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- EMDB-0864: MicroED structure of lysozyme at 1.73A determained using crystal ... -

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Basic information

Entry
Database: EMDB / ID: EMD-0864
TitleMicroED structure of lysozyme at 1.73A determained using crystal lamellas prepared by focused ion beam milling
Map data
Sample
  • Complex: lysozyme
    • Protein or peptide: Lysozyme C
  • Ligand: ACETATE ION
  • Ligand: water
KeywordsLysozyme / MicroED / Focused Ion Beam / crystal lamella / HYDROLASE
Function / homology
Function and homology information


Lactose synthesis / Antimicrobial peptides / Neutrophil degranulation / beta-N-acetylglucosaminidase activity / cell wall macromolecule catabolic process / lysozyme / lysozyme activity / defense response to Gram-negative bacterium / killing of cells of another organism / defense response to Gram-positive bacterium ...Lactose synthesis / Antimicrobial peptides / Neutrophil degranulation / beta-N-acetylglucosaminidase activity / cell wall macromolecule catabolic process / lysozyme / lysozyme activity / defense response to Gram-negative bacterium / killing of cells of another organism / defense response to Gram-positive bacterium / defense response to bacterium / endoplasmic reticulum / extracellular space / identical protein binding / cytoplasm
Similarity search - Function
Glycoside hydrolase, family 22, lysozyme / Glycoside hydrolase family 22 domain / Glycosyl hydrolases family 22 (GH22) domain signature. / Glycoside hydrolase, family 22 / C-type lysozyme/alpha-lactalbumin family / Glycosyl hydrolases family 22 (GH22) domain profile. / Alpha-lactalbumin / lysozyme C / Lysozyme-like domain superfamily
Similarity search - Domain/homology
Biological speciesGallus gallu (chicken) / Gallus gallus (chicken)
Methodelectron crystallography / cryo EM / Resolution: 1.73 Å
AuthorsZhou H / Luo Z
Funding support China, 4 items
OrganizationGrant numberCountry
National Natural Science Foundation of China31570730 China
National Natural Science Foundation of China31722015 China
Ministry of Science and Technology (China)2016YFA0501102 China
Ministry of Science and Technology (China)2016YFA0501902 China
CitationJournal: J Struct Biol / Year: 2019
Title: Using focus ion beam to prepare crystal lamella for electron diffraction.
Authors: Heng Zhou / Zhipu Luo / Xueming Li /
Abstract: Electron diffraction provides a powerful tool to solve the structures of small protein crystals. However, strong interactions between the electrons and the materials limit the application of the ...Electron diffraction provides a powerful tool to solve the structures of small protein crystals. However, strong interactions between the electrons and the materials limit the application of the electron crystallographic method on large protein crystals with micrometer or larger sizes. Here, we used the focused ion beam (FIB) equipped on the scanning electron microscope (SEM) to mill a large crystal to thin lamella. The influences of the milling on the crystal lamella were observed and investigated, including radiation damage on the crystal surface during the FIB imaging, deformation of the thin crystal lamella, and variation in the diffraction intensities under electron radiation. These observations provide important information to optimize the FIB milling, and hence is important to obtain high-quality crystal samples for routine structure determination of protein crystals using the electron cryo-microscope.
History
DepositionNov 13, 2019-
Header (metadata) releaseNov 27, 2019-
Map releaseNov 27, 2019-
UpdateOct 23, 2024-
Current statusOct 23, 2024Processing site: PDBj / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.1
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 0.1
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6lav
  • Surface level: 0.1
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-6lav
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_0864.png

Downloads & links

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Map

FileDownload / File: emd_0864.map.gz / Format: CCP4 / Size: 2.3 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)X (Row.)Y (Col.)
0.59 Å/pix.
x 86 pix.
= 37.889 Å		0.56 Å/pix.
x 88 pix.
= 78.987 Å		0.56 Å/pix.
x 78 pix.
= 78.987 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

generated in cubic-lattice coordinate

Voxel sizeX: 0.56419 Å / Y: 0.56419 Å / Z: 0.59202 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.1 / Movie #1: 0.1
Minimum - Maximum-0.25392598 - 0.76632047
Average (Standard dev.)0.000088706925 (±0.10233926)
SymmetrySpace group: 96
Details

EMDB XML:

Map geometry
Axis orderYXZ
Origin-420-29
Dimensions887886
Spacing14014064
CellA: 78.986595 Å / B: 78.986595 Å / C: 37.88928 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z0.564192857142860.564192857142860.592015625
M x/y/z14014064
origin x/y/z0.0000.0000.000
length x/y/z78.98778.98737.889
α/β/γ90.00090.00090.000
start NX/NY/NZ-420-29
NX/NY/NZ887886
MAP C/R/S213
start NC/NR/NS0-42-29
NC/NR/NS788886
D min/max/mean-0.2540.7660.000

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Supplemental data

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Sample components

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Entire : lysozyme

EntireName: lysozyme
Components
  • Complex: lysozyme
    • Protein or peptide: Lysozyme C
  • Ligand: ACETATE ION
  • Ligand: water

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Supramolecule #1: lysozyme

SupramoleculeName: lysozyme / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Gallus gallu (chicken)

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Macromolecule #1: Lysozyme C

MacromoleculeName: Lysozyme C / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO / EC number: lysozyme
Source (natural)Organism: Gallus gallus (chicken)
Molecular weightTheoretical: 14.33116 KDa
SequenceString:
KVFGRCELAA AMKRHGLDNY RGYSLGNWVC AAKFESNFNT QATNRNTDGS TDYGILQINS RWWCNDGRTP GSRNLCNIPC SALLSSDIT ASVNCAKKIV SDGNGMNAWV AWRNRCKGTD VQAWIRGCRL

UniProtKB: Lysozyme C

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Macromolecule #2: ACETATE ION

MacromoleculeName: ACETATE ION / type: ligand / ID: 2 / Number of copies: 2 / Formula: ACT
Molecular weightTheoretical: 59.044 Da
Chemical component information

ChemComp-ACT:
ACETATE ION

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Macromolecule #3: water

MacromoleculeName: water / type: ligand / ID: 3 / Number of copies: 37 / Formula: HOH
Molecular weightTheoretical: 18.015 Da
Chemical component information

ChemComp-HOH:
WATER

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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: 4.5
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TECNAI F20
Image recordingFilm or detector model: GATAN ULTRASCAN 4000 (4k x 4k) / Average electron dose: 0.057 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: DIFFRACTION / Camera length: 1200 mm
Experimental equipment
Model: Tecnai F20 / Image courtesy: FEI Company

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Image processing

Final reconstructionResolution.type: BY AUTHOR / Resolution: 1.73 Å / Resolution method: DIFFRACTION PATTERN/LAYERLINES
Crystallography statisticsNumber intensities measured: 121300 / Number structure factors: 11548 / Fourier space coverage: 89 / R sym: 0.281 / R merge: 0.281 / Overall phase error: 0 / Overall phase residual: 1 / Phase error rejection criteria: 60 / High resolution: 1.7 Å / Shell - Shell ID: 1 / Shell - High resolution: 1.7 Å / Shell - Low resolution: 1.76 Å / Shell - Number structure factors: 1002 / Shell - Phase residual: 1 / Shell - Fourier space coverage: 79.1 / Shell - Multiplicity: 9.7

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Atomic model buiding 1

RefinementSpace: RECIPROCAL / Protocol: FLEXIBLE FIT
Output model

PDB-6lav:
MicroED structure of lysozyme at 1.73A determained using crystal lamellas prepared by focused ion beam milling

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