family of toxins from E coli whose role is to kill closely related competing bacteria. It has been shown that 40% of enteric gram negative bacteria carry a colicinogenic plasmid. The plasmid also confers upon the host a resistance to the toxic effects of the colicin and thus it was possible for colicins to evolve into a very efficient killing machine. The colicins which are large proteins traverse the outer membrane and deliver a toxic activity to the cytoplasmic compartment of the cells. In doing this they perform several functions which are common to many other cellular systems. They must recognise the target cell by binding to a specific receptor, they must then translocate a sizeable protein domain through a membrane and across the periplasmic space and finally they must penetrate the inner membrane to reach the highly sensitive cytoplasm. Somewhere during this process the immunity protein must be able to act to protect the host cells.
The toxic activity of colicins is of two types, nuclease (in which an RNase or DNase activity is transported to the cytoplasmic space) or pore-forming (in which the toxin forms a pore in the cytoplasmic membrane causing death by ion efflux). Colicins are modular proteins and the C-terminal domain carries the toxic activity. This means that, despite the difference in toxicity, the two groups of colicins share many features of receptor-binding and translocation. Our groups interest is in Colicin N which is the smallest colicin and is a member of the pore-forming division. We are using it to understand the molecular mechanism behind basic cellular processes such as receptor recognition, protein translocation, pore-formation and protein-domain interaction.
Outer membrane proteins: The porins of Escherichia coli are well characterised ion channel forming proteins from the outer membrane. Their structures have been determined to high resolution and are still members of a very small group of structures of channel forming proteins. We investigate these proteins by a mixture of single channel measurements in artificial planar bilayers, spectroscopy (CD, Fluorescence), binding studies using isothermal titration calorimetry or Biacore and protein engineering to alter or define function. We have investigated the voltage gating characteristics, the pore selectivity and their ligand binding activities and their role as colicin N translocators.
Protein structural modelling and solution behaviour: We are interested in applying model building and spectroscopy to the everyday analysis of proteins. Thus we link molecular modelling studies which predict structure or solution characteristics to rapid methods of analysis using circular dichroism, fluorescence, calorimetry etc . These approaches are often extremely helpful in the analysis of data generated by genome projects where predicted characteristics of proteins need to be tested. Another field where this approach is useful is the design and analysis of engineered proteins. This work involves collaboration with both academic and industrial scientists. The three-dimensional folding characteristics of therapeutic proteins in solution are being increasingly seen as an important quality issue.
Bionanotechnology: Nanotechnology will rely upon both top down and bottom up approaches to manufacture small devices. We are harnessing the two dimensional self assembly characteristics of membrane protein arrays to create nanoscale interfaces. By engineering these protein scaffolds supplied by nature we wish to create a toolkit for the rapid inclusion of biology into nanotechnology.