Computational protocol: Stapled BH3 Peptides against MCL-1: Mechanism and Design Using Atomistic Simulations

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Protocol publication

[…] The initial structure of MCL-1 bound to a stapled peptide was taken from the crystal structure 3MK8, resolved at 2.3Å . The missing residues (K194-R201) were modeled using Modeler 9.7 and guided by their positions in the NMR structure of MCL-1 bound to a peptide (PDB code 2KBW ). The starting model included residues 172–320 of human MCL-1, and residues 5–23 of the BH3 peptide . The stapled regions were modeled using the Xleap module of AMBER and the parameters were built using the antechamber module of AMBER , . Only the N- and C- termini of MCL-1 were capped (with acetyl or ACE and N-methyl or NME respectively) to keep them neutral, in accord with the experiments . Molecular dynamics simulations were performed with the SANDER module of the AMBER9 package employing the all-atom ff99SB force field . Simulations were carried out for the complexes of BH3 wild type and eleven stapled peptides bound to MCL-1 (). Each system was solvated with a TIP3P water box whose sides are at a minimum distance of 10 Å to any protein atom. Particle Mesh Ewald method (PME) was used for treating the long range electrostatics. All bonds involving hydrogen atoms were constrained by SHAKE . A time step of 2fs was used. Initially, the whole system was minimized for 4,000 steps, to remove any unfavorable interactions. Subsequently, the systems were each heated to 300 K for 30 ps under NPT conditions. After this, each system was equilibrated for 100 ps and then simulated for 20 ns at constant temperature (300 K) and pressure (1 atm) and structures were stored every 1 ps. The free energy of binding (ΔGbind) of the peptides to MCL-1 was computed using the MM-GBSA (molecular mechanics/Generalized Born surface area) method , using the GB module in Amber while the non-polar component was estimated from the solvent accessible surface area using MOLSURF with ΔGsolv,np = 0.00542*SASA +0.92 . Each energy term was averaged over frames taken every 2 ps over the last 10 ns of each simulation. Vibrational entropy was estimated using normal mode analysis (Nmode module of Amber) and averaged over 200 ps intervals. PyMOL and Visual Molecular Dynamics (VMD) were used for visualizations. […]

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