Measuring nanometre distance changes in biomolecules under pressure

Lead Research Organisation: University of St Andrews
Department Name: Physics and Astronomy

Abstract

Biomolecules, such as proteins, function by means of changes in their structural arrangement. Observing and understanding these changes can allow for detailed elucidation of biochemical mechanisms, which in turn can lead to advances in fields such as nanotechnology and drug discovery.

Distances on a nanometre scale can be measured between pairs of specifically placed spin-label molecules using the pulsed EPR technique, double electron-electron resonance, DEER (PELDOR). Moving the locations of these labels within a protein can enable its structure to be mapped. As well as distances, the relative orientation of these labels can also be determined which can provide further information on the structure of the protein (G. Jeschke, Annu. Rev. Phys. Chem. (2012) 63, 419-446).
While proteins typically exist in well-ordered native states under physiological conditions, their functions may cause them to move to excited states. Due to these states having higher energies, lower equilibrium populations and conformational flexibility, the excited states cannot be measured by traditional spectroscopic techniques. Interestingly, this can be overcome by subjecting the sample to high pressure, which can result in changes to the structural state of the protein and can populate the excited state (K. Akasaka, Biochemistry. (2003) 42, 10875-10885). Combining this with site-directed spin labelling and DEER spectroscopy can allow for these states to be accessed, if the sample can be rapidly frozen (M.T. Lerch et al, Proc. Natl. Acad. Sci. (2014) 111, E1201-E1210).
In this work, we will apply this methodology to the messenger protein calmodulin which undergoes structural changes both with the calcium concentration as well as with the presence of binding proteins. We plan to measure at Q-band frequency and also at W-band using our home-built spectrometer, HiPER (P.A.S. Cruickshank et al, Rev. Sci. Instrum. (2009) 80, 103102).

Agreed training requirements are to learn best research practice, scientific writing for publications and thesis, and to learn various research techniques. On top of these, further academic development is required in the form of QM-CDT training blocks, taught SUPA courses, attendance at the weekly colloquium, and transferrable skill courses. Further, involvement in public engagement and teaching is expected.

Publications

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Studentship Projects

Project Reference Relationship Related To Start End Student Name
EP/R513337/1 01/10/2018 30/09/2023
2276826 Studentship EP/R513337/1 01/09/2019 28/02/2023 Hannah Russell