Computational protocol: Identification and characterization of smallest pore-forming protein in the cell wall of pathogenic Corynebacterium urealyticum DSM 7109

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Protocol publication

[…] Initial models of hexamer and octamer channels were constructed using the CCBuilder tool [] without prior structural information. Both channels were simulated in a POPE bilayer containing a 1 M KCl solution (hexamer system about 66.000 atoms, octamer system about 90.000 atoms) using GROMACS 5.1.2 [] and the CHARMM36 force field [, ]. Long-range electrostatic potentials beyond the cut-off of 12 Å were computed using the particle mesh Ewald algorithm []. Lennard-Jones potentials were computed using the 10–12 Å force-switch scheme. The systems were equilibrated for 16 ns following the protocol described in the CHARMM-GUI Membrane Builder []. Unbiased production runs of 500 ns each were performed for the two systems in the pressure ensemble. The Nosé-Hoover thermostat [] was used to maintain a constant temperature of 303.15 K by coupling the protein, lipid and solvent separately to a temperature bath using a coupling constant of 1 ps. Moreover, the pressure was maintained constant at 1 bar by using the Parrinello-Rahman barostat [] with semi-isotropic scaling with a coupling constant of 5 ps. Covalent bonds containing hydrogen atoms were constrained using the LINCS algorithm [] to be able to use an integration time step of 2 fs. Furthermore, two simulations of the octamer system were performed in the presence of an applied voltage of 100 mV to determine the ion conductance at 1 M KCl. The membrane potential (V = E/Lz) was modelled by applying a constant electric field (E) perpendicular to the membrane [, ]. […]

Pipeline specifications

Software tools CCBuilder, GROMACS, CHARMM-GUI Membrane Builder, P-LINCS
Applications Protein structure analysis, Membrane protein analysis
Organisms Corynebacterium urealyticum, Corynebacterium glutamicum, Escherichia coli