Research Opportunities 2010-11 Dr Roman Tuma

Dr Roman Tuma and Prof Peter G. Stockley

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Development of novel single molecule tools to follow assembly and dynamics of large macromolecular complexes

Dr Roman Tuma and Prof Peter G. Stockley

The purpose of this project is to develop new single molecule technology to follow assembly of macromolecular complexes in vitro and in living cells. The position would suit a students with a strong interest and background in biophysics or optoelectronics who wants to expand their expertise in biology.

Most of cellular processes are performed by large molecular machines that are composed of multiple protein and nucleic acid subunits. Advances in single molecule techniques revolutionized biophysics and enabled unprecedented insights into the function of such complex molecular machines. The main goal of the project is to establish experimental techniques which would enable characterization of large macromolecular assemblies at single molecule level and in living cells. In particular, the focus is on visualizing the self-assembly of large complexes, such as viruses, in vitro and within cells. This project draws on the existing strength and expertise in self-assembly and single molecule techniques in the Astbury Centre (1-3). The project is divided into two phases. During the first phase the existing single molecule technology will be upgraded to allow multiple excitation/multiple fluorophore imaging in vitro and in live cells. The new technology will be validated by following entry of individual in vitro labelled viral capsids into the host cell. During the second phase efficient methods for in vivo labeling of viral protein will be developed. Then the imaging technology will be used to follow interaction of viral proteins in vitro and in cells. New data processing techniques will be adopted and developed at this stage to allow application to virus assembly at the final stages of the project. The technology developed here will help to answer some of outstanding and fundamental questions in virology: How does assembly of viral capsids in cells differ from the defined in vitro conditions? What leads to the formation of “viral factories” (i.e. non-membrane compartments) where assembly of certain viruses takes place? The detailed dissection of viral assembly will provide basis for design of novel antiviral drugs that target the key stages of the process.

This multidisciplinary project will provide opportunities for the student to receive training in biophysics, virology and chemical biology.

Please contact Dr. Roman Tuma () for further details about this opportunity.

References

  1. Basnak, G; Morton, VL; Rolfsson, O; Stonehouse, NJ; Ashcroft, AE; Stockley, PG (2010) Viral genomic single-stranded RNA directs the pathway toward a T=3 capsid.. J Mol Biol 395(5), 924-936A.
  2. Gell, C; Sabir, T; Westwood, J; Rashid, A; Smith, DAM; Harris, SA; Stockley, PG (2008) Single-Molecule Fluorescence Resonance Energy Transfer Assays Reveal Heterogeneous Folding Ensembles in a Simple RNA Stem-Loop. J MOL BIOL384(1), 264-278
  3. E. Wallin, H. Ojala, E. Hæggström R. Tuma(2008). Stiffer optical tweezers through real-time feedback control. Appl. Phys. Lett.92, 224104 (2008).

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