If the binding modes are known, the problem of crossing energy barriers becomes more tractable. Relative alchemical free energy perturbation (AFE) simulations can sometimes be used in such cases to find out the free energy difference between two binding modes. These calculations simulate a series of separate intermediate states to transform one binding mode into another by disappearing atoms from one region and growing them in another region. The resulting free energy difference can be computed via a variety of perturbationbased estimators such as the multistate Bennett acceptance-ratio method (MBAR), or via alternative formulations like thermodynamic integration (TI). The binding modes need to be very similar to each other for relative AFE calculations.