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[…] N) as separate domains and (ii) FimHPD (from PDB ID 1KLF) and FimHLD in the R state (from PDB ID 1KLF) as separate domains. The best solutions were identified as a T conformation for FimGNteH A62S and a bent R conformation (90° between FimHLD and FimHPD) for FimGNteH A27V/V163A. Several rounds of refinements were performed in phenix.refine to improve the final models., Crystal structures of FimGNteH published in this study along with models of FimGNteH based on previous crystal structures (GNte docked on H from CH structure) in the absence or presence of ligands were used to initiate MD simulations. Structural models of FimGNteH variants were created with the in silico mutagenesis wizard in PyMOL after the mutant side-chain rotamer with the fewest steric clashes was selected. Structural models were then prepared within the CHARMM36 force field and the TIP3P water model by the MD software package GROMACS-4.6.7 (). The system was first solvated in a cubic box, with sides equal to 130 Å in the presence of 50 mM NaCl. The structure was then energy-minimized to reduce steric clashes, and the system was finally equilibrated at constant temperature (37°C) and pressure (1 atm). Conventional unrestrained MD simulations were then performed with steps of 2 fs for a total time of ~15 to 20 ns for each run, collecting four replicates for simulations on the T (5JQI), bent R (5JR4), and elongated R (based on 1KLF) conformations for WT FimH. Longer simulations were also performed at 100 to 200 ns for three replicates for all FimH variants in these conformations with similar results. Coordinates wer […]

Pipeline specifications

Software tools PyMOL, CHARMM, GROMACS