The Substrate Import Mechanism of the Human Serotonin Transporter

02 October 2019, Version 1
This content is a preprint and has not undergone peer review at the time of posting.

Abstract

The serotonin transporter, SERT, initiates the reuptake of extracellular serotonin in the synapse to terminate neurotransmission. Recently, the cryo-EM structures of SERT bound to ibogaine resolved in different states provided the first glimpse of functional conformations at atomistic resolution. However, the conformational dynamics and structural transitions to various intermediate states are not fully understood. Furthermore, while experimental SERT structures were complexed with drug molecules and inhibitors, the molecular basis of how the physiological substrate, serotonin, is recognized, bound, and transported remains unclear. In this study, we performed microsecond long simulations of the human SERT to investigate the structural dynamics to various intermediate states and elucidated the complete substrate import pathway. Using Markov state models, we characterized a sequential order of conformational driven ion-coupled substrate binding and transport events and calculated the free energy barriers of conformation transitions associated with the import mechanism. We identified a set of crucial residues that recognize the substrate at the extracellular surface of SERT and our biochemical screening results show that mutations causes dramatic reduction in transport function. Our simulations also revealed a third sodium ion binding site coordinated by Glu136 and Glu508 in a buried cavity which helps maintain the conserved fold adjacent to the orthosteric site for transport function. The mutation of these residues results in a complete loss of transport activity. Our study provides novel insights on the molecular basis of dynamics driven ion-substrate recognition and transport of SERT that can serve as a model for other closely related neurotransmitter transporters.

Keywords

Neurotransmitter
Transporters
Membrane Protein
Markov State Models
Molecular Dynamics
serotonin transporter

Supplementary materials

Title
Description
Actions
Title
SI
Description
Actions

Comments

Comments are not moderated before they are posted, but they can be removed by the site moderators if they are found to be in contravention of our Commenting Policy [opens in a new tab] - please read this policy before you post. Comments should be used for scholarly discussion of the content in question. You can find more information about how to use the commenting feature here [opens in a new tab] .
This site is protected by reCAPTCHA and the Google Privacy Policy [opens in a new tab] and Terms of Service [opens in a new tab] apply.