Award details

How is protein folding coupled to protein binding? Properties of the colicin N translocation domain and its interaction with TolA

Reference	BB/C507402/1
Principal Investigator / Supervisor	Professor Geoffrey Robert Moore
Co-Investigators / Co-Supervisors
Institution	University of East Anglia
Department	Chemistry
Funding type	Research
Value (£)	213,395
Status	Completed
Type	Research Grant
Start date	01/03/2005
End date	29/02/2008
Duration	36 months

Abstract

The focus of this proposal is the characterisation of how the pore-forming colicin N (ColN) produced by some strains of Escherichia coli interacts with the C-terminal domain of E. coli TolA, an interaction that is essential for the passage of ColN into an E. coli cell where it creates a pore in the periplasmic membrane. ColN is a 43 kDa 3-domain protein with its TolA binding site located in its natively unfolded N-terminal 90 amino acid translocation domain (ColN-T). Due to the flexibility of this domain it is not visible in X-ray electron density maps but is detectable by NMR spectroscopy of the intact ColN. NMR studies of ColN and peptides ranging from the entire ColN-T to smaller fragments of it have shown that through flexible and largely disordered, ColN-T does have a compact structure, albeit one lacking secondary structure. When bound to the 13 kDa C-terminal domain of TolA (TolA-III) however, CoIN-T folds into a more structural state. This project will use NMR spectroscopy of 13C/15N and 15N labelled ColN, ColN-T, fragments of ColN-T and TolA-III to determine the ensemble conformational properties of the free translocation domain and the structure of the TolA-III/ColN-T complex. Based on these structures, mutations will be introduced into ColN-T designed to destabilise it and their effect on the conformational properties of ColN-T and its interaction with TolA-III will be studied by CD, NMR and fluorescence spectroscopies. The aim of this will be to determine how the folding of ColN-T is linked to its binding of ToIA-III. The project will provide fundamental information on an important type of colicin protein interaction as well as addressing key important issues concerned with protein folding. (Joint with BB/C507396/1)

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	X - not in an Initiative
Funding Scheme	X – not Funded via a specific Funding Scheme

Associated awards:

BB/C507396/1 How is protein folding coupled to protein binding? Properties of the colicin N translocation domain and its interaction with TolA.

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