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Crystallography by UK scientists may have uncovered the mechanism by which quinolone drugs interact with DNA and bacterial topoisomerase and so point to a better understanding of how resistance to this class of drugs emerges in meningitis and pneumonia. Bacteria evolved resistance to antibiotics the moment the first prescription for the microbicidal drugs was written. However, in recent years an increase in antibiotic use often inappropriately as well as environmental and socioeconomic factors have led the emergence of strains of certain particularly lethal bacteria that are resistant even to the drugs held in reserve as a last line of defence. One particular group of chemicals, the quinolones, are currently used to treat virulent bacteria responsible for some of the most dangerous diseases such as pneumonia and meningitis. Unfortunately, rapid growth and mutability of the pathogens involved in these diseases is leading to emerging resistance to quinolones. In considering quinolone resistance, it is important to note that while it is present, it is not widespread, with only a few percent of pneumococcal isolates. Understanding how the quinolones work and how they interact with bacterial DNA at the molecular level could provide new insights allowing medicinal chemists to find a way to fight back. Research being undertaken at the Diamond national synchrotron light source in Didcot, UK and SOLEIL, its French counterpart in Gif-sur-Yvette, could provide new clues for drug development. Mark Sanderson, Ivan Laponogov, Maninder Sohi, and Dennis Veselkov of the Randall Division of Cell and Molecular Biophysics at King's College London collaborate with Mark Fisher, Xiao-Su Pan, and Ritica Sawhney of the Molecular Genetics Group at St. George's, University of London, Andrew Thompson at SOLEIL and Katherine McAuley of Diamond. The team has now published details of their recent work in the journal Nature Structural and Molecular Biology, which reveals a possible mechanism by which quinolone drugs interact with DNA and the bacterial enzyme, topoisomerase IV. "New antibacterial drugs are desperately needed to combat infections caused by resistant bacteria, such as MRSA, for which conventional antibiotics are ineffective," explains Sanderson, "Using the intense X-rays produced by the Diamond and SOLEIL synchrotrons, we now understand how quinolones interact at a molecular level with their bacterial targets. This knowledge will feed into the R&D programmes of drug companies who are developing the next generation of antibacterial drugs." He explains that in the 1960s, the then new family of antimicrobials known as quinolones was introduced to treat urinary tract infections at first. These compounds are inhibitors of bacterial topoisomerase but had only limited activity and resistant bacterial strains emerged quickly. The next generations of quinolones were more potent and had a better spectrum of activity. Today, quinolones are the second line of defence against S. pneumoniae disease, which includes pneumonia, meningitis, and otitis. This microbe causes hundreds of thousands of cases of pneumonia, millions of ear infections, tens of thousands of cases of invasive disease, including meningitis. "Type II topoisomerases alter DNA topology by forming a covalent DNA-cleavage complex that allows DNA transport through a double-stranded DNA break," the researchers explain. "Bacteria usually express two related type II topoisomerases - DNA gyrase, which regulates DNA supercoiling of the circular bacterial chromosome, and topoisomerase IV (topo IV), a ParC2ParE2 tetramer that mediates topological unlinking of catenated daughter chromosomes." These two enzymes work by splitting double-stranded DNA. The team has now looked at the structures of the cleavage complexes formed by two domains of topoisomerase IV - the Streptococcus pneumoniae ParC breakage-reunion and ParE TOPRIM domain and the effect on this process of each of two important fluoroquinolone drugs, clinafloxacin and moxifloxacin, the latter being used to treat respiratory tract infections. The crystal structures reveal two drug molecules intercalated at the highly bent DNA gate and help explain antibacterial quinolone action and resistance, the team says. The study helps resolve several problems in understanding the activity of quinolones against topoisomerases. First, the structures show that two quinolone molecules are present in the drug-stabilized cleavage complex, each stacked against DNA bases. This agrees with one earlier model but contradicts another that suggested four might be present. Secondly, they explain how mutations in the enzyme might prevent intercalation happening and so lead to bacterial resistance to these drugs. If a mutation prevents the drugs sliding into the DNA then they cannot prevent the supercoiling and unlinking processes used by the bacteria. "These new insights should provide a structural base for antibacterial drug development," the researchers conclude.
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Organised domains interface with quinolones and DNA |
