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• W2536706142 abstract "The plasmalemmal monoamine transporters clear the extracellular space from their cognate substrates and sustain cellular monoamine stores even during neuronal activity. In some instances, however, the transporters enter a substrate-exchange mode, which results in release of intracellular substrate. Understanding what determines the switch between these two transport modes demands time-resolved measurements of intracellular (co-)substrate binding and release. Here, we report an electrophysiological investigation of intracellular solute-binding to the human serotonin transporter (SERT) expressed in HEK-293 cells. We measured currents induced by rapid application of serotonin employing varying intracellular (co-)substrate concentrations and interpreted the data using kinetic modeling. Our measurements revealed that the induction of the substrate-exchange mode depends on both voltage and intracellular Na+ concentrations because intracellular Na+ release occurs before serotonin release and is highly electrogenic. This voltage dependence was blunted by electrogenic binding of intracellular K+ and, notably, also H+. In addition, our data suggest that Cl− is bound to SERT during the entire catalytic cycle. Our experiments, therefore, document an essential role of electrogenic binding of K+ or of H+ to the inward-facing conformation of SERT in (i) cancelling out the electrogenic nature of intracellular Na+ release and (ii) in selecting the forward-transport over the substrate-exchange mode. Finally, the kinetics of intracellular Na+ release and K+ (or H+) binding result in a voltage-independent rate-limiting step where SERT may return to the outward-facing state in a KCl- or HCl-bound form." @default.
• W2536706142 created "2016-10-28" @default.
• W2536706142 creator A5055412961 @default.
• W2536706142 creator A5060360807 @default.
• W2536706142 creator A5086541043 @default.
• W2536706142 date "2016-12-01" @default.
• W2536706142 modified "2023-10-16" @default.
• W2536706142 title "Electrogenic Binding of Intracellular Cations Defines a Kinetic Decision Point in the Transport Cycle of the Human Serotonin Transporter" @default.
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• W2536706142 doi "https://doi.org/10.1074/jbc.m116.753319" @default.
• W2536706142 hasPubMedCentralId "https://www.ncbi.nlm.nih.gov/pmc/articles/5207061" @default.
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