Molecular mechanism of ion-ion and ion-substrate coupling in the Na+-dependent leucine transporter LeuT.

ABSTRACT: Ion-coupled transport of neurotransmitter molecules by neurotransmitter:sodium symporters (NSS) play an important role in the regulation of neuronal signaling. One of the major events in the transport cycle is ion-substrate coupling and formation of the high-affinity occluded state with bound ions and substrate. Molecular mechanisms of ion-substrate coupling and the corresponding ion-substrate stoichiometry in NSS transporters has yet to be understood. The recent determination of a high-resolution structure for a bacterial homolog of Na(+)/Cl(-)-dependent neurotransmitter transporters, LeuT, offers a unique opportunity to analyze the functional roles of the multi-ion binding sites within the binding pocket. The binding pocket of LeuT contains two metal binding sites. The first ion in site NA1 is directly coupled to the bound substrate (Leu) with the second ion in the neighboring site (NA2) only approximately 7 A away. Extensive, fully atomistic, molecular dynamics, and free energy simulations of LeuT in an explicit lipid bilayer are performed to evaluate substrate-binding affinity as a function of the ion load (single versus double occupancy) and occupancy by specific monovalent cations. It was shown that double ion occupancy of the binding pocket is required to ensure substrate coupling to Na(+) and not to Li(+) or K(+) cations. Furthermore, it was found that presence of the ion in site NA2 is required for structural stability of the binding pocket as well as amplified selectivity for Na(+) in the case of double ion occupancy.

SUBMITTER: Caplan DA 

PROVIDER: S-EPMC2576368 | biostudies-literature | 2008 Nov

REPOSITORIES: biostudies-literature