Computational protocol: Molecular Dynamics Simulation Reveals Insights into the Mechanism of Unfolding by the A130T/V Mutations within the MID1 Zinc-Binding Bbox1 Domain

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Protocol publication

[…] All simulations on the native MID1 Bbox1 domain and specific mutations were performed using the GROMACS software package []. The calculations of the native Bbox1 domain used model 1 of the NMR ensemble of structures (PDB code 2FFW) []. The mutant structures were built via homology modeling by changing residue A130 to threonine or valine using I-TASSER []. For the simulation, the Amber03 force field [] and explicit solvent based on the TIP3P model were employed []. The structures were solvated with explicit water in the periodic rectangular boxes under normal (150 mM) saline condition. The LINCS algorithm was used to constrain all bond lengths []. Long-range electrostatic interactions were treated with the particle-mesh Ewald method []. For the MD calculations, the non-bonded (electrostatic and VDW) cutoff range was 8 Å and the time step was 2 fs.Before each MD simulation, the entire system was first minimized by a 1000-step steepest descent calculation followed by a 3000-step of conjugate gradient optimization. We performed three 100 ns-trajectories at 300K for each of the native and two mutant structures, for a total 900ns computer simulation time. The standard deviation (SD) of backbone RMSD among the three repeated MD trajectories for the native, and the A130T/V mutants are 0.2Å, 0.7Å and 0.8Å, respectively. The small SD values (<1Å) indicate that the simulated trajectories are consistent. The pattern of the secondary structure elements were calculated using standard DSSP program []. The structures were visualized and analyzed by VMD and PyMOL []. The pKa values for residue C142 were calculated by PROPKA [,]. The root mean square deviation (RMSD), root mean square fluctuation (RMSF), and angles were carried out by using the gromacs inbuilt tools g_rms, g_rmsf and g_sgangle, respectively. […]

Pipeline specifications

Application Protein structure analysis
Diseases Genetic Diseases, Inborn
Chemicals Cysteine, Zinc