Computational protocol: Insights into the Importance of Hydrogen Bonding in the γ-Phosphate Binding Pocket of Myosin: Structural and Functional Studies of Serine 236†,‡

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Protocol publication

[…] X-ray data for the S236A·MgADP·VO4 and S236A·MgAMPPNP complexes were collected at the Advanced Photon Source (APS) in Argonne, IL, using a MAR-165 detector at COM-CAT 32-ID. A total of 280 frames, each with an oscillation range of 0.5°, were collected at 150 mm. The data were integrated and scaled with the HKL2000 program package (). The structures were determined by molecular replacement with the program Molrep () utilizing the structure of S1dC·MgADP·VO4 (PDB accession number 1VOM) () as the starting model. The structure was refined with Refmac5 in the CCP4 package, and water molecules were located and added using the model building program COOT (,). The conformations for AMPPNP and blebbistatin were obtained from coordinate sets 1MMN and 1YV3, respectively (,). The models were manually adjusted with the model building program COOT (,).Data for S236A·MgPPi were collected at APS using a ADSC Q315 315 mm × 315 mm mosaic CCD detector at SBC-CAT 19-ID. A total of 220 frames, each with an oscillation range of 1°, were collected at 180 mm. Integration and solution of the structure were performed as above with the exception that the starting model was that of S1dC·MgPPi (PDB accession code 1MNE) (). Data collection and refinement statistics are given in Table . […]

Pipeline specifications

Software tools Molrep, REFMAC5, Coot
Application Protein structure analysis
Organisms Dictyostelium discoideum
Chemicals Adenosine Triphosphate, Adenylyl Imidodiphosphate, Hydrogen, Hydroxyl Radical