Computational protocol: Structure of a translocation signal domain mediating conjugative transfer by type IV secretion systems

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[…] The SeMet-containing protein crystals of TSAΔcryst was used for phasing. The data sets of the SeMet-TSA and native-TSA were collected at beamline ID14-2 at the European Synchrotron Radiation Facility (ESRF). A data set was collected at the wavelength of 0.8726 Å, and processed using the XDS suite. The unmerged HKL file was converted into an mtz file using POINTLESS. REINDEX and SCALA (Winn et al., ) were used for scaling and separating the anomalous pairs. The unmerged intensities were then converted into merged amplitudes using C-TRUNCATE (Winn et al., ). Heavy atoms search and SAD phases calculations were carried out using PHASER (McCoy et al., ). Phases were further improved by solvent flattening using DM (Winn et al., ). The solvent flattened phases were used for model building using BUCCANEER (Winn et al., ). Building of the initial model was completed manually using COOT. Restrained refinement and one round of TLS refinement was performed using REFMAC (Winn et al., ). The model was manually inspected and corrected in COOT using 2Fo-Fc maps (Emsley and Cowtan, ). The final model was validated using PROCHECK (Winn et al., ). […]

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