Computational protocol: Functional Role of Histidine in the Conserved His-x-Asp Motif in the Catalytic Core of Protein Kinases

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Protocol publication

[…] Co-crystallization of the proteins with adenosine was performed at 4 °C using the hanging drop vapor diffusion method. Bean-shaped crystals were grown in drops containing equal volumes (2 μL) of the protein mixture solution and the reservoir solution (0.1 M Bis-Tris, pH = 5.5, 0.2 M Ammonium sulfate, 25% w/v PEG 3350) to the maximum size in 1–2 days. Diffraction data were collected from flash-cooled crystals at −176 °C at beamline BL-17U in the Shanghai Synchrotron Radiation Facility (Shanghai, China) and processed with HKL2000 subsequently. The structure was solved with the molecular replacement method implemented in the program suite CCP4 by using the structure of the human Aurora A kinase (PDB code 1MQ4) as search model. The initial structure refinement was carried out with program Phenix and REFMAC5. Model building was performed manually with the program Coot. Throughout the refinement, 5% of randomly chosen reflections were set aside for calculating free R value. The final stereochemical quality of structural models was checked with MolProbity. A summary of structure refinement is listed in . […]

Pipeline specifications

Software tools CCP4, PHENIX, REFMAC5, Coot, MolProbity
Application Protein structure analysis
Diseases Neoplasms
Chemicals Tyrosine