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Title: Conformational Dynamics on the Extracellular Side of LeuT Controlled by Na + and K + Ions and the Protonation State of Glu 290

Journal Article · · Journal of Biological Chemistry
 [1];  [2];  [3];  [4];  [2];  [2];  [1]
  1. Cornell Univ., Ithaca, NY (United States)
  2. Univ. of Copenhagen (Denmark)
  3. Cornell Univ., Ithaca, NY (United States); National Inst. of Health (NIH), Baltimore, MD (United States)
  4. Columbia Univ., New York, NY (United States); New York State Psychiatric Inst., New York, NY (United States)
+ Show Author Affiliations

Ions play key mechanistic roles in the gating dynamics of neurotransmitter:sodium symporters (NSSs). In other microsecond scale molecular dynamics simulations of a complete model of the dopamine transporter, a NSS protein, we observed a partitioning of K+ ions from the intracellular side toward the unoccupied Na2 site of dopamine transporter following the release of the Na2-bound Na+. In this work, we evaluate with computational simulations and experimental measurements of ion affinities under corresponding conditions, the consequences of K+ binding in the Na2 site of LeuT, a bacterial homolog of NSS, when both Na+ ions and substrate have left, and the transporter prepares for a new cycle. We compare the results with the consequences of binding Na+ in the same apo system. Analysis of >50-μs atomistic molecular dynamics and enhanced sampling trajectories of constructs with Glu290, either charged or neutral, point to the Glu290 protonation state as a main determinant in the structural reconfiguration of the extracellular vestibule of LeuT in which a “water gate” opens through coordinated motions of residues Leu25, Tyr108, and Phe253. The resulting water channel allows the binding/dissociation of the Na+ and K+ ions that are prevalent, respectively, in the extracellular and intracellular environments.

Research Organization:
Oak Ridge National Laboratory (ORNL), Oak Ridge, TN (United States). Oak Ridge Leadership Computing Facility (OLCF)
Sponsoring Organization:
USDOE Office of Science (SC)
Grant/Contract Number:
AC05-00OR22725
OSTI ID:
1565483
Journal Information:
Journal of Biological Chemistry, Vol. 291, Issue 38; ISSN 0021-9258
Publisher:
American Society for Biochemistry and Molecular BiologyCopyright Statement
Country of Publication:
United States
Language:
English
Citation Metrics:
Cited by: 16 works
Citation information provided by
Web of Science

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Cited By (7)

A partially-open inward-facing intermediate conformation of LeuT is associated with Na+ release and substrate transport journal January 2018
Conformation space of a heterodimeric ABC exporter under turnover conditions journal July 2019
Locking Two Rigid-body Bundles in an Outward-Facing Conformation: The Ion-coupling Mechanism in a LeuT-fold Transporter journal December 2019
A Markov State-based Quantitative Kinetic Model of Sodium Release from the Dopamine Transporter journal January 2017
A kinetic account for amphetamine-induced monoamine release journal February 2018
X-ray structure of LeuT in an inward-facing occluded conformation reveals mechanism of substrate release journal February 2020
Electrogenic Binding of Intracellular Cations Defines a Kinetic Decision Point in the Transport Cycle of the Human Serotonin Transporter journal December 2016

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Related Subjects

59 BASIC BIOLOGICAL SCIENCES
conformational change
membrane protein
molecular dynamics
monoamine transporter
neurotransmitter transport