Importance of the Extracellular Loop 4 in the Human Serotonin Transporter for Inhibitor Binding and Substrate Translocation.

Research output: Contribution to journal/Conference contribution in journal/Contribution to newspaperJournal articleResearchpeer-review

DOI

  • Hafsteinn Rannversson, Det Sundhedsvidenskabelige Fakultet, Københavns Universitet, København, Denmark
  • Pamela Wilson, Det Sundhedsvidenskabelige Fakultet, Københavns Universitet, København, Denmark
  • Kristina Kristensen, Denmark
  • Steffen Sinning
  • Anders Skov Kristensen, Det Sundhedsvidenskabelige Fakultet, Københavns Universitet, København, Denmark
  • Kristian Strømgaard, Det Sundhedsvidenskabelige Fakultet, Københavns Universitet, København, Denmark
  • Jacob Andersen, Det Sundhedsvidenskabelige Fakultet, Københavns Universitet, København, Denmark
The serotonin transporter (SERT) terminates serotonergic neurotransmission by performing reuptake of released serotonin, and SERT is the primary target for antidepressants. SERT mediates the reuptake of serotonin through an alternating access mechanism, implying that a central substrate site is connected to both sides of the membrane by permeation pathways, of which only one is accessible at a time. The coordinated conformational changes in SERT associated with substrate translocation are not fully understood. Here, we have identified a Leu to Glu mutation at position 406 (L406E) in the extracellular loop 4 (EL4) of human SERT, which induced a remarkable gain-of-potency (up to >40-fold) for a range of SERT inhibitors. The effects were highly specific for L406E relative to six other mutations in the same position, including the closely related L406D mutation, showing that the effects induced by L406E are not simply charge-related effects. Leu(406) is located >10 Å from the central inhibitor binding site indicating that the mutation affects inhibitor binding in an indirect manner. We found that L406E decreased accessibility to a residue in the cytoplasmic pathway. The shift in equilibrium to favor a more outward-facing conformation of SERT can explain the reduced turnover rate and increased association rate of inhibitor binding we found for L406E. Together, our findings show that EL4 allosterically can modulate inhibitor binding within the central binding site, and substantiates that EL4 has an important role in controlling the conformational equilibrium of human SERT.
Original languageEnglish
JournalJournal of Biological Chemistry
Volume290
Issue23
Pages (from-to)14582-94
Number of pages13
ISSN0021-9258
DOIs
Publication statusPublished - 5 Jul 2015

See relations at Aarhus University Citationformats

ID: 96564426