title: Two modes of interaction between the membrane-embedded TARP stargazin's C-terminal domain and the bilayer visualized by electron crystallography.
abstract: Glutamate-mediated neurotransmission through ligand-gated, ionotropic glutamate receptors is the main form of excitatory neurotransmission in the vertebrate central nervous system where it plays central roles in learning, memory and a variety of disorders. Acting as auxiliary subunits, transmembrane α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptor (AMPAR) regulatory proteins (TARPs) are essential regulators for glutamate-mediated neurotransmission in the central nervous system. Here, we report the first electron crystallographic reconstructions of full-length mouse stargazin (γ-2) at ∼20Å resolution in a membrane bilayer environment. Formation of ordered arrays required anionic lipids and was modulated by cholesterol and monovalent cations. Projection structures revealed that the C-termini of stargazin monomers closely interacted with the bilayer surface in an extended conformation that placed the C-terminal PDZ-binding motif ∼100Å away from the transmembrane domain and in close proximity to a membrane re-entrant region. The C-termini interaction with the bilayer was modulated by the ionic strength of the solution and overall protein secondary structure increased when membrane-bound. Our data suggest that stargazin interactions with and within the membrane play significant roles in TARP structure and directly visualize TARP functional mechanisms essential for AMPAR trafficking and clustering.
authors.affiliation: Department of Molecular Biophysics and Biochemistry, Yale University School of Medicine, New Haven, CT 06510, USA.
authors.forename: Matthew F
authors.initials: MF
authors.lastname: Roberts
authors.forename: David W
authors.initials: DW
authors.lastname: Taylor
authors.forename: Vinzenz M
authors.initials: VM
authors.lastname: Unger
mesh_terms: D000818:Animals; D000838:Anions / Q000737:chemistry; D015220:Calcium Channels / Q000737:chemistry* / Q000378:metabolism* / Q000648:ultrastructure; D002784:Cholesterol / Q000737:chemistry; D003461:Crystallography / Q000379:methods*; D019281:Dimerization; D021621:Imaging, Three-Dimensional; D008051:Lipid Bilayers / Q000737:chemistry* / Q000378:metabolism*; D008055:Lipids / Q000737:chemistry; D051379:Mice; D054731:PDZ Domains; D017434:Protein Structure, Tertiary; D018091:Receptors, AMPA / Q000737:chemistry / Q000378:metabolism
chemical_list: D000838:Anions; C113940:Cacng2 protein, mouse; D015220:Calcium Channels; D008051:Lipid Bilayers; D008055:Lipids; D018091:Receptors, AMPA; D002784:Cholesterol