Unraveling Copper Exchange in the Atox1-Cu(I)-Mnk1 Heterodimer: A Simulation Approach.

Copper, an essential metal for various cellular processes, requires tight regulation to prevent cytotoxicity. Intracellular pathways crucial for maintaining optimal copper levels involve soluble and membrane transporters, namely, metallochaperones and -type ATPases, respectively. In this study, we used a simulation workflow based on free-energy perturbation (FEP) theory and parallel bias metadynamics (PBMetaD) to predict the Cu(I) exchange mechanism between the human Cu(I) chaperone, Atox1, and one of its two physiological partners, ATP7A. ATP7A, also known as the Menkes disease protein, is a transmembrane protein and one of the main copper-transporting ATPases. It pumps copper into the trans-Golgi network for the maturation of cuproenzymes and is also essential for the efflux of excess copper across the plasma membrane. In this analysis, we utilized the nuclear magnetic resonance (NMR) structure of the Cu(I)-mediated complex between Atox1 and the first soluble domain of the Menkes protein (Mnk1) as a starting point. Independent free-energy simulations were conducted to investigate the dissociation of both Atox1 and Mnk1. The calculations revealed that the two dissociations require free energy values of 6.3 and 6.2 kcal/mol, respectively, following a stepwise dissociation mechanism.