[English] 日本語
Yorodumi
- EMDB-25185: Ab initio structure of proteinase K from electron-counted MicroED data -

+
Open data


ID or keywords:

Loading...

-
Basic information

Entry
Database: EMDB / ID: EMD-25185
TitleAb initio structure of proteinase K from electron-counted MicroED data
Map data1.5A resolution 2mFo-dFc MicroED map of proteinase K determined ab initio by electron-counted data
Sample
  • Complex: Proteinase K
    • Protein or peptide: Proteinase K
  • Ligand: 5-amino-2,4,6-triiodobenzene-1,3-dicarboxylic acid
  • Ligand: CALCIUM ION
  • Ligand: water
KeywordsHydrolase
Function / homology
Function and homology information


peptidase K / serine-type endopeptidase activity / proteolysis / extracellular space / metal ion binding
Similarity search - Function
Proteinase K-like catalytic domain / Peptidase S8 propeptide/proteinase inhibitor I9 / Peptidase inhibitor I9 / : / Peptidase S8 propeptide/proteinase inhibitor I9 superfamily / Peptidase S8, subtilisin, His-active site / Serine proteases, subtilase family, histidine active site. / Serine proteases, subtilase family, aspartic acid active site. / Peptidase S8, subtilisin, Asp-active site / Serine proteases, subtilase family, serine active site. ...Proteinase K-like catalytic domain / Peptidase S8 propeptide/proteinase inhibitor I9 / Peptidase inhibitor I9 / : / Peptidase S8 propeptide/proteinase inhibitor I9 superfamily / Peptidase S8, subtilisin, His-active site / Serine proteases, subtilase family, histidine active site. / Serine proteases, subtilase family, aspartic acid active site. / Peptidase S8, subtilisin, Asp-active site / Serine proteases, subtilase family, serine active site. / Peptidase S8, subtilisin, Ser-active site / Peptidase S8, subtilisin-related / Serine proteases, subtilase domain profile. / Peptidase S8/S53 domain superfamily / Peptidase S8/S53 domain / Subtilase family
Similarity search - Domain/homology
Biological speciesParengyodontium album (fungus)
Methodelectron crystallography / cryo EM
AuthorsMartynowycz MW / Clabbers MTB
Funding support United States, 2 items
OrganizationGrant numberCountry
Howard Hughes Medical Institute (HHMI) United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)P41GM136508 United States
CitationJournal: Nat Methods / Year: 2022
Title: Ab initio phasing macromolecular structures using electron-counted MicroED data.
Authors: Michael W Martynowycz / Max T B Clabbers / Johan Hattne / Tamir Gonen /
Abstract: Structures of two globular proteins were determined ab initio using microcrystal electron diffraction (MicroED) data that were collected on a direct electron detector in counting mode. Microcrystals ...Structures of two globular proteins were determined ab initio using microcrystal electron diffraction (MicroED) data that were collected on a direct electron detector in counting mode. Microcrystals were identified using a scanning electron microscope (SEM) and thinned with a focused ion beam (FIB) to produce crystalline lamellae of ideal thickness. Continuous-rotation data were collected using an ultra-low exposure rate to enable electron counting in diffraction. For the first sample, triclinic lysozyme extending to a resolution of 0.87 Å, an ideal helical fragment of only three alanine residues provided initial phases. These phases were improved using density modification, allowing the entire atomic structure to be built automatically. A similar approach was successful on a second macromolecular sample, proteinase K, which is much larger and diffracted to a resolution of 1.5 Å. These results demonstrate that macromolecules can be determined to sub-ångström resolution by MicroED and that ab initio phasing can be successfully applied to counting data.
History
DepositionOct 21, 2021-
Header (metadata) releaseJun 8, 2022-
Map releaseJun 8, 2022-
UpdateOct 16, 2024-
Current statusOct 16, 2024Processing site: RCSB / Status: Released

-
Structure visualization

Supplemental images

emd_25185.png

Downloads & links

-
Map

FileDownload / File: emd_25185.map.gz / Format: CCP4 / Size: 10.3 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotation1.5A resolution 2mFo-dFc MicroED map of proteinase K determined ab initio by electron-counted data
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesX (Sec.)Y (Row.)Z (Col.)
0.37 Å/pix.
x 137 pix.
= 67.08 Å		0.37 Å/pix.
x 147 pix.
= 67.08 Å		0.37 Å/pix.
x 134 pix.
= 106.78 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

generated in cubic-lattice coordinate

Voxel sizeX: 0.37267 Å / Y: 0.37267 Å / Z: 0.37076 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 1.5
Minimum - Maximum-2.7613127 - 12.3482485
Average (Standard dev.)-0.0034129387 (±0.9709549)
SymmetrySpace group: 96
Details

EMDB XML:

Map geometry
Axis orderZYX
Origin-30-6953
Dimensions147134137
Spacing180180288
CellA: 67.08 Å / B: 67.08 Å / C: 106.78 Å
α=β=γ: 90.0 °

-
Supplemental data

-
Sample components

-
Entire : Proteinase K

EntireName: Proteinase K
Components
  • Complex: Proteinase K
    • Protein or peptide: Proteinase K
  • Ligand: 5-amino-2,4,6-triiodobenzene-1,3-dicarboxylic acid
  • Ligand: CALCIUM ION
  • Ligand: water

-
Supramolecule #1: Proteinase K

SupramoleculeName: Proteinase K / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Parengyodontium album (fungus)
Molecular weightTheoretical: 28 KDa

-
Macromolecule #1: Proteinase K

MacromoleculeName: Proteinase K / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO / EC number: peptidase K
Source (natural)Organism: Parengyodontium album (fungus)
Molecular weightTheoretical: 28.930783 KDa
SequenceString: AAQTNAPWGL ARISSTSPGT STYYYDESAG QGSCVYVIDT GIEASHPEFE GRAQMVKTYY YSSRDGNGHG THCAGTVGSR TYGVAKKTQ LFGVKVLDDN GSGQYSTIIA GMDFVASDKN NRNCPKGVVA SLSLGGGYSS SVNSAAARLQ SSGVMVAVAA G NNNADARN ...String:
AAQTNAPWGL ARISSTSPGT STYYYDESAG QGSCVYVIDT GIEASHPEFE GRAQMVKTYY YSSRDGNGHG THCAGTVGSR TYGVAKKTQ LFGVKVLDDN GSGQYSTIIA GMDFVASDKN NRNCPKGVVA SLSLGGGYSS SVNSAAARLQ SSGVMVAVAA G NNNADARN YSPASEPSVC TVGASDRYDR RSSFSNYGSV LDIFGPGTSI LSTWIGGSTR SISGTSMATP HVAGLAAYLM TL GKTTAAS ACRYIADTAN KGDLSNIPFG TVNLLAYNNY QA

UniProtKB: Proteinase K

-
Macromolecule #2: 5-amino-2,4,6-triiodobenzene-1,3-dicarboxylic acid

MacromoleculeName: 5-amino-2,4,6-triiodobenzene-1,3-dicarboxylic acid / type: ligand / ID: 2 / Number of copies: 2 / Formula: I3C
Molecular weightTheoretical: 558.835 Da
Chemical component information

ChemComp-I3C:
5-amino-2,4,6-triiodobenzene-1,3-dicarboxylic acid

-
Macromolecule #3: CALCIUM ION

MacromoleculeName: CALCIUM ION / type: ligand / ID: 3 / Number of copies: 5 / Formula: CA
Molecular weightTheoretical: 40.078 Da

-
Macromolecule #4: water

MacromoleculeName: water / type: ligand / ID: 4 / Number of copies: 234 / Formula: HOH
Molecular weightTheoretical: 18.015 Da
Chemical component information

ChemComp-HOH:
WATER

-
Experimental details

-
Structure determination

Methodcryo EM
Processingelectron crystallography
Aggregation state3D array

-
Sample preparation

Concentration5 mg/mL
BufferpH: 7.5
GridModel: Quantifoil R2/2 / Material: COPPER / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY / Support film - Film thickness: 10 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 30 sec. / Pretreatment - Atmosphere: AIR / Details: NEGATIVE
VitrificationCryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 277 K / Instrument: LEICA PLUNGER
DetailsMilled microcrystals

-
Electron microscopy

MicroscopeFEI TITAN KRIOS
TemperatureMin: 77.0 K / Max: 90.0 K
Image recordingFilm or detector model: FEI FALCON IV (4k x 4k) / Digitization - Dimensions - Width: 2048 pixel / Digitization - Dimensions - Height: 2048 pixel / Number grids imaged: 1 / Number real images: 1 / Number diffraction images: 840 / Average exposure time: 0.5 sec. / Average electron dose: 0.001 e/Å2
Details: 0.15 degrees per second, 0.5 second readout, 30 to -30 degrees
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: DIFFRACTION / Cs: 2.7 mm / Camera length: 1738 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

-
Image processing

DetailsBinned by 2.
Final reconstructionResolution method: DIFFRACTION PATTERN/LAYERLINES / Software - Name: REFMAC
Merging software listSoftware - Name: AIMLESS
Crystallography statisticsNumber intensities measured: 416133 / Number structure factors: 39303 / Fourier space coverage: 98.87 / R sym: 0.087 / R merge: 0.277 / Overall phase error: 20 / Overall phase residual: 0 / Phase error rejection criteria: None / High resolution: 1.5 Å
Details: Phases were determined by placing 4 ideal helix fragments. These were extended by chain tracing and density modifications.
Shell - Shell ID: 1 / Shell - High resolution: 1.53 Å / Shell - Low resolution: 1.5 Å / Shell - Number structure factors: 1758 / Shell - Phase residual: 30 / Shell - Fourier space coverage: 89.3 / Shell - Multiplicity: 8.9

-
Atomic model buiding 1

RefinementSpace: RECIPROCAL / Protocol: AB INITIO MODEL / Overall B value: 14.137 / Target criteria: Maximum likelihood
Output model

PDB-7skx:
Ab initio structure of proteinase K from electron-counted MicroED data

+
About Yorodumi

-
News

-
Feb 9, 2022. New format data for meta-information of EMDB entries

New format data for meta-information of EMDB entries

  • Version 3 of the EMDB header file is now the official format.
  • The previous official version 1.9 will be removed from the archive.

Related info.:EMDB header

External links:wwPDB to switch to version 3 of the EMDB data model

-
Aug 12, 2020. Covid-19 info

Covid-19 info

URL: https://pdbj.org/emnavi/covid19.php

New page: Covid-19 featured information page in EM Navigator.

Related info.:Covid-19 info / Mar 5, 2020. Novel coronavirus structure data

+
Mar 5, 2020. Novel coronavirus structure data

Novel coronavirus structure data

Related info.:Yorodumi Speices / Aug 12, 2020. Covid-19 info

External links:COVID-19 featured content - PDBj / Molecule of the Month (242)：Coronavirus Proteases

+
Jan 31, 2019. EMDB accession codes are about to change! (news from PDBe EMDB page)

EMDB accession codes are about to change! (news from PDBe EMDB page)

  • The allocation of 4 digits for EMDB accession codes will soon come to an end. Whilst these codes will remain in use, new EMDB accession codes will include an additional digit and will expand incrementally as the available range of codes is exhausted. The current 4-digit format prefixed with “EMD-” (i.e. EMD-XXXX) will advance to a 5-digit format (i.e. EMD-XXXXX), and so on. It is currently estimated that the 4-digit codes will be depleted around Spring 2019, at which point the 5-digit format will come into force.
  • The EM Navigator/Yorodumi systems omit the EMD- prefix.

Related info.:Q: What is EMD? / ID/Accession-code notation in Yorodumi/EM Navigator

External links:EMDB Accession Codes are Changing Soon! / Contact to PDBj

+
Jul 12, 2017. Major update of PDB

Major update of PDB

  • wwPDB released updated PDB data conforming to the new PDBx/mmCIF dictionary.
  • This is a major update changing the version number from 4 to 5, and with Remediation, in which all the entries are updated.
  • In this update, many items about electron microscopy experimental information are reorganized (e.g. em_software).
  • Now, EM Navigator and Yorodumi are based on the updated data.

External links:wwPDB Remediation / Enriched Model Files Conforming to OneDep Data Standards Now Available in the PDB FTP Archive

-
Yorodumi

Thousand views of thousand structures

  • Yorodumi is a browser for structure data from EMDB, PDB, SASBDB, etc.
  • This page is also the successor to EM Navigator detail page, and also detail information page/front-end page for Omokage search.
  • The word "yorodu" (or yorozu) is an old Japanese word meaning "ten thousand". "mi" (miru) is to see.

Related info.:EMDB / PDB / SASBDB / Comparison of 3 databanks / Yorodumi Search / Aug 31, 2016. New EM Navigator & Yorodumi / Yorodumi Papers / Jmol/JSmol / Function and homology information / Changes in new EM Navigator and Yorodumi

Read more