Development Electrochemical Imaging Techniques for Biophysical Applications

Investigation of Membrane Processes in Chloroplasts and Thylakoids

Supervisors: Julie MacPherson, Patrick Unwin and Colin Robinson

My project concerns the development of new high resolution quantitative electrochemical imaging techniques for the study of physicochemical processes at the micro and nanoscale. These techniques will be used to study important processes at chloroplasts, thylakoid membranes and proteins reconstituted into liposomes, in order to provide a detailed understanding of basic biophysical processes. The methods will have widespread applicability in the life sciences in the future.

Electrochemical imaging techniques involve the use of a tiny, mobile probe electrode, which can be used to monitor and/or perturb the chemical environment adjacent to a sample. Scanning electrochemical microscopy (SECM) and scanning ion conductance microscopy (SICM) provide a means of detecting and visualising a wide range of interfacial physicochemical processes at the micrometre and nanometre scale. One of the primary goals of this project is to develop sophisticated and robust techniques supported by simulations of key physicochemical processes related to membrane processes.

Chloroplasts perform all the primary processes of photosynthesis, such as light capture, electron transport leading to NADPH and ATP synthesis, and also most of the secondary processes, e.g. those involved in carbon formation. Chloroplasts are typically ca. 4 – 10 µm across and 1 µm thick. The main light-harvesting pigments, chlorophyll molecules, are usually found as chlorophyll-protein complexes embedded in, or associated with, the lipid bilayer forming the backbone of the thylakoid membranes. While the basic physicochemical processes associated with chloroplasts, thylakoids and membrane proteins in reconstituted bilayers have been intensively studied, measurements have been averaged over ensembles of material and little is known on the relationship between structure and function at the level of a single organelle or sub-organelle. The electrochemical imaging techniques, SECM and SICM, allow single organelle and sub-organelle measurement to be made. This project will provide this information through studies of ion transport (of key trace-level ions), molecular transport, photo-induced electron transport and membrane insertion processes.

Papers:

Intermittent Contact-Scanning Electrochemical Microscopy (IC-SECM): A New Approach for Tip Positionong and Simultaneous Imaging of Interfacila Topography and Activity
K. McKelvey. M. A. Edwards and P. R. Unwin, Anal. Chem., 2010, ASAP.