Biogenic amine transporters play important roles in the action and recycling of their specific neurotransmitter substrates. They have been well established as the targets for many pharmacological agents that affect brain function. For examples, the rewarding properties ofthe abused drugs, cocaine and amphetamine, are due mainly to its inhibition of dopamine transporter (DAT); the antidepressant effect of imipramine is exerted on serotonin transporter (SERT). These transporters belong to the Neurotransmitter:Sodium Symporter (NSS) family, which has a large number of prokaryotic homologs. A prokaryotic leucine transporter (LeuT) structure has been solved in high resolution, with the substrate Leu bound in an enclosed cavity. However, the determinants of substrate specificity, an understanding of which is important for rational drug design, may lie not only within the binding site crevice revealed by the LeuT structure, but also along the permeation pathway. The long term goal of the present proposal is to elucidate dynamic structural changes of the permeation pathway of NSS family proteins during the translocation cycle and to evaluate competing transport models, e. g., the alternating-access scheme and our two-substrate model. Specifically, using prokaryotic NSS-proteins as model systems, this will be achieved by an integrated, comparative process of iteration between molecular modeling simulations and experimental investigations/validations. With this multidisciplinary protocol we will explore the conserved residue positions and potential auxiliary cavities that line the permeation pathway and reorganize dynamically in different conformational states. The common, and the different features of the permeation pathways, including the roles of sodium ions, will be compared among prokaryotic and eukaryotic NSS-proteins. The knowledge acquired should shed light on the translocation cycles of biogenic amine transporters and will contribute to our understanding of the structural bases of substrate specificities.

Public Health Relevance

Neurotransmitter:Sodium Symporter proteins include many important therapeutic targets, such as those for abused drugs (cocaine and amphetamine) and antidepressants. The molecular mechanism revealed by the proposed study would specifically identify the location(s) and manner that these compounds exert effects on the target proteins and facilitate the rational drug design.

National Institute of Health (NIH)
National Institute on Drug Abuse (NIDA)
Research Transition Award (R00)
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Special Emphasis Panel (NSS)
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Wu, Da-Yu
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Weill Medical College of Cornell University
Schools of Medicine
New York
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Stolzenberg, Sebastian; Li, Zheng; Quick, Matthias et al. (2017) The role of transmembrane segment 5 (TM5) in Na2 release and the conformational transition of neurotransmitter:sodium symporters toward the inward-open state. J Biol Chem 292:7372-7384
Quick, Matthias; Shi, Lei (2015) The sodium/multivitamin transporter: a multipotent system with therapeutic implications. Vitam Horm 98:63-100
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Michino, Mayako; Donthamsetti, Prashant; Beuming, Thijs et al. (2013) A single glycine in extracellular loop 1 is the critical determinant for pharmacological specificity of dopamine D2 and D3 receptors. Mol Pharmacol 84:854-64
Kantcheva, Adriana K; Quick, Matthias; Shi, Lei et al. (2013) Chloride binding site of neurotransmitter sodium symporters. Proc Natl Acad Sci U S A 110:8489-94
Mondal, Sayan; Khelashvili, George; Shi, Lei et al. (2013) The cost of living in the membrane: a case study of hydrophobic mismatch for the multi-segment protein LeuT. Chem Phys Lipids 169:27-38
Quick, Matthias; Shi, Lei; Zehnpfennig, Britta et al. (2012) Experimental conditions can obscure the second high-affinity site in LeuT. Nat Struct Mol Biol 19:207-11

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