Multi-Force-Field Molecular Dynamics Reveals How Cation Hydration Kinetics Dictate Polypeptide Assembly Pathways and Timescales.

Abstract

The distinct roles of Mg2+ and Ca2+ ions in biomolecular assembly can be understood through their different hydration dynamics, but a quantitative, causal link between the water-exchange kinetics of cations and assembly pathways has been lacking. Here, we establish this relationship using multi-force-field all-atom molecular dynamics simulations combined with enhanced sampling and Markov state model analysis. By systematically comparing force fields that inherently encode different hydration exchange rates, we show that the Mg2+ ion, with its kinetically inert hydration shell (microsecond water exchange), must undergo stepwise dehydration to coordinate aspartate residues. This kinetic barrier limits its ability to induce microphase separation on microsecond timescales, positioning the Mg2+ ion as a slow "structural reorganizer" that gradually disrupts preexisting Arg-Asp salt bridges. In contrast, the Ca2+ ion, with its rapidly exchanging hydration shell (picosecond-to-nanosecond water exchange), directly bridges multiple aspartate side-chains and acts as a fast "microphase separation trigger." Quantitative comparison of water-exchange rates across force fields, validated against experimental NMR data, establishes a causal chain from hydration kinetics to coordination modes to assembly timescales. Based on these results, we propose a "hydration clock" model in which the intrinsic water-exchange kinetics of cations dictates the fundamental timescales of biomolecular assembly. This work provides a kinetic framework, grounded in explicit-solvent molecular dynamics and multi-force-field validation, for understanding cation-specific effects and the rational design of time-programmable self-assembling systems.