Computational protocol: Crystal Structure and Functional Characterization of an Esterase (EaEST) from Exiguobacterium antarcticum

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Protocol publication

[…] A single crystal of EaEST was harvested and transferred to Pratone-N oil for cryoprotection. A data set containing 180 images at the resolution of 1.90 Å was collected on a 7A beam line at Pohang Accelerator Laboratory (PAL; Pohang, Korea) at 100K. The diffraction data were indexed, integrated, and scaled using the program HKL-2000 []. An EaEST crystal belongs to the trigonal space group P3, with unit cell parameters of a = 76.765 Å, b = 76.765 Å, c = 68.161 Å, and α = β = 90°, γ = 120°. The volume of the asymmetric unit allows the presence of two copies with a Matthews coefficient of 1.83 Å3Da-1 and a solvent content of 32.84% []. The structure of EaEST was determined by molecular replacement using the program MOLREP []. The coordinates of an esterase from P. fluorescens (PDB cold 3HI4) [], which has 39% identity to EaEST, was used as a model for molecular replacement. The resulting coordinate was rebuilt and refined manually based on electron-density maps using the programs REFMAC5 and COOT [, ]. After multiple rounds of structural refinement, the final structure of EaEST has the Rwork and Rfree of 0.185 and 0.238, respectively, with a total of 542 amino acid residues and 191 water molecules. The statistics of data collection and structure refinement are listed in . The atomic coordinates and structure factors have been deposited in the Protein Data Bank ( under accession code 5H3H. […]

Pipeline specifications

Software tools HKL-2000, Molrep, REFMAC5, Coot
Application Protein structure analysis
Organisms Exiguobacterium antarcticum