Glutamate binding and conformational flexibility of ligand-binding domains are critical early determinants of efficient kainate receptor biogenesis

Martin B. Gill, Pompun Vivithanaporn, Geoffrey T. Swanson*

*Corresponding author for this work

Research output: Contribution to journalArticle

24 Scopus citations

Abstract

Intracellular glutamate binding within the endoplasmic reticulum (ER) is thought to be necessary for plasma membrane expression of ionotropic glutamate receptors. Here we determined the importance of glutamate binding to folding and assembly of soluble ligand-binding domains (LBDs), as well as full-length receptors, by comparing the secretion of a soluble GluR6-S1S2 protein versus the plasma membrane localization of GluR6 kainate receptors following mutagenesis of the LBD. The mutations were designed to either eliminate glutamate binding, thereby trapping the bilobate LBD in an "open" conformation, or "lock" the LBD in a closed conformation with an engineered interdomain disulfide bridge. Analysis of plasma membrane localization, medium secretion of soluble LBD proteins, and measures of folding efficiency suggested that loss of glutamate binding affinity significantly impacted subunit protein folding and assembly. In contrast, receptors with conformationally restricted LBDs also exhibited decreased PM expression and altered oligomeric receptor assembly but did not exhibit any deficits in subunit folding. Secretion of the closed LBD protein was enhanced compared with wild-type GluR6-S1S2. Our results suggest that glutamate acts as a chaperone molecule for appropriate folding of nascent receptors and that relaxation of LBDs from fully closed states during oligomerization represents a critical transition that necessarily engages other determinants within receptor dimers. Glutamate receptor LBDs therefore must access multiple conformations for efficient biogenesis.

Original languageEnglish (US)
Pages (from-to)14503-14512
Number of pages10
JournalJournal of Biological Chemistry
Volume284
Issue number21
DOIs
StatePublished - May 22 2009

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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