From populations to absolute binding affinities in molecular simulations: exact volumetric terms and practical estimators

Abstract

We present a statistical-mechanics framework for computing equilibrium binding constants $K$ in the dilute limit. From first principles, we derive a general expression relating $K$ to the relative populations of the bound and unbound states. Its transparency has twofold advantage: it makes the origin of the unbound-state volumetric term explicit, and it allows one to track exactly how an imposed volume restraint propagates through the expression. This makes $K$ directly computable, as restrained simulations can account for the volumetric contribution exactly, under the physically mild assumption of a homogeneous unbound state. The resulting estimators are computable from histograms of any suitably defined reaction coordinate, and determine unambiguously how the boundaries of the thermodynamic states of interest must be defined. We apply our framework to the cucurbit[7]uril/1-adamantanol host--guest complex and the galactonate--DgoT ligand--protein complex. Our results show that commonly used single-bin estimators depart from the theoretically correct one by $\approx 1$~kcal/mol in both systems. This shift originates in the definition of the bound state: by anchoring that definition to what state-of-the-art experiments resolve, the theory turns it from a hidden assumption into a controlled input, and provides a principled route to absolute binding affinities from molecular simulations.