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TitleEfficient and rapid isolation of native AMPA receptor complexes for cryo-EM.
Journal, issue, pagesProtein Sci, Vol. 35, Issue 2, Page e70483, Year 2026
Publish dateJan 24, 2026
AuthorsJumi Park / Eric Gouaux /
PubMed AbstractIsolating native ion channels for structural characterization is routinely achieved by extraction from membrane fractions of tissue with prolonged mild detergent treatment. AMPA receptors (AMPARs), ...Isolating native ion channels for structural characterization is routinely achieved by extraction from membrane fractions of tissue with prolonged mild detergent treatment. AMPA receptors (AMPARs), glutamatergic receptors that mediate fast excitatory transmission and synaptic plasticity, are coassembled with diverse auxiliary subunits and transiently-interacting partners to finely regulate processes from trafficking to gating kinetics. Previous studies of the composition and architecture of native AMPARs (nAMPARs) isolated from membrane fractions of rodent brain tissue have revealed many different subunit compositions and non-stochastic assemblies of the auxiliary subunits. However, elucidating the molecular architectures of nAMPARs complexed with less populated or transiently bound proteins has proven challenging. Here, we employ strategies for the rapid solubilization and purification of nAMPARs to increase the likelihood of isolating the greatest range of nAMPARs complexes. By utilizing whole brain tissue and reducing solubilization and purification duration, we purify nAMPARs complexed with a wider variety of auxiliary subunits and binding partners in a sufficient quantity and purity for cryo-electron microscopy studies. We resolve previously unreported subunit compositions and conformations that include ones with a half-splayed ATD layer, as well as complexes with four distinct auxiliary subunit arrangements in the TMD layer.
External linksProtein Sci / PubMed:41578975 / PubMed Central
MethodsEM (single particle)
Resolution2.87 - 11.73 Å
Structure data

EMDB-73846: Cryo-EM map of native AMPA receptor A1A2A1A2 subtype
Method: EM (single particle) / Resolution: 3.34 Å

EMDB-73847: Cryo-EM map of native AMPA receptor A3A2A3A2 subtype
Method: EM (single particle) / Resolution: 3.32 Å

EMDB-73848: Cryo-EM map of native AMPA receptor A1A2A3A2 (AS1) subtype
Method: EM (single particle) / Resolution: 3.42 Å

EMDB-73849: Cryo-EM map of native AMPA receptor A1A2A3A2 (AS2) subtype
Method: EM (single particle) / Resolution: 3.38 Å

EMDB-73850: Cryo-EM map of native AMPA receptor AxAxA1A2 subtype
Method: EM (single particle) / Resolution: 8.41 Å

EMDB-73851: Cryo-EM map of native AMPA receptor AxAxA3A2 subtype
Method: EM (single particle) / Resolution: 11.64 Å

EMDB-73852: Cryo-EM map of native AMPA receptor A1A2AxA2 (AS1) subtype
Method: EM (single particle) / Resolution: 8.08 Å

EMDB-73853: Cryo-EM map of native AMPA receptor A1A2AxA2 (AS2) subtype
Method: EM (single particle) / Resolution: 11.73 Å

EMDB-73854: Cryo-EM map of native AMPA receptor A3A2A2A2 (AS1) subtype
Method: EM (single particle) / Resolution: 7.38 Å

EMDB-73855: Cryo-EM map of native AMPA receptor A3A2A2A2 (AS2) subtype
Method: EM (single particle) / Resolution: 8.6 Å

EMDB-73856: Cryo-EM map of native AMPA receptor A3A2AxA2 subtype
Method: EM (single particle) / Resolution: 7.69 Å

EMDB-73858: Cryo-EM map of native AMPA receptor A1A2A2A2 (AS1) subtype
Method: EM (single particle) / Resolution: 8.77 Å

EMDB-73859: Cryo-EM map of native AMPA receptor A1A2A2A2 (AS2) subtype
Method: EM (single particle) / Resolution: 9.64 Å

EMDB-73860: Cryo-EM map of TMD with 2 TARPs from all native AMPA receptor subtypes
Method: EM (single particle) / Resolution: 3.55 Å

EMDB-73861, PDB-9z6u:
The structure of TMD with 2 TARPs and 2 CNIHs from all native AMPA receptor subtypes
Method: EM (single particle) / Resolution: 2.87 Å

EMDB-73862, PDB-9z6v:
The structure of TMD with 3 TARPs and 1 CNIH from all native AMPA receptor subtypes
Method: EM (single particle) / Resolution: 3.0 Å

EMDB-73863, PDB-9z6w:
The structure of TMD with 4 TARPs from all native AMPA receptor subtypes
Method: EM (single particle) / Resolution: 2.99 Å

Chemicals

ChemComp-POV:
(2S)-3-(hexadecanoyloxy)-2-[(9Z)-octadec-9-enoyloxy]propyl 2-(trimethylammonio)ethyl phosphate / phospholipid*YM

ChemComp-C14:
TETRADECANE

ChemComp-XVD:
6-[2-chloro-6-(trifluoromethoxy)phenyl]-1H-benzimidazol-2-ol

ChemComp-D12:
DODECANE

ChemComp-OCT:
N-OCTANE

Source
  • mus musculus (house mouse)
KeywordsMEMBRANE PROTEIN / Ionotropic glutamate receptor AMPA receptor

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