where G complex , G receptor and G ligand are the average values of the Gibbs free energy for the complex, receptor (protein) and ligand respectively. Sampling of the complex and its two components can be performed independently or conformations of the receptor and ligand derived from simulation of the complex. The latter approach is more commonly used due to its improved convergence behaviour, a consequence of cancellation between the noisy terms describing the internal energy of the ligand, receptor and complex . However, recent work has indicated that adaptation energies associated with confining the receptor and ligand in a complex can differ significantly even for closely related complexes . Here we investigate a range of ESMACS protocols incorporating different component sampling strategies. When both the receptor and ligand contributions are computed from the complex trajectory we designate this a "1traj protocol". When all three derive from independent trajectories we refer to this as the "3traj protocol" and when only one or other of the receptor or ligand contributions do so a "2traj protocol". A suffix (either -fl or -fr, for flexible ligand and receptor respectively) is added to the protocol name to signify which component is derived from the independent simulation. Additional variants involve the use of the average receptor contribution across the complex simulations for all comparable ligands, which is indicated with an -ar (averaged receptor) suffix in the protocol name. A summary of all of the protocols, describing from which simulation component data is obtained, is given in Table . It should be noticed that the statistical performance of the pair of protocols 1traj-ar and 2traj-fr, and 2traj-ar and 3traj are the same as the receptor contribution in all cases is a constant. Consequently, we do not analyze the 3traj or 2traj-fr protocols explicitly.