Molecular Mechanism of Chiral Recognition of R- and S-Methamphetamine by the Human Dopamine Transporter: Insights...

Methamphetamine (METH) exerts its stimulant and addictive effects primarily through interactions with the human dopamine transporter (hDAT), yet the molecular origins of the pronounced stereoselectivity between its S- and R-enantiomers remain poorly understood. Here, we employed all-atom molecular dynamics simulations combined with free-energy calculations and kinetic analyses to elucidate the atomic-level mechanisms underlying the stereoselective recognition of METH by hDAT. Our results reveal that S-METH binds more strongly and forms more stable interaction networks with outward-facing and occluded conformations of hDAT than R-METH. Kinetic analyses further show that S-METH associates with the outward-facing transporter more rapidly than R-METH. Importantly, S-METH promotes a thermodynamically favorable and kinetically accelerated transition from the outward-facing to the occluded state while simultaneously increasing the free-energy barrier and slowing the transition toward the inward-facing state. This dual effect effectively traps hDAT in a long-lived occluded state. In contrast, R-METH exhibits weaker binding interactions and less pronounced modulation of transporter conformational dynamics. Together, these findings provide a comprehensive thermodynamic and kinetic framework for understanding the chiral selectivity of hDAT toward METH enantiomers and offer mechanistic insight into the markedly different pharmacological potencies of S- and R-METH. The mechanistic principles revealed here may inform the rational design of dopamine transporter modulators with optimized binding thermodynamics and conformational kinetic profiles.