Title

Dopamine Transporter Amino and Carboxyl Termini Synergistically Contribute to Substrate and Inhibitor Affinities

UMMS Affiliation

Brudnick Neuropsychiatric Research Institute, Department of Psychiatry

Date

1-27-2017

Document Type

Article

Disciplines

Biochemistry | Mental and Social Health | Neuroscience and Neurobiology | Psychiatry | Psychiatry and Psychology

Abstract

Extracellular dopamine and serotonin concentrations are determined by the presynaptic dopamine (DAT) and serotonin (SERT) transporters, respectively. Numerous studies have investigated the DAT and SERT structural elements contributing to inhibitor and substrate binding. To date, crystallographic studies have focused on conserved transmembrane domains, where multiple substrate binding and translocation features are conserved. However, it is unknown what, if any, role the highly divergent intracellular N and C termini contribute to these processes. Here, we used chimeric proteins to test whether DAT and SERT N and C termini contribute to transporter substrate and inhibitor affinities. Replacing the DAT N terminus with that of SERT had no effect on DA transport Vmax but significantly decreased DAT substrate affinities for DA and amphetamine. Similar losses in uptake inhibition were observed for small DAT inhibitors, whereas substituting the DAT C terminus with that of SERT affected neither substrate nor inhibitor affinities. In contrast, the N-terminal substitution was completely tolerated by the larger DAT inhibitors, which exhibited no loss in apparent affinity. Remarkably, all affinity losses were rescued in DAT chimeras encoding both SERT N and C termini. The sensitivity to amino-terminal substitution was specific for DAT, because replacing the SERT N and/or C termini affected neither substrate nor inhibitor affinities. Taken together, these findings provide compelling experimental evidence that DAT N and C termini synergistically contribute to substrate and inhibitor affinities.

Rights and Permissions

Citation:J Biol Chem. 2017 Jan 27;292(4):1302-1309. doi: 10.1074/jbc.M116.762872. Epub 2016 Dec 16. Link to article on publisher's site.

Related Resources

Link to Article in PubMed

Keywords

amphetamine, cocaine, dopamine transporter, kinetics, monoamine transporter, psychostimulants, reuptake, selective serotonin reuptake inhibitor (SSRI), serotonin transporter

PubMed ID

27986813