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- EMDB-45752: Structure of human Argonaute2-guide-target complex in a fully pai... -

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Basic information

Entry
Database: EMDB / ID: EMD-45752
TitleStructure of human Argonaute2-guide-target complex in a fully paired, slicing-competent conformation
Map data
Sample
  • Complex: miR7-HsAGO2 RISC
    • RNA: RNA (5'-R(P*UP*GP*GP*AP*AP*GP*AP*CP*UP*AP*GP*UP*GP*AP*UP*UP*UP*UP*GP*UP*U)-3')
    • RNA: RNA (5'-R(*CP*AP*AP*CP*AP*AP*AP*AP*UP*CP*AP*CP*UP*AP*GP*UP*CP*UP*UP*CP*CP*A)-3')
    • Protein or peptide: Protein argonaute-2
  • Ligand: MAGNESIUM ION
KeywordsRNAi / Argonaute / Slicing / HYDROLASE-RNA complex
Function / homology
Function and homology information


: / endoribonuclease activity, cleaving miRNA-paired mRNA / endoribonuclease activity, cleaving siRNA-paired mRNA / siRNA-mediated gene silencing by mRNA destabilization / miRNA-mediated gene silencing by mRNA destabilization / Post-transcriptional silencing by small RNAs / Competing endogenous RNAs (ceRNAs) regulate PTEN translation / Regulation of CDH11 mRNA translation by microRNAs / Regulation of NPAS4 mRNA translation / Regulation of PTEN mRNA translation ...: / endoribonuclease activity, cleaving miRNA-paired mRNA / endoribonuclease activity, cleaving siRNA-paired mRNA / siRNA-mediated gene silencing by mRNA destabilization / miRNA-mediated gene silencing by mRNA destabilization / Post-transcriptional silencing by small RNAs / Competing endogenous RNAs (ceRNAs) regulate PTEN translation / Regulation of CDH11 mRNA translation by microRNAs / Regulation of NPAS4 mRNA translation / Regulation of PTEN mRNA translation / negative regulation of amyloid precursor protein biosynthetic process / Small interfering RNA (siRNA) biogenesis / positive regulation of trophoblast cell migration / Transcriptional Regulation by MECP2 / miRNA metabolic process / RISC-loading complex / mRNA cap binding / regulatory ncRNA-mediated post-transcriptional gene silencing / RISC complex assembly / miRNA processing / miRNA-mediated gene silencing by inhibition of translation / pre-miRNA processing / RNA 7-methylguanosine cap binding / siRNA processing / regulation of synapse maturation / siRNA binding / Regulation of MITF-M-dependent genes involved in apoptosis / mRNA 3'-UTR AU-rich region binding / M-decay: degradation of maternal mRNAs by maternally stored factors / RISC complex / regulatory ncRNA-mediated gene silencing / TGFBR3 expression / Regulation of RUNX1 Expression and Activity / P-body assembly / miRNA binding / positive regulation of nuclear-transcribed mRNA catabolic process, deadenylation-dependent decay / MicroRNA (miRNA) biogenesis / positive regulation of nuclear-transcribed mRNA poly(A) tail shortening / RNA polymerase II complex binding / Regulation of MECP2 expression and activity / core promoter sequence-specific DNA binding / Nuclear events stimulated by ALK signaling in cancer / NR1H3 & NR1H2 regulate gene expression linked to cholesterol transport and efflux / negative regulation of translational initiation / translation initiation factor activity / RNA endonuclease activity / post-embryonic development / positive regulation of translation / TP53 Regulates Metabolic Genes / Transcriptional regulation by small RNAs / P-body / MAPK6/MAPK4 signaling / Pre-NOTCH Transcription and Translation / cytoplasmic ribonucleoprotein granule / positive regulation of angiogenesis / double-stranded RNA binding / Ca2+ pathway / Estrogen-dependent gene expression / single-stranded RNA binding / postsynapse / translation / dendrite / glutamatergic synapse / positive regulation of transcription by RNA polymerase II / RNA binding / extracellular exosome / nucleoplasm / metal ion binding / nucleus / membrane / cytosol / cytoplasm
Similarity search - Function
Protein argonaute-2 / Protein argonaute, Mid domain / Mid domain of argonaute / Argonaute linker 2 domain / Protein argonaute, N-terminal / Argonaute-like, PIWI domain / N-terminal domain of argonaute / Argonaute linker 2 domain / DUF1785 / Argonaute, linker 1 domain ...Protein argonaute-2 / Protein argonaute, Mid domain / Mid domain of argonaute / Argonaute linker 2 domain / Protein argonaute, N-terminal / Argonaute-like, PIWI domain / N-terminal domain of argonaute / Argonaute linker 2 domain / DUF1785 / Argonaute, linker 1 domain / Argonaute linker 1 domain / Piwi domain profile. / Piwi domain / Piwi domain / Piwi / PAZ domain superfamily / PAZ / PAZ domain / PAZ domain profile. / PAZ domain / Ribonuclease H superfamily / Ribonuclease H-like superfamily
Similarity search - Domain/homology
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.3 Å
AuthorsMohamed AA / Wang PY / Bartel DP / Vos SM
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)DP2 GM146254 United States
CitationJournal: bioRxiv / Year: 2024
Title: The structural basis for RNA slicing by human Argonaute2.
Authors: Abdallah A Mohamed / Peter Y Wang / David P Bartel / Seychelle M Vos /
Abstract: Argonaute (AGO) proteins associate with guide RNAs to form complexes that slice transcripts that pair to the guide. This slicing drives post-transcriptional gene-silencing pathways that are essential ...Argonaute (AGO) proteins associate with guide RNAs to form complexes that slice transcripts that pair to the guide. This slicing drives post-transcriptional gene-silencing pathways that are essential for many eukaryotes and the basis for new clinical therapies. Despite this importance, structural information on eukaryotic AGOs in a fully paired, slicing-competent conformation-hypothesized to be intrinsically unstable-has been lacking. Here we present the cryogenic-electron microscopy structure of a human AGO-guide complex bound to a fully paired target, revealing structural rearrangements that enable this conformation. Critically, the N domain of AGO rotates to allow the RNA full access to the central channel and forms contacts that license rapid slicing. Moreover, a conserved loop in the PIWI domain secures the RNA near the active site to enhance slicing rate and specificity. These results explain how AGO accommodates targets possessing the pairing specificity typically observed in biological and clinical slicing substrates.
History
DepositionJul 15, 2024-
Header (metadata) releaseDec 11, 2024-
Map releaseDec 11, 2024-
UpdateMay 14, 2025-
Current statusMay 14, 2025Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_45752.png

Downloads & links

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Map

FileDownload / File: emd_45752.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.65 Å/pix.
x 300 pix.
= 196.2 Å		0.65 Å/pix.
x 300 pix.
= 196.2 Å		0.65 Å/pix.
x 300 pix.
= 196.2 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.654 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.338
Minimum - Maximum-2.0067601 - 3.60319
Average (Standard dev.)0.004738944 (±0.09351491)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions300300300
Spacing300300300
CellA=B=C: 196.2 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_45752_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_45752_half_map_1.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_45752_half_map_2.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : miR7-HsAGO2 RISC

EntireName: miR7-HsAGO2 RISC
Components
  • Complex: miR7-HsAGO2 RISC
    • RNA: RNA (5'-R(P*UP*GP*GP*AP*AP*GP*AP*CP*UP*AP*GP*UP*GP*AP*UP*UP*UP*UP*GP*UP*U)-3')
    • RNA: RNA (5'-R(*CP*AP*AP*CP*AP*AP*AP*AP*UP*CP*AP*CP*UP*AP*GP*UP*CP*UP*UP*CP*CP*A)-3')
    • Protein or peptide: Protein argonaute-2
  • Ligand: MAGNESIUM ION

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Supramolecule #1: miR7-HsAGO2 RISC

SupramoleculeName: miR7-HsAGO2 RISC / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#3
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 113 KDa

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Macromolecule #1: RNA (5'-R(P*UP*GP*GP*AP*AP*GP*AP*CP*UP*AP*GP*UP*GP*AP*UP*UP*UP*UP...

MacromoleculeName: RNA (5'-R(P*UP*GP*GP*AP*AP*GP*AP*CP*UP*AP*GP*UP*GP*AP*UP*UP*UP*UP*GP*UP*U)-3')
type: rna / ID: 1 / Number of copies: 1
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 7.078188 KDa
SequenceString:
UGGAAGACUA GUGAUUUUGU UG

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Macromolecule #2: RNA (5'-R(*CP*AP*AP*CP*AP*AP*AP*AP*UP*CP*AP*CP*UP*AP*GP*UP*CP*UP*...

MacromoleculeName: RNA (5'-R(*CP*AP*AP*CP*AP*AP*AP*AP*UP*CP*AP*CP*UP*AP*GP*UP*CP*UP*UP*CP*CP*A)-3')
type: rna / ID: 2 / Number of copies: 1
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 8.813276 KDa
SequenceString:
UUUCAACAAA AUCACUAGUC UUCCAAAU

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Macromolecule #3: Protein argonaute-2

MacromoleculeName: Protein argonaute-2 / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
EC number: Hydrolases; Acting on ester bonds; Endoribonucleases producing 5'-phosphomonoesters
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 97.319102 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: GSYSGAGPAL APPAPPPPIQ GYAFKPPPRP DFGTSGRTIK LQANFFEMDI PKIDIYHYEL DIKPEKCPRR VNREIVEHMV QHFKTQIFG DRKPVFDGRK NLYTAMPLPI GRDKVELEVT LPGEGKDRIF KVSIKWVSCV SLQALHDALS GRLPSVPFET I QALDVVMR ...String:
GSYSGAGPAL APPAPPPPIQ GYAFKPPPRP DFGTSGRTIK LQANFFEMDI PKIDIYHYEL DIKPEKCPRR VNREIVEHMV QHFKTQIFG DRKPVFDGRK NLYTAMPLPI GRDKVELEVT LPGEGKDRIF KVSIKWVSCV SLQALHDALS GRLPSVPFET I QALDVVMR HLPSMRYTPV GRSFFTASEG CSNPLGGGRE VWFGFHQSVR PSLWKMMLNI DVSATAFYKA QPVIEFVCEV LD FKSIEEQ QKPLTDSQRV KFTKEIKGLK VEITHCGQMK RKYRVCNVTR RPASHQTFPL QQESGQTVEC TVAQYFKDRH KLV LRYPHL PCLQVGQEQK HTYLPLEVCN IVAGQRCIKK LTDNQTSTMI RATARSAPDR QEEISKLMRS ASFNTDPYVR EFGI MVKDE MTDVTGRVLQ PPSILYGGRN KAIATPVQGV WDMRNKQFHT GIEIKVWAIA CFAPQRQCTE VHLKSFTEQL RKISR DAGM PIQGQPCFCK YAQGADSVEP MFRHLKNTYA GLQLVVVILP GKTPVYAEVK RVGDTVLGMA TQCVQMKNVQ RTTPQT LSN LCLKINVKLG GVNNILLPQG RPPVFQQPVI FLGADVTHPP AGDGKKPSIA AVVGSMDAHP NRYCATVRVQ QHRQEII QD LAAMVRELLI QFYKSTRFKP TRIIFYRAGV SEGQFQQVLH HELLAIREAC IKLEKDYQPG ITFIVVQKRH HTRLFCTD K NERVGKSGNI PAGTTVDTKI THPTEFDFYL CSHAGIQGTS RPSHYHVLWD DNRFSSDELQ ILTYQLCHTY VRCTRSVSI PAPAYYAHLV AFRARYHLVD KEHDSAEGSH TSGQSNGRDH QALAKAVQVH QDTLRTMYFA

UniProtKB: Protein argonaute-2

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Macromolecule #4: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 4 / Number of copies: 1 / Formula: MG
Molecular weightTheoretical: 24.305 Da

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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

BufferpH: 7.5
Component:
ConcentrationNameFormula
50.0 mMTris
150.0 mMsodium chlorideNaCl
1.0 mMmagnesium chlorideMgCl2
GridModel: Quantifoil R2/2 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: CONTINUOUS / Support film - Film thickness: 0.2 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 12 sec. / Pretreatment - Atmosphere: OTHER
VitrificationCryogen name: ETHANE / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeTFS KRIOS
Specialist opticsEnergy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 51.0 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: 1.4000000000000001 µm / Nominal defocus min: 0.2 µm / Nominal magnification: 130000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 1427326
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: INSILICO MODEL
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.3 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 69008
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: AB INITIO MODEL
Output model

PDB-9cmp:
Structure of human Argonaute2-guide-target complex in a fully paired, slicing-competent conformation

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