Key resources table.
| Reagent type (species) or resource |
Designation | Source or reference |
Identifiers | Additional information |
|---|---|---|---|---|
| Cell line (Spodoptera frugiperda) | Sf9 | ATCC | CTL-1711 (RRID:CVCL_0549) |
Insect cells used for baculovirus production |
| Cell line (Homo sapiens) | HEK293S GnTI- | Gift from Prof. H Gobind Khorana (PMID:12370423) | Mammalian cells used for baculovirus expression of hFzd5FL and hFzd5ICL3BRIL | |
| Cell line (Homo sapiens) | HEK293T with Fzd1/2/4/5/7/8 KO | Gift from Prof. Michael Boutros (PMID:28733458) | For signaling assay | |
| Cell line (Mus musculus) | Mouse monoclonal M1 hybridoma | Gift from Prof. Brian K Kobilka (PMID:17962520) | To purify anti-FLAG M1 (mouse IgG2a) antibody to prepare FLAG affinity column (15 mg/ml resin). This was used for purification of hFzd5FL and hFzd5ICL3BRIL. | |
| Strain, strain background (Escherichia coli) | BL21(DE3) | New England Biolabs | C2527I | E. coli strain for expression of the nanobody |
| Strain, strain background (Escherichia coli) | BL21 (gold) | Agilent Technologies | 230130 | E. coli strain for expression of the Fabs |
| Transfected construct (Homo sapiens) | pRK5 hFzd5 wild-type |
Yu et al., 2012
(PMID:23095888) |
For signaling assay | |
| Transfected construct (Homo sapiens) | pRK5 hFzd5 mutants | this study | For signaling assay | |
| Recombinant DNA reagent | 7xTCF-Ffluc | Addgene | 24308 | For signaling assay |
| Recombinant DNA reagent | BestBac Linearized Baculovirus DNA 2.0 |
Expression Systems | 91–002 | For baculovirus production |
| Peptide, recombinant protein | Wnt3a | R and D Systems | 5036-WN | For signaling assay |
| Peptide, recombinant protein | Surrogate Wnt | Miao et al. Cell Stem Cell 2020 |
For signaling assay | |
| Peptide, recombinant protein | DRPB_Fz8 |
Dang et al., 2019
(PMID:31086346) |
For signaling assay | |
| Commercial assay or kit | Luciferase assay kit | Promega | E152A | For signaling assay |
| Chemical compound, drug | n-Dodecyl-β-D-Maltoside (DDM) | Anatrace | D310S | Membrane protein purification |
| Chemical compound, drug | Lauryl Maltose Neopentyl Glycol (LMNG) | Anatrace | NG310 | Membrane protein purification |
| Chemical compound, drug | Glyco-Diosgenin (GDN) | Anatrace | GDN101 | Membrane protein purification |
| Chemical compound, drug | Cholesterol Hemisuccinate tris Salt (CHS) | Anatrace | CH210 | Membrane protein purification |
| Chemical compound, drug | Digitonin | EMD Millipore | 300410 | Cryo-EM specimen freezing additive |
| Software, algorithm | DigitalMicrograph | Gatan | Microscope alignment and cryo-EM data collection | |
| Software, algorithm | serialEM |
https://bio3d.colorado.edu/SerialEM/
(PMID:16182563) |
version 3.6 | Cryo-EM data collection |
| Software, algorithm | MotionCor2 |
https://emcore.ucsf.edu/ucsf-motioncor2/
(PMID:28250466) |
Motion correction of cryo-EM movies | |
| Software, algorithm | cryoSPARC |
https://cryosparc.com/
(PMID:28165473) |
version 2.12.14 | Cryo-EM data processing |
| Software, algorithm | Coot |
https://www2.mrc-lmb.cam.ac.uk/personal/pemsley/coot/
(PMID:20383002) |
Structure model building | |
| Software, algorithm | Isolde |
https://isolde.cimr.cam.ac.uk/
(PMID:29872003) |
Structure model building/refinement | |
| Software, algorithm | Phenix suite |
https://www.phenix-online.org/
(PMID:20124702) |
Structure refinement | |
| Software, algorithm | UCSF Chimera |
https://www.cgl.ucsf.edu/chimera/
(PMID:15264254) |
Initial homology model docking | |
| Software, algorithm | UCSF ChimeraX |
https://www.rbvi.ucsf.edu/chimerax/
(PMID:28710774) |
Structural visualization/figure preparation/Isolde execution | |
| Software, algorithm | PyMol | Schrödinger | Structural visualization/figure preparation | |
| Software, algorithm | GraphPad Prism 7 | GraphPad | analysis of signaling data | |
| Other | CNBr-Activated Sepharose 4 Fast Flow | GE Healthcare | 17098101 | For preparation of anti-FLAG sepharose |
| Other | Lipofectamine 2000 | Invitrogen | 11668030 | Transfection reagent for signaling assay |