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Molecular mechanism of triggering of a bacterial transcription activator

Reference	BB/C501484/1
Principal Investigator / Supervisor	Professor Steve Busby
Co-Investigators / Co-Supervisors
Institution	University of Birmingham
Department	Sch of Biosciences
Funding type	Research
Value (£)	208,593
Status	Completed
Type	Research Grant
Start date	01/12/2004
End date	30/09/2008
Duration	46 months

Abstract

The aim of the proposed work is to identify determinants in the E. coli MeIR transcription activator that are important when it is functioning (I) as a transcription activator, or (ii) as a transcription repressor. To do this, 2-hybrid screen will be used to identify different surfaces responsible for MeIR dimension that are important either for activation or repression. Concerning activation, known to be dependent on the trigger ligand, melibiose, genetic analyses will be used to identify the melibiose site in the MeIR N-terminal domain, they key sidechains in the dimerisation surfaces, and sidechains that are involved in relaying the activation message to the DNA-binding C-terminal domain. This will involve the isolation and characterisation of both mutants that are defective in activation and mutants that are able to activate in the absence of melibiose. The isolation will involve screens of mutant libraries of the meIR gene generated by PCR and the characterisation will exploit in vitro systems that we previously developed to study MeIR. Concerning repression, a parallel genetic analysis will be used to identify the key sidechains specific for repression, and to locate the dimerisation surface that permits DNA looping, which has been proposed to be essential for repression. An in vitro system will be exploited to study MeIR-dependent repression and to investigate the role of the Fis protein in maintaining this repression. In the final part of the work, the organisation of the activatory ternary MeIR-CRP-promoter DNA complex will be investigated, using genetic analysis to pinpoint the CRP-MeIR interface and chemical probes to trace the path of the DNA.

Summary

unavailable

Committee	Closed Committee - Plant & Microbial Sciences (PMS)
Research Topics	Microbiology
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

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