Structural and biophysical studies of transcriptional regulatory and anti-termination complexes

Lead Research Organisation: MRC National Inst for Medical Research

Abstract

The work involves the application of biophysical and structural methods to investigate the mechanism of transcriptional regulation.

Technical Summary

Transcriptional anti-termination is a ubiquitous mechanism responsible for regulation and control of gene expression in eubacteria, archaebacteria, eukaryotic viruses and probably also in eukaryotes. The basic tenet of any antitermination mechanism is that the RNA polymerase (RNAP) becomes modified in such a way that it is able to readthrough termination signals allowing it to transcribe into a downstream gene, or further into an operon. The long term goal and objective of this work is to obtain detailed biochemical, biophysical and structural information concerning transcriptional anti-termination complexes in an attempt to provide insight into this important and ubiquitous mechanism of gene regulation. To do this we plan to undertake two main areas of study. The first will concentrate on the identification of components of anti-termination complexes using immunoprecipitation methods combined with mass spectroscopy. The second will focus on the detailed in vitro characterisation of the proteins and complexes involved in antitermination. This will combine structural data obtained from X-ray crystallographic, cryo-electron microscopy and magnetic resonance methods with kinetic and equilibrium binding data obtained from biophysical and biochemical measurements.||Initially, we will focus on the identification and characterisation of the components of antitermination complexes derived from E. coli and M. tuberculosis. However, this will be quickly extended to look at complexes from other organisms such as Mycobacterium leprae and the related but non-pathogenic Mycobacterium phlei and Mycobacterium smegmatis. As rRNA synthesis is intimately linked with bacterial growth rate we will be able to make a valuable comparison between the composition of antitermination complexes from slow growing pathogenic bacteria and the fast growing non-pathogenic varieties. We will also investigate sensor kinase directed antitermination mechanisms, initially concentrating on the proteins from the well-characterised E. coli and B. subtilis Bgl and Sac systems. However, we would also like to look at these systems in slow growing pathogenic mycobacteria, where the control of nutrient uptake is an important issue and sensor kinase directed antitermination mechanisms may well be involved.

Publications

10 25 50
 
Description Structural studies of JSRV Gag proteins 
Organisation University of Glasgow
Department School of Veterinary Medicine Glasgow
Country United Kingdom 
Sector Academic/University 
PI Contribution structural analysis of the JSRV gag proteins
Collaborator Contribution undertook complementary virological studies and together with the structural studies in my lab resulted in publication
Impact publication of paper in 2009 Pubmed ID 19007792
Start Year 2007
 
Description Structure and function of SAMHD1 
Organisation University of Manchester
Department School of Medicine Manchester
Country United Kingdom 
Sector Academic/University 
PI Contribution My team determined the structure and elucidated the mechanism of the HIV-1 restriction factor SAMHD1
Collaborator Contribution identifying the SamHD1 gene and demonstrating it's role in Aicardi-Goutieres syndrome
Impact Structure of SAMHD1 determined findings published in Nature 2011 and Plos pathogens 2015
Start Year 2016