Award details

Structural studies on prion proteins and related molecules

Reference	BS308136
Principal Investigator / Supervisor	Professor George Dodson
Co-Investigators / Co-Supervisors	Dr Peter Bayley, Professor Christopher Bostock
Institution	The Francis Crick Institute
Department	Division of Protein Structure
Funding type	Research
Value (£)	346,883
Status	Completed
Type	Research Grant
Start date	05/01/1998
End date	05/01/2001
Duration	36 months

Abstract

This proposal aims to obtain structural information to elucidate the role of the prion protein PrP in the molecular pathology of spongiform encephalopathies, using spectroscopic methods and X-ray crystallography. Particular emphasis is given to understanding the physical properties of the protein, and to developing new approaches involving protein engineering and novel crystallographic techniques. Intact murine PrPc and selected mutants will be expressed in an optimised CHO system and in E. coli systems. The recombinant full-length aglycosyl protein is currently secreted from the CHO cells at ca 5mg/L, and can be purified in non- denaturing conditions. A variety of constructs coding for intact PrPc and fragments will be expressed from E. coli in experiments in which selected and random mutations will be screened for high and soluble expression. The three-dimensional structure of PrPc 121-231 will be used to design point mutations, and to identify suitable sites for the deletion of loops and making other modifications. Biophysical studies on the recombinant proteins will be carried out to characterise and to improve their solubility, monodispersity and stability. Spectroscopic experiments will be used to characterise the folded states of the intact PrPc and its mutant derivatives, in relation to the PrPc > PrPsc switch. The effects of organic solvents, metal ions, antibodies and of heparin (and other anti-scrapie prophylactic molecules) will be studied as possible stabilisers of the conformation of PrPc. These results will provide the basis for the systematic programme to crystallise purified and soluble PrPc and fragments for X- ray analysis.

Summary

unavailable

Committee	Closed Committee - Agri-food (AF)
Research Topics	X – not assigned to a current Research Topic
Research Priority	X – Research Priority information not available
Research Initiative	Biology of Spongiform Encephalopathies - Phase 3 (BS3) [1996-1997]
Funding Scheme	X – not Funded via a specific Funding Scheme

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