) for samples drawn from the equilibrium distribution π(x; λ = 0) and vice versa for samples drawn from the equilibrium distribution π(x; λ = 1). Free energy estimates were obtained using the EXP (exponential averaging) free energy estimator and the BAR (Bennett's acceptance ratio) estimator; both were used as implemented in the pymbar package. To estimate uncertainties for FEP and NEQ free energy estimates, the 90% confidence interval is calculated using bootstrapping. For the MFES estimate, we calculate the 90% confidence interval by calculating the standard deviation of the three independent calculations and using the t-distribution.