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- EMDB-9553: Single particle reconstruction of CNTNAP2: conformation #3 -

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Basic information

Entry
Database: EMDB / ID: EMD-9553
TitleSingle particle reconstruction of CNTNAP2: conformation #3
Map dataSingle particle reconstruction of CNTNAP2: conformation #3
Sample
  • Complex: CNTNAP2
Function / homology
Function and homology information


limbic system development / clustering of voltage-gated potassium channels / neuron recognition / vocal learning / protein localization to juxtaparanode region of axon / paranodal junction / superior temporal gyrus development / thalamus development / vocalization behavior / paranode region of axon ...limbic system development / clustering of voltage-gated potassium channels / neuron recognition / vocal learning / protein localization to juxtaparanode region of axon / paranodal junction / superior temporal gyrus development / thalamus development / vocalization behavior / paranode region of axon / striatum development / juxtaparanode region of axon / adult behavior / transmission of nerve impulse / social behavior / positive regulation of gap junction assembly / axolemma / prepulse inhibition / voltage-gated potassium channel complex / neuron projection morphogenesis / learning / synaptic membrane / brain development / cerebral cortex development / neuron projection development / protease binding / perikaryon / transmembrane transporter binding / cell population proliferation / early endosome / cell adhesion / axon / neuronal cell body / dendrite / Golgi apparatus / enzyme binding / cell surface / membrane
Similarity search - Function
Neurexin/syndecan/glycophorin C / putative band 4.1 homologues' binding motif / : / Laminin G domain / Laminin G domain profile. / Laminin G domain / Laminin G domain / Coagulation factors 5/8 type C domain (FA58C) signature 2. / Fibrinogen, alpha/beta/gamma chain, C-terminal globular domain / Fibrinogen-like, C-terminal ...Neurexin/syndecan/glycophorin C / putative band 4.1 homologues' binding motif / : / Laminin G domain / Laminin G domain profile. / Laminin G domain / Laminin G domain / Coagulation factors 5/8 type C domain (FA58C) signature 2. / Fibrinogen, alpha/beta/gamma chain, C-terminal globular domain / Fibrinogen-like, C-terminal / Fibrinogen C-terminal domain profile. / Coagulation factors 5/8 type C domain (FA58C) signature 1. / Coagulation factor 5/8 C-terminal domain, discoidin domain / Coagulation factors 5/8 type C domain (FA58C) profile. / F5/8 type C domain / Coagulation factor 5/8 C-terminal domain / Epidermal growth factor-like domain. / EGF-like domain profile. / EGF-like domain / Galactose-binding-like domain superfamily / Concanavalin A-like lectin/glucanase domain superfamily
Similarity search - Domain/homology
Contactin-associated protein-like 2
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / negative staining / Resolution: 15.9 Å
AuthorsLu Z / Reddy MS / Liu JF / Kalichava A / Liu JK / Zhang L / Chen F / Wang Y / Holthauzen LM / Seshadrinathan S ...Lu Z / Reddy MS / Liu JF / Kalichava A / Liu JK / Zhang L / Chen F / Wang Y / Holthauzen LM / Seshadrinathan S / Zhong XY / Ren G / Rudenko G
Funding support United States, 2 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of Mental HealthR01MH077303 United States
Department of Energy (United States)DE-AC02-05CH11231 United States
CitationJournal: J Biol Chem / Year: 2016
Title: Molecular Architecture of Contactin-associated Protein-like 2 (CNTNAP2) and Its Interaction with Contactin 2 (CNTN2).
Authors: Zhuoyang Lu / M V V V Sekhar Reddy / Jianfang Liu / Ana Kalichava / Jiankang Liu / Lei Zhang / Fang Chen / Yun Wang / Luis Marcelo F Holthauzen / Mark A White / Suchithra Seshadrinathan / ...Authors: Zhuoyang Lu / M V V V Sekhar Reddy / Jianfang Liu / Ana Kalichava / Jiankang Liu / Lei Zhang / Fang Chen / Yun Wang / Luis Marcelo F Holthauzen / Mark A White / Suchithra Seshadrinathan / Xiaoying Zhong / Gang Ren / Gabby Rudenko /
Abstract: Contactin-associated protein-like 2 (CNTNAP2) is a large multidomain neuronal adhesion molecule implicated in a number of neurological disorders, including epilepsy, schizophrenia, autism spectrum ...Contactin-associated protein-like 2 (CNTNAP2) is a large multidomain neuronal adhesion molecule implicated in a number of neurological disorders, including epilepsy, schizophrenia, autism spectrum disorder, intellectual disability, and language delay. We reveal here by electron microscopy that the architecture of CNTNAP2 is composed of a large, medium, and small lobe that flex with respect to each other. Using epitope labeling and fragments, we assign the F58C, L1, and L2 domains to the large lobe, the FBG and L3 domains to the middle lobe, and the L4 domain to the small lobe of the CNTNAP2 molecular envelope. Our data reveal that CNTNAP2 has a very different architecture compared with neurexin 1α, a fellow member of the neurexin superfamily and a prototype, suggesting that CNTNAP2 uses a different strategy to integrate into the synaptic protein network. We show that the ectodomains of CNTNAP2 and contactin 2 (CNTN2) bind directly and specifically, with low nanomolar affinity. We show further that mutations in CNTNAP2 implicated in autism spectrum disorder are not segregated but are distributed over the whole ectodomain. The molecular shape and dimensions of CNTNAP2 place constraints on how CNTNAP2 integrates in the cleft of axo-glial and neuronal contact sites and how it functions as an organizing and adhesive molecule.
History
DepositionOct 3, 2016-
Header (metadata) releaseOct 19, 2016-
Map releaseOct 19, 2016-
UpdateNov 6, 2019-
Current statusNov 6, 2019Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 1
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 1
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_9553.png

Downloads & links

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Map

FileDownload / File: emd_9553.map.gz / Format: CCP4 / Size: 8 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationSingle particle reconstruction of CNTNAP2: conformation #3
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.88 Å/pix.
x 128 pix.
= 240.64 Å		1.88 Å/pix.
x 128 pix.
= 240.64 Å		1.88 Å/pix.
x 128 pix.
= 240.64 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.88 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 1.0 / Movie #1: 1
Minimum - Maximum-0.9394105 - 3.1539044
Average (Standard dev.)-0.000000001024257 (±0.20710926)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-34-34-34
Dimensions128128128
Spacing128128128
CellA=B=C: 240.64 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.881.881.88
M x/y/z128128128
origin x/y/z0.0000.0000.000
length x/y/z240.640240.640240.640
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS-34-34-34
NC/NR/NS128128128
D min/max/mean-0.9393.154-0.000

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Supplemental data

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Additional map: Single particle reconstruction of CNTNAP2: conformation #3, additional...

Fileemd_9553_additional.map
AnnotationSingle particle reconstruction of CNTNAP2: conformation #3, additional map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : CNTNAP2

EntireName: CNTNAP2
Components
  • Complex: CNTNAP2

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Supramolecule #1: CNTNAP2

SupramoleculeName: CNTNAP2 / type: complex / ID: 1 / Parent: 0
Details: Human contactin-associated protein-like 2 (CNTNAP2) ectodomain (or fragments) with a C-terminal ASTSHHHHHH tag were produced using baculovirus-mediated overexpression in High Five cells in ...Details: Human contactin-associated protein-like 2 (CNTNAP2) ectodomain (or fragments) with a C-terminal ASTSHHHHHH tag were produced using baculovirus-mediated overexpression in High Five cells in Insect-XPRESS+L-Glutamine medium (Lonza).
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper) / Recombinant cell: High Five / Recombinant plasmid: baculovirus
Molecular weightTheoretical: 134 kDa/nm

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Experimental details

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Structure determination

Methodnegative staining
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration0.005 mg/mL
BufferpH: 8
Component:
ConcentrationFormulaName
25.0 mMC4H11NO3Tris
100.0 mMNaClsodium chloride
3.0 mMCaCl2calcium chloride
StainingType: NEGATIVE / Material: uranium formate / Details: 1% (w/v) uranyl formate
GridModel: EMSCF, 200-Cu / Material: COPPER / Mesh: 200 / Support film - Material: CARBON / Support film - topology: CONTINUOUS / Support film - Film thickness: 5.0 nm / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: OTHER
DetailsThis sample was monodisperse.

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Electron microscopy

MicroscopeZEISS LIBRA120PLUS
Specialist opticsEnergy filter - Name: Zeiss in-column Omega filter / Energy filter - Lower energy threshold: 0 eV / Energy filter - Upper energy threshold: 20 eV
Image recordingFilm or detector model: GATAN ULTRASCAN 4000 (4k x 4k) / Average electron dose: 3.0 e/Å2
Electron beamAcceleration voltage: 120 kV / Electron source: LAB6
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.2 mm / Nominal defocus max: 0.6 µm / Nominal defocus min: 0.1 µm / Nominal magnification: 125000
Sample stageSpecimen holder model: GATAN 626 SINGLE TILT LIQUID NITROGEN CRYO TRANSFER HOLDER

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Image processing

Particle selectionNumber selected: 53774
CTF correctionSoftware - Name: SPIDER (ver. 1.8)
Startup modelType of model: OTHER / Details: IPET reconstructed density map (low pass filtered)
Final reconstructionResolution.type: BY AUTHOR / Resolution: 15.9 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: IPET / Number images used: 53774
Initial angle assignmentType: ANGULAR RECONSTITUTION / Software - Name: EMAN (ver. 1.9)
Final angle assignmentType: ANGULAR RECONSTITUTION / Software - Name: EMAN (ver. 1.9)
Final 3D classificationNumber classes: 200 / Software - Name: EMAN (ver. 1.9)

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