Computational protocol: The PHCCEx domain of Tiam1/2 is a novel protein- and membrane-binding module

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Protocol publication

[…] Using data from the Tiam2 P4122/P4322 crystals, the positions of 19 selenium sites were determined using SHELX-D (). Phasing and density modification were carried out using SHARP () and SOLOMON (), respectively. Model building was performed from the obtained electron density map using the graphic program COOT (). The structure was refined by the simulated annealing method using the program CNS () to 3.2-Å resolution. The model was used for structural determination of the Tiam2 P21 crystal by the molecular replacement method using the program MOLREP (). The structure was refined using CNS to 2.1-Å resolution (). The crystal contains four Tiam2 PH–CC–Ex regions in the asymmetric units and the current structure includes 920 residues (232 residues of Mol_1, 231 of Mol_2, 229 of Mol_3 and 228 of Mol_4). No models were built for β3–β4 loop residues (6, 8, 9 and 12 residues for Mol_1, 2, 3 and 4, respectively), which were not observed in the electron density map. This analysis revealed no residual electron density that corresponded to the CD44 peptide. The Tiam1 PH–CC–Ex structure was determined by molecular replacement using MOLREP with the Tiam2 structure used as a search model. The structure was refined by the simulated annealing method using the program CNS to 4.5-Å resolution. […]

Pipeline specifications

Software tools Coot, CNS, Molrep
Application Protein structure analysis
Chemicals Phosphatidylinositols, N-Methylaspartate