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- EMDB-45428: MicroED structure of the human MP20 protein -

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Basic information

Entry
Database: EMDB / ID: EMD-45428
TitleMicroED structure of the human MP20 protein
Map data2mfo-fc map for MP20
Sample
  • Organelle or cellular component: MP20 crystals grown in lipidic cubic phase
    • Protein or peptide: Lens fiber membrane intrinsic protein
KeywordsLens / Cataract / MicroED / Tetraspanin / CELL ADHESION
Function / homology
Function and homology information


structural constituent of eye lens / cell-cell junction assembly / lens development in camera-type eye / cell junction / vesicle / plasma membrane
Similarity search - Function
Lens fibre membrane intrinsic protein / PMP-22/EMP/MP20 / PMP-22 / EMP / MP20 family signature 2. / : / PMP-22/EMP/MP20/Claudin family / PMP-22/EMP/MP20/Claudin superfamily
Similarity search - Domain/homology
Lens fiber membrane intrinsic protein
Similarity search - Component
Biological speciesSpodoptera frugiperda (fall armyworm) / Homo sapiens (human)
Methodelectron crystallography / cryo EM / Resolution: 3.5 Å
AuthorsNicolas WJ / Shiriaeva A / Martynowycz MW / Grey AC / Ruma Y / Donaldson PJ / Gonen T
Funding support United States, 2 items
OrganizationGrant numberCountry
National Institutes of Health/National Eye Institute (NIH/NEI)P41GM136508 United States
Howard Hughes Medical Institute (HHMI) United States
CitationJournal: bioRxiv / Year: 2024
Title: Structure of the lens MP20 mediated adhesive junction.
Authors: William J Nicolas / Anna Shiriaeva / Michael W Martynowycz / Angus C Grey / Yasmeen Ruma / Paul J Donaldson / Tamir Gonen /
Abstract: Human lens fiber membrane intrinsic protein MP20 is the second most abundant membrane protein of the human eye lens. Despite decades of effort its structure and function remained elusive. Here, we ...Human lens fiber membrane intrinsic protein MP20 is the second most abundant membrane protein of the human eye lens. Despite decades of effort its structure and function remained elusive. Here, we determined the MicroED structure of full-length human MP20 in lipidic-cubic phase to a resolution of 3.5 Å. MP20 forms tetramers each of which contain 4 transmembrane α-helices that are packed against one another forming a helical bundle. Both the N- and C- termini of MP20 are cytoplasmic. We found that each MP20 tetramer formed adhesive interactions with an opposing tetramer in a head-to-head fashion. These interactions were mediated by the extracellular loops of the protein. The dimensions of the MP20 adhesive junctions are consistent with the 11 nm thin lens junctions. Investigation of MP20 localization in human lenses indicated that in young fiber cells MP20 was stored intracellularly in vesicles and upon fiber cell maturation MP20 inserted into the plasma membrane and restricted the extracellular space. Together these results suggest that MP20 forms lens thin junctions in vivo confirming its role as a structural protein in the human eye lens, essential for its optical transparency.
History
DepositionJun 20, 2024-
Header (metadata) releaseMay 21, 2025-
Map releaseMay 21, 2025-
UpdateMay 21, 2025-
Current statusMay 21, 2025Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_45428.png

Downloads & links

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Map

FileDownload / File: emd_45428.map.gz / Format: CCP4 / Size: 897.5 KB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotation2mfo-fc map for MP20
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesX (Sec.)Y (Row.)Z (Col.)
0.78 Å/pix.
x 47 pix.
= 56.18 Å		0.78 Å/pix.
x 52 pix.
= 56.18 Å		0.79 Å/pix.
x 94 pix.
= 142.5 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

generated in cubic-lattice coordinate

Voxel sizeX: 0.78028 Å / Y: 0.78028 Å / Z: 0.79167 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 1.0
Minimum - Maximum-4.349026 - 6.6047854
Average (Standard dev.)-0.043197606 (±0.9574159)
SymmetrySpace group: 90
Details

EMDB XML:

Map geometry
Axis orderZYX
Origin-38-12-11
Dimensions529447
Spacing7272180
CellA: 56.18 Å / B: 56.18 Å / C: 142.5 Å
α=β=γ: 90.0 °

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Supplemental data

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Sample components

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Entire : MP20 crystals grown in lipidic cubic phase

EntireName: MP20 crystals grown in lipidic cubic phase
Components
  • Organelle or cellular component: MP20 crystals grown in lipidic cubic phase
    • Protein or peptide: Lens fiber membrane intrinsic protein

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Supramolecule #1: MP20 crystals grown in lipidic cubic phase

SupramoleculeName: MP20 crystals grown in lipidic cubic phase / type: organelle_or_cellular_component / ID: 1 / Parent: 0 / Macromolecule list: all
Details: MP20 constructs were expressed in SF9 insect (Spodoptera frugiperda) cells using the Bac-to-bac 251 baculovirus expression system (Invitrogen).
Source (natural)Organism: Spodoptera frugiperda (fall armyworm)
Molecular weightTheoretical: 22 KDa

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Macromolecule #1: Lens fiber membrane intrinsic protein

MacromoleculeName: Lens fiber membrane intrinsic protein / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 25.160506 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: MKTIIALSYI FCLVFADYKD DDDAKLQTMH HHHHHHHHHL EVLFQMYSFM GGGLFCAWVG TILLVVAMAT DHWMQYRLSG SFAHQGLWR YCLGNKCYLQ TDSIAYWNAT RAFMILSALC AISGIIMGIM AFAHQPTFSR ISRPFSAGIM FFSSTLFVVL A LAIYTGVT ...String:
MKTIIALSYI FCLVFADYKD DDDAKLQTMH HHHHHHHHHL EVLFQMYSFM GGGLFCAWVG TILLVVAMAT DHWMQYRLSG SFAHQGLWR YCLGNKCYLQ TDSIAYWNAT RAFMILSALC AISGIIMGIM AFAHQPTFSR ISRPFSAGIM FFSSTLFVVL A LAIYTGVT VSFLGRRFGD WRFSWSYILG WVAVLMTFFA GIFYMCAYRV HECRRLSTPR

UniProtKB: Lens fiber membrane intrinsic protein

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Experimental details

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Structure determination

Methodcryo EM
Processingelectron crystallography
Aggregation state3D array

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Sample preparation

BufferpH: 7.5
GridModel: Quantifoil R2/2 / Material: COPPER / Mesh: 200 / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: OTHER / Details: Frozen in LN2 at ambiant humidity (~40%).
Crystal formationLipid protein ratio: 3.2
Lipid mixture: Molten lipid mix (90% w/w monoolein and 10% w/w cholesterol)
Instrument: syringe coupling system (Hamilton)

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsEnergy filter - Name: TFS Selectris / Energy filter - Slit width: 5 eV
Image recordingFilm or detector model: TFS FALCON 4i (4k x 4k) / Digitization - Dimensions - Width: 4096 pixel / Digitization - Dimensions - Height: 4096 pixel / Number real images: 130000 / Number diffraction images: 130000 / Average exposure time: 0.00325 sec. / Average electron dose: 0.00361 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Calibrated magnification: 2941 / Illumination mode: FLOOD BEAM / Imaging mode: DIFFRACTION / Nominal defocus max: 0.0 µm / Nominal defocus min: 0.0 µm / Nominal magnification: 2500 / Camera length: 1202 mm
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN / Tilt angle: -30.0, 30.0
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.5 Å / Resolution method: DIFFRACTION PATTERN/LAYERLINES / Software - Name: PHENIX
CTF correctionType: NONE
Molecular replacementSoftware - Name: PHENIX
Crystallography statisticsNumber intensities measured: 58259 / Number structure factors: 2785 / Fourier space coverage: 86.6 / R merge: 0.3316 / Overall phase error: 28.84 / Overall phase residual: 0 / Phase error rejection criteria: NONE / High resolution: 3.5 Å / Shell - Shell ID: 1 / Shell - High resolution: 3.5 Å / Shell - Low resolution: 52.26 Å / Shell - Number structure factors: 2785 / Shell - Phase residual: 28.84 / Shell - Fourier space coverage: 86.6 / Shell - Multiplicity: 20.7

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Atomic model buiding 1

Initial modelChain - Source name: AlphaFold / Chain - Initial model type: in silico model / Details: Initially truncated to Poly-A chain
RefinementSpace: RECIPROCAL / Protocol: OTHER / Overall B value: 88.55
Output model

PDB-9cbv:
MicroED structure of the human MP20 protein

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