[English] 日本語
Yorodumi
- EMDB-17973: Archaeoglobus fulgidus AfAgo complex with AfAgo-N protein (fAfAgo... -

+
Open data


ID or keywords:

Loading...

-
Basic information

Entry
Database: EMDB / ID: EMD-17973
TitleArchaeoglobus fulgidus AfAgo complex with AfAgo-N protein (fAfAgo) bound with 30 nt RNA guide and 51 nt DNA target
Map dataphenix.auto_sharped of cryosparc local refinement map
Sample
  • Complex: AfAgo and AfAgo-N complex with 30 nt RNA guide and 51 nt DNA target
    • Complex: Piwi protein
      • Protein or peptide: Piwi protein
    • Complex: Archaeoglobus fulgidus AfAgo-N protein
      • Protein or peptide: Archaeoglobus fulgidus AfAgo-N protein
    • Complex: RNA (30-MER)
      • RNA: RNA (30-MER)
    • Complex: DNA (51-MER)
      • DNA: DNA (51-MER)
  • Ligand: MAGNESIUM ION
  • Ligand: water
KeywordsARGONAUTE / PIWI DOMAIN / PROTEIN-DNA COMPLEX / DNA BINDING PROTEIN
Function / homologyPiwi domain / Piwi domain profile. / Piwi domain / Piwi / Ribonuclease H superfamily / nucleic acid binding / Ribonuclease H-like superfamily / Uncharacterized protein / Piwi protein
Function and homology information
Biological speciesArchaeoglobus fulgidus (archaea) / Archaeoglobus fulgidus DSM 8774 (archaea) / Escherichia coli (E. coli)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.81 Å
AuthorsManakova EN / Zaremba M / Pocevicuite R / Golovinas E / Sasnauskas G / Zagorskaite E / Silanskas A
Funding supportLithuania, 2 items
OrganizationGrant numberCountry
Research Council of LithuaniaS-MIP-23-131Lithuania
Research Council of LithuaniaS-MIP-20-37Lithuania
CitationJournal: Nucleic Acids Res / Year: 2024
Title: The missing part: the Archaeoglobus fulgidus Argonaute forms a functional heterodimer with an N-L1-L2 domain protein.
Authors: Elena Manakova / Edvardas Golovinas / Reda Pocevičiūtė / Giedrius Sasnauskas / Arunas Silanskas / Danielis Rutkauskas / Marija Jankunec / Evelina Zagorskaitė / Edvinas Jurgelaitis / ...Authors: Elena Manakova / Edvardas Golovinas / Reda Pocevičiūtė / Giedrius Sasnauskas / Arunas Silanskas / Danielis Rutkauskas / Marija Jankunec / Evelina Zagorskaitė / Edvinas Jurgelaitis / Algirdas Grybauskas / Česlovas Venclovas / Mindaugas Zaremba
Abstract: Argonaute (Ago) proteins are present in all three domains of life (bacteria, archaea and eukaryotes). They use small (15-30 nucleotides) oligonucleotide guides to bind complementary nucleic acid ...Argonaute (Ago) proteins are present in all three domains of life (bacteria, archaea and eukaryotes). They use small (15-30 nucleotides) oligonucleotide guides to bind complementary nucleic acid targets and are responsible for gene expression regulation, mobile genome element silencing, and defence against viruses or plasmids. According to their domain organization, Agos are divided into long and short Agos. Long Agos found in prokaryotes (long-A and long-B pAgos) and eukaryotes (eAgos) comprise four major functional domains (N, PAZ, MID and PIWI) and two structural linker domains L1 and L2. The majority (∼60%) of pAgos are short pAgos, containing only the MID and inactive PIWI domains. Here we focus on the prokaryotic Argonaute AfAgo from Archaeoglobus fulgidus DSM4304. Although phylogenetically classified as a long-B pAgo, AfAgo contains only MID and catalytically inactive PIWI domains, akin to short pAgos. We show that AfAgo forms a heterodimeric complex with a protein encoded upstream in the same operon, which is a structural equivalent of the N-L1-L2 domains of long pAgos. This complex, structurally equivalent to a long PAZ-less pAgo, outperforms standalone AfAgo in guide RNA-mediated target DNA binding. Our findings provide a missing piece to one of the first and the most studied pAgos.
History
DepositionJul 18, 2023-
Header (metadata) releaseNov 29, 2023-
Map releaseNov 29, 2023-
UpdateMar 27, 2024-
Current statusMar 27, 2024Processing site: PDBe / Status: Released

-
Structure visualization

Supplemental images

emd_17973.png

Downloads & links

-
Map

FileDownload / File: emd_17973.map.gz / Format: CCP4 / Size: 27 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationphenix.auto_sharped of cryosparc local refinement map
Voxel sizeX=Y=Z: 1.1 Å
Density
Contour LevelBy AUTHOR: 6.0
Minimum - Maximum-18.852810000000002 - 42.545949999999998
Average (Standard dev.)0.000000000002529 (±1.0)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderZYX
Origin000
Dimensions192192192
Spacing192192192
CellA=B=C: 211.20001 Å
α=β=γ: 90.0 °

-
Supplemental data

-
Half map: cryosparc local refinement half-map

Fileemd_17973_half_map_1.map
Annotationcryosparc local refinement half-map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

-
Half map: cryosparc local refinement half-map

Fileemd_17973_half_map_2.map
Annotationcryosparc local refinement half-map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

-
Sample components

+
Entire : AfAgo and AfAgo-N complex with 30 nt RNA guide and 51 nt DNA target

EntireName: AfAgo and AfAgo-N complex with 30 nt RNA guide and 51 nt DNA target
Components
  • Complex: AfAgo and AfAgo-N complex with 30 nt RNA guide and 51 nt DNA target
    • Complex: Piwi protein
      • Protein or peptide: Piwi protein
    • Complex: Archaeoglobus fulgidus AfAgo-N protein
      • Protein or peptide: Archaeoglobus fulgidus AfAgo-N protein
    • Complex: RNA (30-MER)
      • RNA: RNA (30-MER)
    • Complex: DNA (51-MER)
      • DNA: DNA (51-MER)
  • Ligand: MAGNESIUM ION
  • Ligand: water

+
Supramolecule #1: AfAgo and AfAgo-N complex with 30 nt RNA guide and 51 nt DNA target

SupramoleculeName: AfAgo and AfAgo-N complex with 30 nt RNA guide and 51 nt DNA target
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#4
Molecular weightTheoretical: 90 KDa

+
Supramolecule #2: Piwi protein

SupramoleculeName: Piwi protein / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1
Source (natural)Organism: Archaeoglobus fulgidus (archaea)

+
Supramolecule #3: Archaeoglobus fulgidus AfAgo-N protein

SupramoleculeName: Archaeoglobus fulgidus AfAgo-N protein / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #2
Source (natural)Organism: Archaeoglobus fulgidus DSM 8774 (archaea)

+
Supramolecule #4: RNA (30-MER)

SupramoleculeName: RNA (30-MER) / type: complex / ID: 4 / Parent: 1 / Macromolecule list: #3
Source (natural)Organism: Escherichia coli (E. coli)

+
Supramolecule #5: DNA (51-MER)

SupramoleculeName: DNA (51-MER) / type: complex / ID: 5 / Parent: 1 / Macromolecule list: #4
Source (natural)Organism: Escherichia coli (E. coli)

+
Macromolecule #1: Piwi protein

MacromoleculeName: Piwi protein / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Archaeoglobus fulgidus (archaea) / Strain: DSM4304
Molecular weightTheoretical: 49.302434 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MMEYKIVENG LTYRIGNGAS VPISNTGELI KGLRNYGPYE VPSLKYNQIA LIHNNQFSSL INQLKSQISS KIDEVWHIHN INISEFIYD SPHFDSIKSQ VDNAIDTGVD GIMLVLPEYN TPLYYKLKSY LINSIPSQFM RYDILSNRNL TFYVDNLLVQ F VSKLGGKP ...String:
MMEYKIVENG LTYRIGNGAS VPISNTGELI KGLRNYGPYE VPSLKYNQIA LIHNNQFSSL INQLKSQISS KIDEVWHIHN INISEFIYD SPHFDSIKSQ VDNAIDTGVD GIMLVLPEYN TPLYYKLKSY LINSIPSQFM RYDILSNRNL TFYVDNLLVQ F VSKLGGKP WILNVDPEKG SDIIIGTGAT RIDNVNLFCF AMVFKKDGTM LWNEISPIVT SSEYLTYLKS TIKKVVYGFK KS NPDWDVE KLTLHVSGKR PKMKDGETKI LKETVEELKK QEMVSRDVKY AILHLNETHP FWVMGDPNNR FHPYEGTKVK LSS KRYLLT LLQPYLKRNG LEMVTPIKPL SVEIVSDNWT SEEYYHNVHE ILDEIYYLSK MNWRGFRSRN LPVTVNYPKL VAGI IANVN RYGGYPINPE GNRSLQTNPW FL

UniProtKB: Piwi protein

+
Macromolecule #2: Archaeoglobus fulgidus AfAgo-N protein

MacromoleculeName: Archaeoglobus fulgidus AfAgo-N protein / type: protein_or_peptide / ID: 2 / Details: N-terminal His tag for purification / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Archaeoglobus fulgidus DSM 8774 (archaea)
Molecular weightTheoretical: 31.356576 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MGGSHHHHHH GMASENLYFQ GGGGEIPLSS GNVNTPDVRS SGILYINIYP IVNYPETIKV SAIPYYEEFL PGKWKKRIGD LIYLYGYGI ENEFDEIDNS NALFGKIFRK YLLDILSENI ATPWQLKELG STLRLVKEIT ENYEFSNIIK LQYELIINVH H WQNTNFGI ...String:
MGGSHHHHHH GMASENLYFQ GGGGEIPLSS GNVNTPDVRS SGILYINIYP IVNYPETIKV SAIPYYEEFL PGKWKKRIGD LIYLYGYGI ENEFDEIDNS NALFGKIFRK YLLDILSENI ATPWQLKELG STLRLVKEIT ENYEFSNIIK LQYELIINVH H WQNTNFGI IVDLKINILD RENNQRISYT KIKDKYGESV KKKIWVSVQA FHRHLTPEGK KYATAMRDKF NLLTGLLKEA FG SSEDEKT FSTPDGEIKI VFKPLEIVEV SNNDGI

UniProtKB: Uncharacterized protein

+
Macromolecule #3: RNA (30-MER)

MacromoleculeName: RNA (30-MER) / type: rna / ID: 3 / Number of copies: 1
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 9.792971 KDa
SequenceString:
AGGAGGGCGG AGCCUAUGGA AAAACGCCAC

+
Macromolecule #4: DNA (51-MER)

MacromoleculeName: DNA (51-MER) / type: dna / ID: 4 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 15.579906 KDa
SequenceString:
(DG)(DC)(DC)(DG)(DC)(DG)(DC)(DC)(DG)(DG) (DT)(DG)(DG)(DC)(DG)(DT)(DT)(DT)(DT)(DT) (DC)(DC)(DA)(DT)(DA)(DG)(DG)(DC)(DT) (DC)(DC)(DG)(DC)(DC)(DC)(DT)(DC)(DC)(DT) (DG) (DC)(DC)(DA)(DG)(DA)(DG)(DT)(DT) (DC)(DG)(DC)

+
Macromolecule #5: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 5 / Number of copies: 1 / Formula: MG
Molecular weightTheoretical: 24.305 Da

+
Macromolecule #6: water

MacromoleculeName: water / type: ligand / ID: 6 / Number of copies: 36 / Formula: HOH
Molecular weightTheoretical: 18.015 Da
Chemical component information

ChemComp-HOH:
WATER / Water

-
Experimental details

-
Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

-
Sample preparation

Concentration1 mg/mL
BufferpH: 8.5
Component:
ConcentrationFormulaName
125.0 mMKClKCl
20.0 mMC4H11NO3Tris hydrochloride pH8.5
2.0 mMMgCl2MgCl2
2.0 mMC4H10O2S2DTT

Details: fAfAgo complex with 30/51 guide-target heteroduplexes was mixed and applied on grid
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 45 sec. / Pretreatment - Atmosphere: AIR
VitrificationCryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV
DetailsfAfAgo complex with 30/51 guide-target heteroduplexes was mixed and applied on grid

-
Electron microscopy

MicroscopeTFS GLACIOS
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 100.0 µm / Illumination mode: SPOT SCAN / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 2.0 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 92000
Sample stageSpecimen holder model: OTHER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: FEI FALCON III (4k x 4k) / Detector mode: COUNTING / Digitization - Dimensions - Width: 4000 pixel / Digitization - Dimensions - Height: 4000 pixel / Number grids imaged: 1 / Number real images: 2152 / Average exposure time: 46.33 sec. / Average electron dose: 31.0 e/Å2

-
Image processing

Particle selectionNumber selected: 2326555 / Details: Blob particle picking
Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

8ok9


Details: Protein only chains
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. v.4.2.1)
Final 3D classificationNumber classes: 105 / Avg.num./class: 14000 / Software - Name: cryoSPARC (ver. v.4.2.1)
Details: Particles of selected classes were further sorted by heterogenous refinement
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. v.4.2.1)
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 2.81 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. v.4.2.1) / Number images used: 498038
FSC plot (resolution estimation)

-
Atomic model buiding 1

Initial model
PDB IDChainDetails

8ok9

chain_id: A, residue_range: 1-427, source_name: PDB, initial_model_type: experimental modelfAfAgo complex was used as initial model

8ok9

chain_id: C, residue_range: 20-245, source_name: PDB, initial_model_type: experimental modelfAfAgo complex was used as initial model
RefinementSpace: REAL / Protocol: RIGID BODY FIT
Output model

PDB-8pvv:
Archaeoglobus fulgidus AfAgo complex with AfAgo-N protein (fAfAgo) bound with 30 nt RNA guide and 51 nt DNA target

+
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