Computational protocol: A cholesterol-binding domain in STIM1 modulates STIM1-Orai1 physical and functional interactions

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Protocol publication

[…] All-atom simulations were carried out with the CHARMM36 force field models for the protein, membrane and solvent environments. The initial coordinates of the monomeric peptide were generated using the available coordinates of the SOAR fragment from Homo sapiens (3TEQ), the protein orientation to the membrane was assigned using the OPM database and embedded in the corresponding lipid bilayers and 0.15 KCl ion concentrations using the CHARMM-GUI membrane builder. The final composition for the SOAR-POPC simulation consists of the protein fragment, 299 POPC lipids, 13,123 water molecules, 41 K+ and 44 Cl− atoms. For the 40% cholesterol simulations the system consists of 144 cholesterol molecules, 216 POPC lipids 14,383 water molecules, 43 K+ and 46 Cl− atoms.The system was energy minimized using steepest descent method and equilibrated at 310 K using NVT and NPT ensembles in which water, lipids, ions and the protein were constrained using a harmonic potential. At the final equilibration step all restraints were removed.For the 400 ns production runs periodic boundary conditions were employed, the short-range electrostatic interactions were cut-off at 1.2 nm and the full electrostatics were treated using PME algorithm. Van der Waal interactions were evaluated using a cutoff switch from 10 to 12 nm. To control the temperature and pressure at 310 K and 1 atm the Nosé-Hoover thermostat with a coupling constant of 1.0 ps and the Parrinello-Rahman barostat with a coupling constant of 1 ps and a compressibility of 4.5 × 10−5 bar −1 were employed using a semi-isotropic coupling for pressure control. The linear constraint solver (LINCS) method was used to constrain all bond lengths, and a 2 fs integration step was used. The simulations were carried out using GROMACS (v5.0.5) and the analyses were performed using python and the MDAnalysis libraries. All molecular images were created using VMD. […]

Pipeline specifications

Software tools CHARMM, CHARMM-GUI Membrane Builder, P-LINCS, GROMACS, MDAnalysis, VMD
Applications Protein structure analysis, Membrane protein analysis
Chemicals Calcium, Cholesterol