| Gene (Rattus norvegicus) |
GRIK1-1a, GRIK1-2a, Neto1, Neto2 |
This paper |
|
GRIK1 was used with mutations in the TM1 region to improve protein expression and stability |
| Strain, strain background (Escherichia coli) |
BL21(DE3) |
Sigma-Aldrich |
CMC0016 |
competent cells |
| Cell line (Homo sapiens) |
HEK293 GnTI- suspension-adapted cells |
ATCC |
CRL-3022 |
Used for expression of GluK1-1aEM for large-scale purification |
| Cell line (Homo sapiens) |
HEK293 WT cells |
ATCC |
CRL-1573 |
Used for whole-cell patch-clamp electrophysiology |
| Cell line (Homo sapiens) |
HEK293-T/17 cells |
ATCC |
CRL-11268 |
Used for outside-out patch-clamp electrophysiology |
| Antibody (Rabbit monoclonal) |
Anti-His monoclonal antibody |
Cell Signaling Technology |
Cat. No. 12698 |
Used for co-immunoprecipitation/Western Blotting |
| Antibody (Rabbit polyclonal) |
Anti-Neto1 polyclonal antibody |
Sigma-Aldrich |
SAB3500679 |
Used for co-immunoprecipitation/Western Blotting |
| Antibody (Mouse Monoclonal) |
Anti-GFP |
Sigma-Aldrich |
G1546 |
Used for co-immunoprecipitation/Western Blotting |
| Antibody (Mouse Monoclonal) |
Anti-Actin |
Sigma-Aldrich |
A3853 |
Used for co-immunoprecipitation/Western Blotting |
| Chemical compound |
SYM2081 (2 S, 4R-4-methyl glutamate) |
Tocris Bioscience |
31137-74-3 |
Used to stabilize the receptor and electrophysiology experiments |
| Chemical compound |
UBP301 |
Tocris Bioscience |
569371-10-4 |
Used to stabilize the receptor and electrophysiology experiments |
| Chemical compound |
Kainic acid |
Tocris Bioscience |
487-79-6 |
Used to stabilize the receptor and electrophysiology experiments |
| Chemical compound |
L-Glutamic acid |
Sigma-Aldrich |
49449 |
Used to stabilize the receptor and electrophysiology experiments |
| Chemical compound |
Sodium Butyrate |
Sigma-Aldrich |
8.17500 |
Added to boost protein production |
| Commercial assay kit |
Bio-Beads SM-2 |
Bio-Rad |
1523920 |
Used for detergent removal during nanodisc reconstitution |
| Recombinant DNA reagent |
pEGBacMam vector |
Eric Gouaux's lab (shared) |
|
Used for protein expression |
| Recombinant DNA reagent |
GluK1-1a and GluK1-2a in pRK7 vector |
Mark Mayer’s lab (shared) |
|
Used for electrophysiology experiments |
| Software algorithm |
cryoSPARCv3 |
Nature Methods Punjani et al., 2017
|
DOI:10.1038/nmeth.4169
|
Used for single-particle data processing |
| Software algorithm |
UCSF Motioncor2 |
Nature Methods Zheng et al., 2017
|
DOI 10.1038/nmeth.4193
|
Used for motion correction of cryo-EM data |
| Software, algorithm |
UCSF ChimeraX |
Goddard et al., 2018
|
RRID:SCR_015872
|
Molecular |
| Software, algorithm |
UCSF Chimera |
Pettersen et al., 2004
|
RRID:SCR_004097
|
Molecular |
| Software, algorithm |
Coot |
Emsley and Cowtan, 2004
|
RRID:SCR_014222
|
Protein Model |
| Software, algorithm |
Phenix |
Adams et al., 2010
|
RRID:SCR_014224
|
Protein Model |
| Software, algorithm |
Clampfit |
Molecular Devices |
11.2 |
Electrophysiology data analysis |
| Software, algorithm |
Fitmaster |
HEKA Elektronik |
v2x90.4 |
Electrophysiology data analysis |
| Software, algorithm |
Dotmatics |
GraphPad Prism |
version 8.0.1 |
Used for statistical analysis and graphs/plots |
| Sequence-based reagent |
Primers |
This paper |
PCR primers |
Sequences given in Figure 4—source data 2
|
| Other |
Soybean polar lipids |
Avanti Polar Lipids |
541602 P |
Used for the reconstitution of GluK1-1aEM in nanodiscs |
| Other |
Dodecyl-β-maltoside (DDM) |
Anatrace |
D310LA |
Used for solubilization of membrane fractions |
| Other |
TALON Cobalt Resin |
Clontech Takara |
635653 |
Used for IMAC purification |
| Other |
Protein A Agarose Beads |
Thermo Scientific |
20334 |
Used for pull-down assays |
| Other |
cOmplete Protease Inhibitor Cocktail |
Roche (Merck) |
11697498001 |
Added to buffers used for protein purification |
