Award details

Structure-function relationships in an AAA+ transcription activator

Reference	BB/D521922/1
Principal Investigator / Supervisor	Professor Martin Buck
Co-Investigators / Co-Supervisors	Mr Mathieu Rappas, Professor Xiaodong Zhang
Institution	Imperial College London
Department	Biological Sciences
Funding type	Research
Value (£)	226,322
Status	Completed
Type	Research Grant
Start date	01/11/2005
End date	31/10/2008
Duration	36 months

Abstract

The bacterial RNA polymerase containing the sigma 54 promoter specificity factor is functionally very distinct from the common cellular RNAPs (i.e. those associated with sigma factors of the sigma 70 type). Hence, unlike the common cellular RNAP form, the sigma 54-RNAP forms a complex with promoter DNA (the closed complex) that is initially inhibited for transcription. To initiate transcription, the sigma 54-RNAP must collaborate with a specialised form of transcription activator protein that belongs to the AAA+ (ATPases Associated with various cellular Activities) protein family. In an ATP-dependent reaction, collectively termed energy coupling, the AAA+ activator utilises the energy derived from ATP hydrolysis to interact with the closed complex and brings about conformational changes in the sigma 54-RNAP and the promoter DNA which subsequently lead to DNA opening and transcription by the RNAP. The major binding target for the AAA+ activator during energy coupling is the amino terminal regulatory domain of sigma 54, known as Region I. However, emerging proximity-based studies suggest that during the energy coupling process, the AAA+ activator also interacts with promoter DNA sequences downstream of the sigma54-RNAP binding site which are normally melted out during transcription initiation. We have recently determined the crystal structure of one model AAA+ activator of the sigma 54-RNAP and have identified two distinct structural features, referred to as Loops 1 and 2, which function in energy coupling in a nucleotide dependent manner. In this proposal, we wish to study the nucleotide dependent relationship between Loops 1 and 2 and in so doing want to elucidate the conformational signalling pathway(s) that operate during energy coupling. We will also investigate the residues in PspF that interact with the to be melted DNA sequences during energy coupling and determine the functional significance of this interaction. Finally, we want to identify the residues in PspF that are involved in nucleotide hydrolysis, in particular the so-called R-finger.

Summary

Many essential processes in the living cell are carried out by specialised small machines, called molecular machines, which alter the materials they interact with. We are interested in the machinery in bacteria that controls the flow of information from the DNA of the cell into the formation of the components of the cell, called proteins, which allow the cell to function and grow. In particular, we wish to understand how one molecular machine uses a small molecule called ATP as fuel to interact with and bring about transformations in the structure of another larger molecular machine that reads the information contained in the DNA. This process is important for controlling whether the information contained in the DNA is used or not and ultimately is important for the survival of the cell.

Committee	Closed Committee - Genes & Developmental Biology (GDB)
Research Topics	Microbiology, Structural Biology
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

I accept the terms and conditions of use (opens in new window)

export PDF file

back to list new search