ATP-bound conformation of topoisomerase IV: a possible target for quinolones in Streptococcus pneumoniae.
Topoisomerase IV, a C(2)E(2) tetramer, is involved in the topological changes of DNA during replication. This enzyme is the target of antibacterial compounds, such as the coumarins, which target the ATP binding site in the ParE subunit, and the quinolones, which bind, outside the active site, to the quinolone resistance-determining region (QRDR). After site-directed and random mutagenesis, we found some mutations in the ATP binding site of ParE near the dimeric interface and outside the QRDR that conferred quinolone resistance to Streptococcus pneumoniae, a bacterial pathogen. Modeling of the N-terminal, 43-kDa ParE domain of S. pneumoniae revealed that the most frequent mutations affected conserved residues, among them His43 and His103, which are involved in the hydrogen bond network supporting ATP hydrolysis, and Met31, at the dimeric interface. All mutants showed a particular phenotype of resistance to fluoroquinolones and an increase in susceptibility to novobiocin. All mutations in ParE resulted in resistance only when associated with a mutation in the QRDR of the GyrA subunit. Our models of the closed and open conformations of the active site indicate that quinolones preferentially target topoisomerase IV of S. pneumoniae in its ATP-bound closed conformation.
Sifaoui
Farid
F
INSERM E0004, Laboratoire de Recherche Moléculaire sur les Antibiotiques, UFR Broussais-Hôtel-Dieu, Université Paris VI, 75270 Paris Cedex 06, France.
Lamour
Valérie
V
Varon
Emmanuelle
E
Moras
Dino
D
Gutmann
Laurent
L
D000255:Adenosine Triphosphate / Q000737:chemistry / Q000378:metabolism*; D000595:Amino Acid Sequence; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D000890:Anti-Infective Agents / Q000494:pharmacology*; D001372:Aza Compounds; D001665:Binding Sites / Q000187:drug effects; D027101:DNA Topoisomerase IV / Q000737:chemistry* / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D024881:Drug Resistance, Bacterial; D024841:Fluoroquinolones; D008958:Models, Molecular; D008969:Molecular Sequence Data; D000077266:Moxifloxacin; D016296:Mutagenesis; D016297:Mutagenesis, Site-Directed; D009675:Novobiocin / Q000494:pharmacology; D011487:Protein Conformation / Q000187:drug effects; D011804:Quinolines; D013296:Streptococcus pneumoniae / Q000201:enzymology* / Q000235:genetics
D000900:Anti-Bacterial Agents; D000890:Anti-Infective Agents; D001372:Aza Compounds; D024841:Fluoroquinolones; D011804:Quinolines; D009675:Novobiocin; D000255:Adenosine Triphosphate; D027101:DNA Topoisomerase IV; D000077266:Moxifloxacin