Computational protocol: Structure of l-rhamnose isomerase in complex with l-rhamnopyranose demonstrates the sugar-ring opening mechanism and the role of a substrate sub-binding site☆

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Protocol publication

[…] Crystals were flash-cooled in liquid nitrogen at 100 K and X-ray diffraction data were collected on PF-AR NE3A and NW12A beam lines in the Photon Factory (Tsukuba, Japan), and the BL26B1 beam line in SPring-8. Diffraction data were processed using the program HKL2000 [] and CCP4 program suite []. Data collection statistics and scaling results are listed in . Initial phases were determined by a molecular replacement method with the program MOLREP [] in the CCP4 program suite, using the structure of the wild-type P. stutzeri l-RhI (PDB code 2HCV) as a probe model []. Further model building was performed with the programs Coot [], in the CCP4 program suite, and X-fit [], in the XtalView program system [], and the structure was refined using the programs Refmac5 [] and CNS []. Water molecules were gradually introduced if peaks above 3.5σ in the (Fo− Fc) electron density map were in the range of a hydrogen bond. The number of residues in the most favored regions of a Ramachandran plot [] was determined by the program PROCHECK []. Refinement statistics are listed in . were illustrated by the program PyMol []. […]

Pipeline specifications

Software tools CCP4, Molrep, Coot, REFMAC5, CNS, PROCHECK, PyMOL
Applications Small-angle scattering, Protein structure analysis
Organisms Dipturus trachyderma