REAGENT or RESOURCE	SOURCE	IDENTIFIER
Experimental models: Organisms/strains
FreeStyle HEK293-F Cells	Thermo Fisher	Cat# R79007
HEK293T	ATCC	RRID: CRL-3216
Chemicals, peptides, and recombinant proteins
Turbofect transfection reagent	Thermo Fisher	Cat# R0533
polyethyleneimine	Polysciences	Cat# 24765
protease inhibitor cocktail	Roche	Cat# 04693124001
DNase	Sigma	Cat# DN25
RNase	Sigma	Cat# R4875
PMSF	Sigma	Cat# P7626
LMNG	Anatrace	Cat# NG310
CHS	Anatrace	Cat# CH210
Amphipol A8-35	Anatrace	Cat# A835
Adenosine 3′,5′-cyclophosphate	Sigma	Cat# A9501
Critical commercial assays
QuikChange Lightning Site-Directed Mutagenesis	Thermo Fisher	Cat# 210518
Recombinant DNA plasmids
pEGA: HCN4ΔC	This paper	N/A
pEGA: HCN4ΔC (H407A, H553A)	This paper	N/A
pCI:TRIP8b(1a-4)	Porro et al., 2020	N/A
pCI:rbHCN4	Porro et al., 2019	N/A
pCI:rbHCN4 (H407A)	This paper	N/A
pCI:rbHCN4 (H553A)	This paper	N/A
pCI:rbHCN4 (E557A)	This paper	N/A
pCI:rbHCN4 (H407A, H553A)	This paper	N/A
pCI:rbHCN4 (H407A, H553A, E557A)	This paper	N/A
pCI:hHCN1	Porro et al., 2019	N/A
pCI:hHCN1 (E436A)	This paper	N/A
pCI:mHCN2	Porro et al., 2019	N/A
Deposited data
CryoEM map of HCN4holo	This paper	EMDB: EMD-12513
CryoEM map of HCN4apo/LC	This paper	EMDB: EMD-12512
CryoEM map of HCN4apo/AM	This paper	EMDB: EMD-12466
Atomic model ofHCN4holo	This paper	PDB: 7NP4
Atomic model ofHCN4apo/LC	This paper	PDB: 7NP3
Atomic model ofHCN4apo/AM	This paper	PDB: 7NMN
Software and algorithms
RELION-3.0	Zivanov et al., 2018	https://www2.mrc-lmb.cam.ac.uk/relion/index.php/Main_Page
CryoSPARC v.2	Punjani et al., 2017	https://cryosparc.com
PHENIX	Adams et al., 2010	https://phenix-online.org
Coot	Emsley et al., 2010	https://www2.mrc-lmb.cam.ac.uk/personal/pemsley/coot/
UCSF CHIMERA	Pettersen et al., 2004	http://www.cgl.ucsf.edu/chimera
MOLPROBITY	Chen et al., 2010	http://molprobity.biochem.duke.edu/
HOLE	Smart et al., 1996	http://www.holeprogram.org/
AutoDock 4.2.6 in conjunction with AutoDockTools 1.5.6	Morris et al., 2009	http://autodock.scripps.edu/
GROMACS 2019	Abraham et al., 2015; Van Der Spoel et al., 2005	https://manual.gromacs.org/documentation/2019/index.html#
LINCS	Hess et al., 1997	https://dblp.uni-trier.de/rec/journals/jcc/HessBBF97.html
Biotite	Kunzmann and Hamacher, 2018	https://github.com/biotite-dev/biotite
Other
Grids	Quantifoil	Three-hundred mesh holey gold R0.6/1 and R1.2/1.3