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- EMDB-26748: Cryogenic electron microscopy 3D map of alpha-catenin ortholog, HMP1 -

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Basic information

Entry
Database: EMDB / ID: EMD-26748
TitleCryogenic electron microscopy 3D map of alpha-catenin ortholog, HMP1
Map dataCryo-EM 3D map of HMP1
Sample
  • Complex: alpha-catenin ortholog, HMP1
    • Protein or peptide: alpha-catenin ortholog, HMP1
Biological speciesCaenorhabditis elegans (invertebrata)
Methodsingle particle reconstruction / cryo EM / Resolution: 7.8 Å
AuthorsSmith EW / Izard T / Rangarajan ES
Funding support United States, 1 items
OrganizationGrant numberCountry
National Science Foundation (NSF, United States) United States
CitationJournal: J Biol Chem / Year: 2023
Title: The nematode α-catenin ortholog, HMP1, has an extended α-helix when bound to actin filaments.
Authors: Erumbi S Rangarajan / Emmanuel W Smith / Tina Izard /
Abstract: The regulation of cell-cell junctions during epidermal morphogenesis ensures tissue integrity, a process regulated by α-catenin. This cytoskeletal protein connects the cadherin complex to ...The regulation of cell-cell junctions during epidermal morphogenesis ensures tissue integrity, a process regulated by α-catenin. This cytoskeletal protein connects the cadherin complex to filamentous actin at cell-cell junctions. The cadherin-catenin complex plays key roles in cell physiology, organism development, and disease. While mutagenesis of Caenorhabditis elegans cadherin and catenin shows that these proteins are key for embryonic morphogenesis, we know surprisingly little about their structure and attachment to the cytoskeleton. In contrast to mammalian α-catenin that functions as a dimer or monomer, the α-catenin ortholog from C. elegans, HMP1 for humpback, is a monomer. Our cryogenic electron microscopy (cryoEM) structure of HMP1/α-catenin reveals that the amino- and carboxy-terminal domains of HMP1/α-catenin are disordered and not in contact with the remaining HMP1/α-catenin middle domain. Since the carboxy-terminal HMP1/α-catenin domain is the F-actin-binding domain (FABD), this interdomain constellation suggests that HMP1/α-catenin is constitutively active, which we confirm biochemically. Our perhaps most surprising finding, given the high sequence similarity between the mammalian and nematode proteins, is our cryoEM structure of HMP1/α-catenin bound to F-actin. Unlike the structure of mammalian α-catenin bound to F-actin, binding to F-actin seems to allosterically convert a loop region of the HMP1/α-catenin FABD to extend an HMP1/α-catenin FABD α-helix. We use cryoEM and bundling assays to show for the first time how the FABD of HMP1/α-catenin bundles actin in the absence of force. Collectively, our data advance our understanding of α-catenin regulation of cell-cell contacts and additionally aid our understanding of the evolution of multicellularity in metazoans.
History
DepositionApr 26, 2022-
Header (metadata) releaseApr 26, 2023-
Map releaseApr 26, 2023-
UpdateApr 26, 2023-
Current statusApr 26, 2023Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_26748.png

Downloads & links

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Map

FileDownload / File: emd_26748.map.gz / Format: CCP4 / Size: 12.9 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationCryo-EM 3D map of HMP1
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.48 Å/pix.
x 150 pix.
= 222. Å		1.48 Å/pix.
x 150 pix.
= 222. Å		1.48 Å/pix.
x 150 pix.
= 222. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.48 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.135
Minimum - Maximum-0.29033676 - 0.39546028
Average (Standard dev.)4.383967e-05 (±0.015847584)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions150150150
Spacing150150150
CellA=B=C: 222.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: half map A

Fileemd_26748_half_map_1.map
Annotationhalf map A
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: half map B

Fileemd_26748_half_map_2.map
Annotationhalf map 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 : alpha-catenin ortholog, HMP1

EntireName: alpha-catenin ortholog, HMP1
Components
  • Complex: alpha-catenin ortholog, HMP1
    • Protein or peptide: alpha-catenin ortholog, HMP1

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Supramolecule #1: alpha-catenin ortholog, HMP1

SupramoleculeName: alpha-catenin ortholog, HMP1 / type: complex / ID: 1 / Chimera: Yes / Parent: 0 / Macromolecule list: all / Details: Full-length alpha-catenin ortholog, HMP1
Source (natural)Organism: Caenorhabditis elegans (invertebrata)
Molecular weightTheoretical: 103.99428 KDa

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Macromolecule #1: alpha-catenin ortholog, HMP1

MacromoleculeName: alpha-catenin ortholog, HMP1 / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
Source (natural)Organism: Caenorhabditis elegans (invertebrata)
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MPANGNSHAY FNIDEVRSKN VLKQITQLIN EVTNITETFP LKPGQTTEGL VATLDAAVAN FLQTGSFAI SKCPIANSDP RAIDLLHEAL GAVQDTGQVM IQTGRDFVRD STSTNKRAIA T NSGRNLLT AVAKFLILAD SIDVKVIVDK VDEVRETAHQ MIEADTKIKV ...String:
MPANGNSHAY FNIDEVRSKN VLKQITQLIN EVTNITETFP LKPGQTTEGL VATLDAAVAN FLQTGSFAI SKCPIANSDP RAIDLLHEAL GAVQDTGQVM IQTGRDFVRD STSTNKRAIA T NSGRNLLT AVAKFLILAD SIDVKVIVDK VDEVRETAHQ MIEADTKIKV DDLYNLLISQ IE ELDITVR RRAIDLVKPN QRDDLLAARS ALRQTAPLLY TSTRTFVRHP EHEEARRNRD YTA DEMHSA LNALESVLNG QQPKVTFSEY GRIGDLINEI DTFQNRIEID PAHYRRGTDR PDLE GHCER IVSGSASIAD AESTRENRKQ KIVAECNNLR QALQELLTEY EKSTGRRDDN DDIPL GIAE VHKRTKDLRR HLRRAIVDHI SDAFLDTRTP LILLIEAAKE GHEENTRYRS KMFQEH ANE IVSVARLSCQ LSSDVESVSV IQHTAAQLEK LAPQVAQAAI LLCHQPTSKT AQENMET YK NAWFDKVRLL TTALDNITTL DDFLAVSEAH IVEDCERGIK GITANASTPD ENAANCET V DCAAGSIRGR ALRVCDVVDA EMDFLQNSEY TETVKQAVRI LKTQRVDQFA ERASALANR QEAHGLTWDP KTKEEEMNEF INACTLVHDA VKDIRHALLM NRSMNDVDSD VEYEADGVGA ANADANRTI SEQENQQNLM RRLPEEEKKK IQAQIDIFKV TQTRFEREVA KWDETGNDII S LANNMCKI MMSMTEFTRG CGPLKTTMDV IRAAQEISLN GSKLNALARQ IGEESADSQT KK DLLAYLS QITLYCQQLN ICSKVKADVT QVGNELVVSA LDSAMSLIQT ARNLLTAVVQ TVK AAYIAS TKFRRPNANS VRVEWRMAPP KKQPLIRPQK NNAIIRRASE RRPLQPAKVL AEFT RNEIE TGRDSDDEEL DRRHQQRING RL

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration0.3 mg/mL
BufferpH: 7.5
Component:
ConcentrationFormulaName
150.0 mMNaClsodium Chloride
20.0 mMTrisTris
1.0 mMDTTDithiothreitol
GridModel: UltrAuFoil R1.2/1.3 / Material: GOLD / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 240 sec.
VitrificationCryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 277 K / Instrument: LEICA EM GP

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Electron microscopy

MicroscopeJEOL CRYO ARM 300
Specialist opticsEnergy filter - Name: In-column Omega Filter / Energy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Number grids imaged: 1 / Number real images: 1294 / Average exposure time: 0.1 sec. / Average electron dose: 60.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 3.0 µm / Nominal defocus min: 0.5 µm / Nominal magnification: 60000
Sample stageSpecimen holder model: JEOL CRYOSPECPORTER / Cooling holder cryogen: NITROGEN

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Image processing

Particle selectionNumber selected: 3543206
Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

5h5m

Final reconstructionNumber classes used: 5 / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 7.8 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC / Number images used: 149802
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

5h5m

RefinementProtocol: FLEXIBLE FIT

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