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- EMDB-21188: human delta protocadherin 1 full ectodomains on membranes of lipo... -

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Basic information

Entry
Database: EMDB / ID: EMD-21188
Titlehuman delta protocadherin 1 full ectodomains on membranes of liposomes tomogram 1
Map datahuman delta protocadherin 1 full ectodomains on membranes tomogram 1
Sample
  • Complex: trans dimers of protocadherin 1 extracellular domains formed between membranes of liposomes
    • Complex: human delta protocadherin 1
      • Protein or peptide: human delta protocadherin 1
    • Complex: liposomes
Biological speciesHomo sapiens (human)
Methodelectron tomography / cryo EM
AuthorsBrasch J / Harrisson OJ / Noble AJ / Carragher B / Potter CS / Shapiro LS
Funding support United States, 6 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical SciencesR01GM118584 United States
National Institutes of Health/National Institute of General Medical SciencesR01GM103310 United States
National Institutes of Health/National Institute of Mental HealthR01MH114817 United States
Other governmentNYSTAR United States
Other privatethe Simons Foundation (SF349247) United States
National Institutes of Health/National Institute of General Medical SciencesF32GM128303 United States
CitationJournal: Cell Rep / Year: 2020
Title: Family-wide Structural and Biophysical Analysis of Binding Interactions among Non-clustered δ-Protocadherins.
Authors: Oliver J Harrison / Julia Brasch / Phinikoula S Katsamba / Goran Ahlsen / Alex J Noble / Hanbin Dan / Rosemary V Sampogna / Clinton S Potter / Bridget Carragher / Barry Honig / Lawrence Shapiro /
Abstract: Non-clustered δ1- and δ2-protocadherins, close relatives of clustered protocadherins, function in cell adhesion and motility and play essential roles in neural patterning. To understand the ...Non-clustered δ1- and δ2-protocadherins, close relatives of clustered protocadherins, function in cell adhesion and motility and play essential roles in neural patterning. To understand the molecular interactions underlying these functions, we used solution biophysics to characterize binding of δ1- and δ2-protocadherins, determined crystal structures of ectodomain complexes from each family, and assessed ectodomain assembly in reconstituted intermembrane junctions by cryoelectron tomography (cryo-ET). Homophilic trans (cell-cell) interactions were preferred for all δ-protocadherins, with additional weaker heterophilic interactions observed exclusively within each subfamily. As expected, δ1- and δ2-protocadherin trans dimers formed through antiparallel EC1-EC4 interfaces, like clustered protocadherins. However, no ectodomain-mediated cis (same-cell) interactions were detectable in solution; consistent with this, cryo-ET of reconstituted junctions revealed dense assemblies lacking the characteristic order observed for clustered protocadherins. Our results define non-clustered protocadherin binding properties and their structural basis, providing a foundation for interpreting their functional roles in neural patterning.
History
DepositionJan 6, 2020-
Header (metadata) releaseFeb 5, 2020-
Map releaseMar 11, 2020-
UpdateMar 11, 2020-
Current statusMar 11, 2020Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Solid view (volume rendering)
  • Imaged by UCSF Chimera
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  • Solid view (volume rendering)
  • Imaged by UCSF Chimera
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Movie viewer
Supplemental images

emd_21188.png

Downloads & links

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Map

FileDownload / File: emd_21188.map.gz / Format: CCP4 / Size: 1.7 GB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationhuman delta protocadherin 1 full ectodomains on membranes tomogram 1
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
9.19 Å/pix.
x 314 pix.
= 2885.66 Å		9.19 Å/pix.
x 1226 pix.
= 11266.939 Å		9.19 Å/pix.
x 1152 pix.
= 10586.88 Å

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

generated in cubic-lattice coordinate

Voxel sizeX=Y=Z: 9.19 Å
Density

Histogram

Histogram (log scale)
Minimum - Maximum-11.651117 - 13.447574
Average (Standard dev.)-0.0000000051 (±0.99999964)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin00-382
Dimensions12261152314
Spacing11521226314
CellA: 10586.88 Å / B: 11266.939 Å / C: 2885.66 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z9.199.18999918433939.19
M x/y/z11521226314
origin x/y/z0.0000.0000.000
length x/y/z10586.88011266.9392885.660
α/β/γ90.00090.00090.000
start NX/NY/NZ-200-200-200
NX/NY/NZ401401401
MAP C/R/S123
start NC/NR/NS00-382
NC/NR/NS11521226314
D min/max/mean-11.65113.448-0.000

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Supplemental data

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Sample components

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Entire : trans dimers of protocadherin 1 extracellular domains formed betw...

EntireName: trans dimers of protocadherin 1 extracellular domains formed between membranes of liposomes
Components
  • Complex: trans dimers of protocadherin 1 extracellular domains formed between membranes of liposomes
    • Complex: human delta protocadherin 1
      • Protein or peptide: human delta protocadherin 1
    • Complex: liposomes

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Supramolecule #1: trans dimers of protocadherin 1 extracellular domains formed betw...

SupramoleculeName: trans dimers of protocadherin 1 extracellular domains formed between membranes of liposomes
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Details: purified full ectodomains of protocadherin 1 are tethered to Ni-NTA lipid head groups presented on the liposome surface through binding of C-terminal hexa-histidine tags
Source (natural)Organism: Homo sapiens (human)

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Supramolecule #2: human delta protocadherin 1

SupramoleculeName: human delta protocadherin 1 / type: complex / ID: 2 / Parent: 1 / Macromolecule list: all
Details: full ectodomain of human delta protocadherin 1 with a C-termin hexa histidine tag

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Supramolecule #3: liposomes

SupramoleculeName: liposomes / type: complex / ID: 3 / Parent: 1
Details: liposomes composed of DOPC and DOGS-NTA (8:2 ratio) with 100nm diameter

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Macromolecule #1: human delta protocadherin 1

MacromoleculeName: human delta protocadherin 1 / type: protein_or_peptide / ID: 1
Details: BiP-signal sequence and hexa histidine C-terminal tag.
Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MKLSLVAAML LLLSAARATR VVYKVPEEQP PNTLIGSLAA DYGFPDVGHL YKLEVGAPYL RVDGKTGDIF TTETSIDR E GLRECQNQLP GDPCILEFEV SITDLVQNGS PRLLEGQIEV QDINDNTPNF ASPVITLAI PENTNIGSLF PIPLASDRDA GPNGVASYEL ...String:
MKLSLVAAML LLLSAARATR VVYKVPEEQP PNTLIGSLAA DYGFPDVGHL YKLEVGAPYL RVDGKTGDIF TTETSIDR E GLRECQNQLP GDPCILEFEV SITDLVQNGS PRLLEGQIEV QDINDNTPNF ASPVITLAI PENTNIGSLF PIPLASDRDA GPNGVASYEL QAGPEAQELF GLQVAEDQEE KQPQLIVMGN LDRERWDSY DLTIKVQDGG SPPRASSALL RVTVLDTNDN APKFERPSYE AELSENSPIG H SVIQVKAN DSDQGANAEI EYTFHQAPEV VRRLLRLDRN TGLITVQGPV DREDLSTLRF SV LAKDRGT NPKSARAQVV VTVKDMNDNA PTIEIRGIGL VTHQDGMANI SEDVAEETAV ALV QVSDRD EGENAAVTCV VAGDVPFQLR QASETGSDSK KKYFLQTTTP LDYEKVKDYT IEIV AVDSG NPPLSSTNSL KVQVVDVNDN APVFTQSVTE VAFPENNKPG EVIAEITASD ADSGS NAEL VYSLEPEPAA KGLFTISPET GEIQVKTSLD REQRESYELK VVAADRGSPS LQGTAT VLV NVLDCNDNDP KFMLSGYNFS VMENMPALSP VGMVTVIDGD KGENAQVQLS VEQDNGD FV IQNGTGTILS SLSFDREQQS TYTFQLKAVD GGVPPRSAYV GVTINVLDEN DNAPYITA P SNTSHKLLTP QTRLGETVSQ VAAEDFDSGV NAELIYSIAG GNPYGLFQIG SHSGAITLE KEIERRHHGL HRLVVKVSDR GKPPRYGTAL VHLYVNETLA NRTLLETLLG HSLDTPLDID IAGDPEYER SKQRGNHHHH HH

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Experimental details

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Structure determination

Methodcryo EM
Processingelectron tomography
Aggregation state3D array

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Sample preparation

Concentration0.7 mg/mL
BufferpH: 7.4
Component:
ConcentrationFormulaName
100.0 mMKClpotassium chloride
25.0 mMHEPES4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid
3.0 mMCaCl2Calcium chloride
5.0 v/v %glycerolglycerol
GridSupport film - Material: CARBON / Support film - topology: LACEY / Details: unspecified
VitrificationCryogen name: ETHANE / Chamber humidity: 65 % / Chamber temperature: 298.15 K / Instrument: GATAN CRYOPLUNGE 3 / Details: 2.5 second blot before plunging..
Detailsaggregated liposomes were deposited onto EM grids.
SectioningOther: NO SECTIONING

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsPhase plate: VOLTA PHASE PLATE / Spherical aberration corrector: Cs corrector (CEOS GmbH) / Energy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K2 QUANTUM (4k x 4k) / Detector mode: COUNTING / Number grids imaged: 1 / Number real images: 47 / Average exposure time: 1.52 sec. / Average electron dose: 1.72 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 0.001 mm
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Final reconstructionAlgorithm: SIMULTANEOUS ITERATIVE (SIRT) / Software - Name: TOMO3D
Software - details: part of Appion-protomo 2.4.1 TOMO3D SIRT
Number images used: 38

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