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- EMDB-18680: FZD3 in complex with nanobody 9 -

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Basic information

Entry
Database: EMDB / ID: EMD-18680
TitleFZD3 in complex with nanobody 9
Map dataFZD3 main map
Sample
  • Cell: FZD3 Nanobody Nb9 complex
    • Protein or peptide: Frizzled-3
    • Protein or peptide: Nanobody Nb9
KeywordsFZD3 Nanobody / CELL ADHESION
Function / homology
Function and homology information


dopaminergic neuron axon guidance / serotonergic neuron axon guidance / cell proliferation in midbrain / establishment of planar polarity / negative regulation of mitotic cell cycle, embryonic / Wnt receptor activity / midbrain morphogenesis / motor neuron migration / non-canonical Wnt signaling pathway / sympathetic ganglion development ...dopaminergic neuron axon guidance / serotonergic neuron axon guidance / cell proliferation in midbrain / establishment of planar polarity / negative regulation of mitotic cell cycle, embryonic / Wnt receptor activity / midbrain morphogenesis / motor neuron migration / non-canonical Wnt signaling pathway / sympathetic ganglion development / filopodium tip / Wnt-protein binding / post-anal tail morphogenesis / negative regulation of execution phase of apoptosis / commissural neuron axon guidance / PCP/CE pathway / Class B/2 (Secretin family receptors) / Wnt signaling pathway, planar cell polarity pathway / inner ear morphogenesis / positive regulation of neuroblast proliferation / presynaptic active zone / hair follicle development / lateral plasma membrane / canonical Wnt signaling pathway / response to electrical stimulus / neural tube closure / Asymmetric localization of PCP proteins / PDZ domain binding / G protein-coupled receptor activity / neuron differentiation / Ca2+ pathway / response to xenobiotic stimulus / apical plasma membrane / axon / neuronal cell body / dendrite / cell surface / plasma membrane / cytoplasm
Similarity search - Function
Frizzled 3, cysteine-rich domain / Frizzled/Smoothened, transmembrane domain / Frizzled/Smoothened family membrane region / Frizzled/Smoothened family membrane region / Frizzled/secreted frizzled-related protein / Frizzled / Frizzled domain / Frizzled cysteine-rich domain superfamily / Fz domain / Frizzled (fz) domain profile. ...Frizzled 3, cysteine-rich domain / Frizzled/Smoothened, transmembrane domain / Frizzled/Smoothened family membrane region / Frizzled/Smoothened family membrane region / Frizzled/secreted frizzled-related protein / Frizzled / Frizzled domain / Frizzled cysteine-rich domain superfamily / Fz domain / Frizzled (fz) domain profile. / GPCR, family 2-like / G-protein coupled receptors family 2 profile 2.
Similarity search - Domain/homology
Biological speciesHomo sapiens (human) / Lama glama (llama)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.9 Å
AuthorsZhao Y / Jones EY
Funding support United Kingdom, 1 items
OrganizationGrant numberCountry
Wellcome Trust223133/Z/21/Z United Kingdom
CitationJournal: Nat Commun / Year: 2024
Title: Structural insights into Frizzled3 through nanobody modulators.
Authors: James Hillier / Yuguang Zhao / Loic Carrique / Tomas Malinauskas / Reinis R Ruza / Tao-Hsin Chang / Gangshun Yi / Helen M E Duyvesteyn / Jing Yu / Weixian Lu / Els Pardon / Jan Steyaert / ...Authors: James Hillier / Yuguang Zhao / Loic Carrique / Tomas Malinauskas / Reinis R Ruza / Tao-Hsin Chang / Gangshun Yi / Helen M E Duyvesteyn / Jing Yu / Weixian Lu / Els Pardon / Jan Steyaert / Yanan Zhu / Tao Ni / E Yvonne Jones /
Abstract: The Wnt receptor Frizzled3 (FZD3) is important for brain axonal development and cancer progression. We report structures of FZD3 in complex with extracellular and intracellular binding nanobodies (Nb) ...The Wnt receptor Frizzled3 (FZD3) is important for brain axonal development and cancer progression. We report structures of FZD3 in complex with extracellular and intracellular binding nanobodies (Nb). The crystal structure of Nb8 in complex with the FZD3 cysteine-rich domain (CRD) reveals that the nanobody binds at the base of the lipid-binding groove and can compete with Wnt5a. Nb8 fused with the Dickkopf-1 C-terminal domain behaves as a FZD3-specific Wnt surrogate, activating β-catenin signalling. The cryo-EM structure of FZD3 in complex with Nb9 reveals partially resolved density for the CRD, which exhibits positional flexibility, and a transmembrane conformation that resembles active GPCRs. Nb9 binds to the cytoplasmic region of FZD3 at the putative Dishevelled (DVL) or G protein-binding site, competes with DVL binding, and inhibits GαS coupling. In combination, our FZD3 structures with nanobody modulators map extracellular and intracellular interaction surfaces of functional, and potentially therapeutic, relevance.
History
DepositionOct 18, 2023-
Header (metadata) releaseSep 4, 2024-
Map releaseSep 4, 2024-
UpdateOct 23, 2024-
Current statusOct 23, 2024Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_18680.png

Downloads & links

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Map

FileDownload / File: emd_18680.map.gz / Format: CCP4 / Size: 512 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationFZD3 main map
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.83 Å/pix.
x 512 pix.
= 425.472 Å		0.83 Å/pix.
x 512 pix.
= 425.472 Å		0.83 Å/pix.
x 512 pix.
= 425.472 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.831 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.4
Minimum - Maximum-1.6515971 - 2.4066694
Average (Standard dev.)-0.0006204813 (±0.018870935)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions512512512
Spacing512512512
CellA=B=C: 425.472 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_18680_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: FZD3 DeepEMhancer

Fileemd_18680_additional_1.map
AnnotationFZD3 DeepEMhancer
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: FZD3 3Dflex

Fileemd_18680_additional_2.map
AnnotationFZD3 3Dflex
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: FZD3 half A

Fileemd_18680_half_map_1.map
AnnotationFZD3 half A
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: FZD3 half B

Fileemd_18680_half_map_2.map
AnnotationFZD3 half B
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : FZD3 Nanobody Nb9 complex

EntireName: FZD3 Nanobody Nb9 complex
Components
  • Cell: FZD3 Nanobody Nb9 complex
    • Protein or peptide: Frizzled-3
    • Protein or peptide: Nanobody Nb9

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Supramolecule #1: FZD3 Nanobody Nb9 complex

SupramoleculeName: FZD3 Nanobody Nb9 complex / type: cell / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Homo sapiens (human)

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Macromolecule #1: Frizzled-3

MacromoleculeName: Frizzled-3 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 60.55927 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MAMTWIVFSL WPLTVFMGHI GGHSLFSCEP ITLRMCQDLP YNTTFMPNLL NHYDQQTAAL AMEPFHPMVN LDCSRDFRPF LCALYAPIC MEYGRVTLPC RRLCQRAYSE CSKLMEMFGV PWPEDMECSR FPDCDEPYPR LVDLNLAGEP TEGAPVAVQR D YGFWCPRE ...String:
MAMTWIVFSL WPLTVFMGHI GGHSLFSCEP ITLRMCQDLP YNTTFMPNLL NHYDQQTAAL AMEPFHPMVN LDCSRDFRPF LCALYAPIC MEYGRVTLPC RRLCQRAYSE CSKLMEMFGV PWPEDMECSR FPDCDEPYPR LVDLNLAGEP TEGAPVAVQR D YGFWCPRE LKIDPDLGYS FLHVRDCSPP CPNMYFRREE LSFARYFIGL ISIICLSATL FTFLTFLIDV TRFRYPERPI IF YAVCYMM VSLIFFIGFL LEDRVACNAS IPAQYKASTV TQGSHNKACT MLFMILYFFT MAGSVWWVIL TITWFLAAVP KWG SEAIEK KALLFHASAW GIPGTLTIIL LAMNKIEGDN ISGVCFVGLY DVDALRYFVL APLCLYVVVG VSLLLAGIIS LNRV RIEIP LEKENQDKLV KFMIRIGVFS ILYLVPLLVV IGCYFYEQAY RGIWETTWIQ ERCREYHIPC PYQVTQMSRP DLILF LMKY LMALIVGIPS VFWVGSKKTC FEWASFFHGR RKKGTLEVLF Q

UniProtKB: Frizzled-3

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Macromolecule #2: Nanobody Nb9

MacromoleculeName: Nanobody Nb9 / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Lama glama (llama)
Molecular weightTheoretical: 14.084658 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
QVQLVESGGG LVQSGDSLRL SCTASGRIFN LDVMGWYRQA PGKRRELVAD ITSGGKINYA DSVKGRFTIS RDNTKDTVYL QMNSLKPED TAVYYCNAEV EWLDMDYWGK GTQVTVSSHH HHHHHH

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation statecell

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Sample preparation

BufferpH: 8
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 40.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.0 µm / Nominal defocus min: 1.0 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: NONE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 2.9 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 330717
Initial angle assignmentType: NOT APPLICABLE
Final angle assignmentType: NOT APPLICABLE
FSC plot (resolution estimation)

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