Computational protocol: Structural insights into the functions of the FANCM-FAAP24 complex in DNA repair

Similar protocols

Protocol publication

[…] Crystallization was performed using the hanging drop vapor diffusion method at 20°C. Crystals of both native and Se-Met FANCM-FAAP24 were grown from drops consisting of 1 µl of protein solution (∼6 mg/ml) and 1 µl of reservoir solution containing 1.0 M (NH4)2SO4 and 0.1 M HEPES (pH 7.2). For diffraction data collection, the crystals were cryoprotected using the reservoir solution supplemented with 30% glycol and then flash-cooled into liquid nitrogen. Three Se-derivative datasets of 3.4 Å resolution and a native data set of 2.9 Å resolution were collected at 100 K at beamline 17 U of Shanghai Synchrotron Radiation Facility, China. The diffraction data were processed with HKL2000 (). The statistics of the diffraction data are summarized in . The structure of FANCM-FAAP24 was solved using the multi-wavelength anomalous dispersion method using Phenix (), which identified 7 Se atoms and yielded a figure of merit of 0.44. The structure model was refined against the native data set using Phenix () and Refmac5 (). Model building was performed using Coot (). The final structure model contains residues 1815–2030 of FANCM and residues 18–213 of FAAP24, except that three surface exposed loops (residues 1903–1915 and 1965–1969 of FANCM, and residues 147–152 of FAAP24) could not be modeled owing to poor electron density. Structure analysis was carried out using programs in CCP4 () and the PISA server (). All the structure figures were prepared using Pymol (http://www.pymol.org). The statistics of the structure refinement and the quality of the final structure model are also summarized in . […]

Pipeline specifications

Software tools PHENIX, REFMAC5, Coot, CCP4, PyMOL
Application Protein structure analysis