Recognition, opening and stabilization of DNA gates by topo IV the chromosome decatenase

The enzyme topo IV mediates the untangling of bacterial chromosomes prior to their segregation at cell division. Topo IV crosses one DNA helix through another allowing the separation of topologically interlocked chromosomes. It does this by an unusual mechanism: it makes a transient double-stranded DNA break (often called a 'gate') in one molecule through which the second DNA molecule is passed. Antibacterial quinolone drugs act by stabilising the open DNA gate which interferes with DNA replication causing cell death. Indeed, quinolones are now important drugs for the treatment of pneumonia and other infections caused by Streptococcus pneumoniae and othe Gram-positive pathogens. Despite it scientific interest and medical importance, little is known about the mechanism of topo IV and its drug interactions. In this study, we aim to examine (i)how S. pneumoniae topo IV recognises and opens potential gate sequences in DNA, and (ii) how quinolones stabilize such gates. The work will make use of synthetic and altered versions of a known topo IV gate which we identified on the S. pneumoniae chromosome. We shall also study the binding of mutant topo IV complexes to identify key protein residues that function to contact and open the DNA gate. Progress on topo IV and its DNA gates will advance fundamental understanding of chromosome biology and will aid the future design of more effective quinolones.

Topo IV, the chromosome decatenase, is the target of antimicrobial quinolones used against Streptococcus pneumoniae and other Gram-positive pathogens. The enzyme acts by introducing a transient double-strand DNA break into one DNA helix (the G or gate segment)through which is passed a second helix (the T or translocated segment). Antipneumococcal quinolones exert their cytotoxic effects by stabilising the open gate form of the G segment, the crucial first step in topo IV action. We aim to establish the DNA and protein determinants for recognition, opening and stabilization of the DNA gate formed by S. pneumoniae topo IV, the gram-positive topo IV paradigm. These studies will provide important new insights into the mechanism of topo IV and its interactions with quinolones.