Accurate calculation of affinity changes to the close state of influenza A M2 transmembrane domain in response to subtle structural changes of adamantyl amines using free energy perturbation methods in different lipid bilayers

Experimental binding free energies of 27 adamantyl amines against the influenza M2(22-46) WT tetramer, in its closed form at pH 8, were measured by ITC in DPC micelles. The measured K_d's range is ~44 while the antiviral potencies (IC₅₀) range is ~750 with a good correlation between binding free energies computed with K_d and IC₅₀ values (r = 0.76). We explored with MD simulations (ff19sb, CHARMM36m) the binding profile of complexes with strong, moderate and weak binders embedded in DMPC, DPPC, POPC or a viral mimetic membrane and using different experimental starting structures of M2. To predict accurately differences in binding free energy in response to subtle changes in the structure of the ligands, we performed 18 alchemical perturbative single topology FEP/MD NPT simulations (OPLS2005) using the BAR estimator (Desmond software) and 20 dual topology calculations TI/MD NVT simulations (ff19sb) using the MBAR estimator (Amber software) for adamantyl amines in complex with M2(22-46) WT in DMPC, DPPC, POPC. We observed that both methods with all lipids show a very good correlation between the experimental and calculated relative binding free energies (r = 0.77-0.87, mue = 0.36-0.92 kcal mol^-1) with the highest performance achieved with TI/MBAR and lowest performance with FEP/BAR in DMPC bilayers. When antiviral potencies are used instead of the K_d values for computing the experimental binding free energies we obtained also good performance with both FEP/BAR (r = 0.83, mue = 0.75 kcal mol^-1) and TI/MBAR (r = 0.69, mue = 0.77 kcal mol^-1).