Award details

The study of membrane protein organisation by atomic force microscopy

Reference	BB/C511156/1
Principal Investigator / Supervisor	Krebs Professor of B Christopher Hunter
Co-Investigators / Co-Supervisors
Institution	University of Sheffield
Department	Molecular Biology and Biotechnology
Funding type	Research
Value (£)	169,720
Status	Completed
Type	Research Grant
Start date	24/06/2005
End date	23/03/2006
Duration	9 months

Abstract

The overall aim is to use atomic force microscopy to study the organisation and topology of membrane protein complexes, initially using photosynthetic membranes as model systems. Given the universal importance of membrane proteins it is essential to understand the macromolecular arrangements of membrane proteins in their native membrane environment and the forces that stabilise these associations. This is significant since many membrane proteins do not work in isolation, but instead they must cooperate, for example in mitochondrial electron flow, so that electrons are transferred efficiently between complexes. The structures of almost all of the major energy transducing complexes in biology are known to atomic resolution, and the direct visualisation of these complexes within their native membrane environment is timely and the next step to take. In the longer term this methodology will provide researchers in the Department of Molecular Biology and Biotechnology at Sheffield with the capability of using Atomic Force Microscope (AFM) to study the topology and organisation of protein complexes within biological membranes, for research projects in molecular medicine, molecular microbiology and molecular plant science. This proposal exploits the fact that photosynthetic membranes, within which many membrane proteins associate in order to harvest light and to convert it to chemical energy, are arguably the best characterised of all membrane systems. It also exploits existing expertise in molecular genetic and biochemical analyses of bacterial photosynthetic membranes, and proposes to advance the direct analysis of membrane protein organisation and topology by AFM. The proposal employs a multidisciplinary approach, employing molecular genetics, biochemistry, structural biology, and state of the art physical imaging technology. The specific objectives are: (1) to map the organisation, distribution and interactions between complexes in native photosynthetic membranes, and (2) to study the interaction forces between a membrane bound and a soluble protein using the interaction between the reaction centre and cytochrome c2 as a model system. We believe that the proposed work will have a wider relevance to the study of other active aggregations of membrane proteins in, for example, plant thylakoids and mitochondrial membranes. Funding is sought for an AFM, which will allow the topography of biological samples to be acquired at less then 10 Angstrom lateral resolution and vertical resolution of approximately 1 Angstrom under near physiological conditions, in order to reveal molecule-to-molecule variations in structure, and details of the way in which complexes sit in the membrane, in terms of the height of their components above the membrane bilayer. There are no facilities for examining membrane proteins in their native environment in Biosciences at Sheffield; the acquisition of a versatile high performance AFM will bring to the Department a completely new set of methodologies for examining this biologically crucial class of proteins, as well as forming part of an interdisciplinary project, being pursued jointly with Professor Graham Leggett in the Department of Chemistry, which is at the physical sciences-life science interface, and so research training for this and other projects will be considerably enhanced.

Summary

unavailable

Committee	Closed Committee - Biomolecular Sciences (BMS)
Research Topics	X – not assigned to a current Research Topic
Research Priority	X – Research Priority information not available
Research Initiative	Research Equipment Initiative 2004 (RE4) [2004]
Funding Scheme	X – not Funded via a specific Funding Scheme

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