Structural basis for ion selectivity in potassium-selective channelrhodopsins

. Author manuscript; available in PMC: 2023 Oct 12.

Published in final edited form as: Cell. 2023 Aug 30;186(20):4325–4344.e26. doi: 10.1016/j.cell.2023.08.009

REAGENT or RESOURCE	SOURCE	IDENTIFIER
Bacterial and virus strains
One Shot Stbl3 Chemically Competent E. coli	Thermo Fisher Scientific	Cat# C737303
E. coli BL21 (DE3)	Thermo Fisher Scientific	Cat# EC0114
E. coli C41 (DE3)	Sigma-Aldrich	Cat# CMC0017
E. coli Mach1	Thermo Fisher Scientific	Cat# C862003
E. coli DH10Bac	Thermo Fisher Scientific	Cat# 10361012
Adeno-Associated Virus Coat Protein 2/8	Constructed in the Deisseroth lab; packaged at the Stanford Gene Vector and Virus Core (GVVC)	AAV8
AAV8-CaMKIIa-HcKCR1 (WT)-EYFP	Constructed in the Deisseroth lab; packaged at the GVVC	N/A
AAV8-CaMKIIa-HcKCR1 (H225F)-EYFP	Constructed in the Deisseroth lab; packaged at the GVVC	N/A
AAV8-CaMKIIa-HcKCR2 (WT)-EYFP	Constructed in the Deisseroth lab; packaged at the GVVC	N/A
AAV8-CaMKIIa-HcKCR2 (H225F)-EYFP	Constructed in the Deisseroth lab; packaged at the GVVC	N/A
AAV8-CaMKIIα-XCaMP-R	Constructed in the Deisseroth lab; packaged at the GVVC	N/A
AAV8-CaMKIIα-sRGECO	Constructed in the Deisseroth lab; packaged at the GVVC	N/A
AAV8-CaMKIIα-NpHR3.3-mCherry	Constructed in the Deisseroth lab; packaged at the GVVC	N/A
AAV8-CaMKIIα-GCaMP6m	Constructed in the Deisseroth lab; packaged at the GVVC	N/A
Chemicals, peptides, and recombinant proteins
APV	Tocris	Cat# 0106
CNQX	Tocris	Cat# 0190
Tetrodotoxin	Tocris	Cat# 1078
Fluorodeoxyuridine	Sigma	Cat# F0503
Lipofectamine 2000	Thermo Fisher Scientific	Cat# 11668027
FuGENE	Promega	Cat# E2311
Benzamidine	Nacalai tesque	Cat# M9G4533
Leupeptin	Peptide Institute	Cat# 4041
n-Dodecyl-β-D-maltoside (DDM)	EMD Millipore	Cat# 324355
n-Dodecyl-β-D-maltoside (DDM)	EMD Millipore	Cat# D97002-C
Ni-NTA Superflow	QIAGEN	Cat# 30430
Superdex 200 Increase 10/300 GL	Cytiva	Cat# 28990944
TSKgel3000SWXL column	TOSOH bioscience	Cat# 0008541
All-trans-retinal (ATR)	Sigma-Aldrich	Cat# R2500
StockOptions pH buffer kit	Hampton Research	Cat# HR2-241
ESF 921 Insect Cell Culture Medium, Protein Free	Expression systems	Cat# 96-001-01
Monolith His-Tag Labeling Kit RED-tris-NTA 2^ND Generation	NanoTemper Technologies	Cat# MO-L018
Deposited data
HcKCR1 – atomic model	From this paper	PDB: 8H86
HcKCR2 – atomic model	From this paper	PDB: 8H87
HcKCR1 H225F mutant– atomic model	From this paper	PDB: 8IU0
HcKCR1 – Cryo-EM map	From this paper	EMDB-34530
HcKCR2 – Cryo-EM map	From this paper	EMDB-34531
HcKCR1 H225F mutant – Cryo-EM map	From this paper	EMDB-35713
HcKCR1 – Cryo-EM movie frames	From this paper	EMPIAR-11558
HcKCR2 – Cryo-EM movie frames	From this paper	EMPIAR-11558
HcKCR1 H225F mutant – Cryo-EM movie frames	From this paper	EMPIAR-11558
Experimental models: Cell lines
Spodoptera frugiperda Sf9 cells	Expression systems	Cat# 94-001F
HEK293T cells	ATCC	Cat# CRL-3216
Primary rat hippocampal cultured neuron	This paper	N/A
Experimental models: Organisms/strains
C57BL/6J	Jackson	000664
Sprague-Dawley rat pups	Charles River	N/A
Recombinant DNA
pFastBac-HcKCR1 (WT)-EGFP-His10	This paper	N/A
pFastBac-HcKCR1 (WT) -His10	This paper	N/A
pFastBac-HcKCR1 (D105N)-EGFP-His10	This paper	N/A
pFastBac-HcKCR1 (D229N)-EGFP-His10	This paper	N/A
pFastBac-HcKCR1 (H225F)-EGFP-His10	This paper	N/A
pFastBac-HcKCR1 (T136A/G140A)-EGFP-His10	This paper	N/A
pFastBac-HcKCR2 (WT)-EGFP-His8	This paper	N/A
pFastBac-HcKCR2 (D105N)-EGFP-His8	This paper	N/A
pFastBac-HcKCR2 (D229N)-EGFP-His8	This paper	N/A
pFastBac-HcKCR2 (A136T/A140G)-EGFP-His8	This paper	N/A
pET-43a(+)-MSP1E3D1	This paper	N/A
pcDNA 3.1-HcKCR1 (WT)-eYFP	This paper	N/A
pcDNA 3.1-HcKCR2 (WT)-eYFP	This paper	N/A
pET-43a(+)-KR2	This paper	N/A
pAAV-HcKCR1 (WT)-EYFP	This paper	N/A
pAAV-HcKCR1 (H225F)-EYFP	This paper	N/A
Software and algorithms
Serial EM software	Mastronarde⁸⁴	https://bio3d.colorado.edu/SerialEM/
MotionCor2	Zheng et al.⁸⁵	https://emcore.ucsf.edu/ucsf-software
RELION 4.0	Zivanov et al.⁸⁶	https://github.com/3dem/relion
UCSF Chimera	Pettersen et al.⁸⁷	https://www.cgl.ucsf.edu/chimera/
Chimera X	Goddard et al.⁸⁸	https://www.rbvi.ucsf.edu/chimerax/
Coot	Emsley and Cowtan⁸⁹	https://www2.mrc-lmb.cam.ac.uk/personal/pemsley/coot/
Cuemol2	N/A	http://www.cuemol.org/ja/index.php?cuemol2
Servalcat	Yamashita et al.⁹⁰	https://github.com/keitaroyam/servalcat
GraphPad Prism	GraphPad	Graphpad.com
MATLAB	Mathworks	Mathworks.com
pClamp 10.6	Molecular Devices	https://www.moleculardevices.com
Other
Optic fiber implants	Doric lenses	Ordering Code: MFC_400/430-0.66_2.5mm_MF2.5_FLT
MultiClamp700B Amplifier	Molecular Devices	https://www.moleculardevices.com
DigiData 1440A	Molecular Devices	https://www.moleculardevices.com
DM-LFSA	Leica	N/A
Digital Optical Power and Energy Meter	Thorlabs	PM100D
SPECTRA-X Light Engine	Lumencor	https://lumencor.com
R 1.2/1.3 grid: Au 300 mesh	QUANTIFOIL	Cat# N1-C14nAu30-01
AMICON MWCO 10,000	Merck/Millipore	Cat# UFC801024
AMICON MWCO 50,000	Merck/Millipore	Cat# UFC805024
AKTA pure 25 L1	Cytiva	Cat# 29018225
Fluorescence detection HPLC system	Shimadzu	Model Prominence
V-750 UV-Visible Spectrophotometer	JASCO	https://jascoinc.com/products/spectroscopy/
Vitrobot Mark IV	FEI/Thermo Fisher Scientific	https://www.thermofisher.com/jp/ja/home/electron-microscopy/products/sample-preparation-equipment-em/vitrobot-system.html
300 kV Titan Krios microscope	FEI/Thermo Fisher Scientific	Out of production; https://www.thermofisher.com/jp/ja/home/electron-microscopy/products/transmission-electron-microscopes.html
K3 Summit camera with post-column energy filter	Gatan/Quantum	https://www.gatan.com/K3
Innova S44i R	Eppendorf	Cat# S44I010226
NanoTemper Monolith	NanoTemper Technologies	https://nanotempertech.com/monolith/