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• W2057264691 abstract "Streptococcus pneumoniae is the major cause of community-acquired pneumonia and is also associated with bronchitis, meningitis, otitis and sinusitis. The emergence and increasing prevalence of resistance to penicillin and other antibiotics has led to interest in other anti-pneumonococcal drugs such as quinolones that target the enzymes DNA gyrase and topoisomerase IV. During crystallization and in the avenues to finding a method to determine phases for the structure of the ParC55 breakage-reunion domain of topoisomerase IV from Streptococcus pneumoniae, obstacles were faced at each stage of the process. These problems included: majority of the crystals being twinned, either non-diffracting or exhibiting a high mosaic spread. The crystals, which were grown under conditions that favoured diffraction, were difficult to flash-freeze without loosing diffraction. The initial structure solution by molecular replacement failed and the approach proved to be unviable due to the complexity of the problem. In the end the successful structure solution required an in-depth data analysis and a very detailed molecular replacement search.Crystal anti-twinning agents have been tested and two different methods of flash freezing have been compared. The fragility of the crystals did not allow the usual method of transferring the crystals into the heavy atom solution. Consequently, it was necessary to co-crystallize in the presence of the heavy atom compound. The multiple isomorphous replacement approach was unsuccessful because the 7 cysteine mutants which were engineered could not be successfully derivatized. Ultimately, molecular replacement was used to solve the structure by sorting through a large number of solutions in space group P1 using CNS.The main objective of this paper is to describe the obstacles which were faced and overcome in order to acquire data sets on such difficult crystals and determine phases for successful structure solution." @default.
• W2057264691 created "2016-06-24" @default.
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• W2057264691 date "2008-09-12" @default.
• W2057264691 modified "2023-09-24" @default.
• W2057264691 title "The Difficult Case of Crystallization and Structure Solution for the ParC55 Breakage-Reunion Domain of Topoisomerase IV from Streptococcus pneumoniae" @default.
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• W2057264691 doi "https://doi.org/10.1371/journal.pone.0003201" @default.
• W2057264691 hasPubMedCentralId "https://www.ncbi.nlm.nih.gov/pmc/articles/2527675" @default.
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