Award details

Molecular basis of DNA gating by topo IV and gyrase and its inhibition by antimicrobial drugs

Reference	BB/K010069/1
Principal Investigator / Supervisor	Professor Larry Mark Fisher
Co-Investigators / Co-Supervisors	Dr Mark Sanderson
Institution	St George's University of London
Department	Basic Medical Sciences
Funding type	Research
Value (£)	633,049
Status	Completed
Type	Research Grant
Start date	01/12/2012
End date	30/11/2015
Duration	36 months

Abstract

Topo IV and gyrase mediate chromosome unlinking and DNA supercoiling and are targeted by clinically important antibacterial quinolones and other agents. Each enzyme transports a DNA duplex (the T segment) through a DNA gate made in a second DNA helix (the G-gate) in a process that involves the reversible formation of a covalent enzyme-DNA intermediate called the cleavage complex. In recent advances, we have determined the X-ray crystal structures of cleavage complexes of Streptococcus pneumoniae topo IV formed with quinolone and quinazolinediones. Building on these and other structures and new biochemical tools, we aim to examine how DNA is gated through the DNA enzyme complex and how novel drugs interfere with this process. The work will involve a mutidisciplinary approach employing biochemical, X-ray structure, EM and SAXS approaches. Completion of the study will provide new insights on DNA gating during catalysis by topo IV and gyrase and its arrest by anti-topoisomerase therapeutics.

Summary

Most life-forms whether humans, mammals, yeast or bacteria have a DNA genome. The DNA consists of two strands that wrap around each other to form the iconic DNA helix. During DNA replication, the helix must be unwound to allow copying and chromosome segregation. This process generates DNA supercoils and DNA tangles that can block replication. Topoisomerases solve the 'unwinding problem' by introducing transient single- or double-stranded breaks into DNA that dissipate supercoils and allow DNA synthesis to proceed. It is not surprising that topoisomerases are essential enzymes but they also turn out to be vital targets of antibacterial and anticancer drugs that inhibit cell growth. We aim to use biochemical and X-ray crystallographic approaches to study two bacterial topoisomerases called topo IV and gyrase. We wish to understand how these proteins act at the molecular level and how clinically important drugs such as fluoroquinolones target their functions. The work will focus on the well-characterised topo IV and gyrase from Streptococcus pneumoniae, a dangerous human pathogen that causes life-threatening pneumonia and meningitis. However the research has wider applications to other bacterial and microbial systems. Progress in this key area should aid development of new drugs that work against antibiotic-resistant organisms, a growing threat to our healthcare system.

Impact Summary

The pursuit of excellent basic research and its translation to drugs is an important feature of the proposal. The major beneficiaries who will be interested in or will benefit from the research are: The pharmaceutical industry developing new antibacterial topo IV/gyrase inhibitors as anti-infectives Microbiologists and drug developers interested in combatting the mechanisms of drug resistance Structural biologists and nano-engineers studying molecular machines A range of scientific disciplines with a focus on DNA-protein interactions The very young and elderly with pneumococcal disease Society at large- through effective disease management reducing the societal and economic costs of healthcare and hospitalisation.

Committee	Research Committee D (Molecules, cells and industrial biotechnology)
Research Topics	Microbiology, Pharmaceuticals, 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

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