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- PDB-21cq: Cryo-EM structure of DICER-D991G-H992G/26S-GU complex in dicing state -

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Basic information

Entry
Database: PDB / ID: 21cq
TitleCryo-EM structure of DICER-D991G-H992G/26S-GU complex in dicing state
Components
  • Endoribonuclease Dicer
  • RNA (64-MER)
KeywordsHYDROLASE/RNA / Endoribonuclease Dicer / RNA / miRNA / precursor miRNA / HYDROLASE / HYDROLASE-RNA complex
Function / homology
Function and homology information


peripheral nervous system myelin formation / global gene silencing by mRNA cleavage / negative regulation of Schwann cell proliferation / pre-miRNA binding / tRNA-derived small RNA (tsRNA or tRNA-related fragment, tRF) biogenesis / Small interfering RNA (siRNA) biogenesis / tRNA decay / apoptotic DNA fragmentation / ribonuclease III / deoxyribonuclease I activity ...peripheral nervous system myelin formation / global gene silencing by mRNA cleavage / negative regulation of Schwann cell proliferation / pre-miRNA binding / tRNA-derived small RNA (tsRNA or tRNA-related fragment, tRF) biogenesis / Small interfering RNA (siRNA) biogenesis / tRNA decay / apoptotic DNA fragmentation / ribonuclease III / deoxyribonuclease I activity / nerve development / positive regulation of Schwann cell differentiation / RISC-loading complex / positive regulation of myelination / miRNA metabolic process / RISC complex assembly / miRNA processing / ribonuclease III activity / pre-miRNA processing / siRNA processing / siRNA binding / M-decay: degradation of maternal mRNAs by maternally stored factors / Regulation of MITF-M-dependent genes involved in apoptosis / RISC complex / MicroRNA (miRNA) biogenesis / negative regulation of tumor necrosis factor production / negative regulation of tumor necrosis factor-mediated signaling pathway / neuron projection morphogenesis / RNA endonuclease activity / helicase activity / double-stranded RNA binding / protein domain specific binding / negative regulation of gene expression / perinuclear region of cytoplasm / negative regulation of transcription by RNA polymerase II / RNA binding / extracellular exosome / ATP binding / metal ion binding / nucleus / cytoplasm / cytosol
Similarity search - Function
Dicer, double-stranded RNA-binding domain / Dicer, partner-binding domain / Dicer, dsRNA-binding domain / : / : / Dicer, platform domain / Dicer dimerisation domain / Dicer dimerisation domain superfamily / Dicer dimerisation domain / Dicer double-stranded RNA-binding fold domain profile. ...Dicer, double-stranded RNA-binding domain / Dicer, partner-binding domain / Dicer, dsRNA-binding domain / : / : / Dicer, platform domain / Dicer dimerisation domain / Dicer dimerisation domain superfamily / Dicer dimerisation domain / Dicer double-stranded RNA-binding fold domain profile. / Ribonuclease III family signature. / Ribonuclease III domain / Ribonuclease III family domain profile. / Ribonuclease III family / Ribonuclease III domain / PAZ domain superfamily / PAZ / PAZ domain / PAZ domain profile. / PAZ domain / Double-stranded RNA binding motif / Ribonuclease III, endonuclease domain superfamily / Double stranded RNA-binding domain (dsRBD) profile. / Double-stranded RNA-binding domain / DEAD/DEAH box helicase domain / DEAD/DEAH box helicase / Helicase conserved C-terminal domain / helicase superfamily c-terminal domain / Superfamilies 1 and 2 helicase C-terminal domain profile. / Superfamilies 1 and 2 helicase ATP-binding type-1 domain profile. / DEAD-like helicases superfamily / Helicase, C-terminal / Helicase superfamily 1/2, ATP-binding domain / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
RNA / RNA (> 10) / Endoribonuclease Dicer
Similarity search - Component
Biological speciesHomo sapiens (human)
synthetic construct (others)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.29 Å
AuthorsNgo, M.K.
Funding support Hong Kong, 1items
OrganizationGrant numberCountry
Other government16103023 Hong Kong
CitationJournal: Nature / Year: 2026
Title: DICER cleavage fidelity is governed by 5'-end binding pockets.
Authors: Minh Khoa Ngo / Cong Truc Le / Tuan Anh Nguyen /
Abstract: RNA interference (RNAi) depends on DICER, an essential enzyme that processes RNA precursors into small regulatory RNAs. DICER cleaves RNA precursors according to the 5'-end counting rule, in which ...RNA interference (RNAi) depends on DICER, an essential enzyme that processes RNA precursors into small regulatory RNAs. DICER cleaves RNA precursors according to the 5'-end counting rule, in which RNA length is measured from the 5'-end. Previous work proposed a single 5'-end binding pocket that disfavours guanosine (5'-G), leading to cleavage inaccuracies. Here we show that 5'-G promotes precise cleavage for many substrates. Using massively parallel dicing assays and cryo-electron microscopy, we identify a conserved guanosine-favoured (G-favoured) binding pocket in DICER, distinct from the previously described uridine-favoured (U-favoured) pocket. Together, these pockets influence the alignment between 21-nucleotide and 22-nucleotide cleavage registers, expanding the mechanism of small-RNA biogenesis in metazoan DICERs. We also find that conflicts between 5'-end binding and RNA-motif recognition can trigger RNA conformational adjustments that preserve accurate cleavage-site selection. In addition, conformational adjustments of the double-stranded RNA-binding domain (dsRBD) and PAZ domain help to align substrates with the catalytic centres for precise double-strand cleavage. These results show that the DICER cleavage mechanism integrates dual 5'-end binding pockets, RNA-motif influence and domain motions, advancing our understanding of microRNA biogenesis.
History
DepositionDec 8, 2025Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Mar 11, 2026Provider: repository / Type: Initial release
Revision 1.0Mar 11, 2026Data content type: EM metadata / Data content type: EM metadata / Provider: repository / Type: Initial release
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Revision 1.1Mar 18, 2026Group: Data collection / Database references / Category: citation / citation_author / em_admin
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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
R: RNA (64-MER)
D: Endoribonuclease Dicer
hetero molecules


Theoretical massNumber of molelcules
Total (without water)238,4754
Polymers238,3952
Non-polymers802
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: RNA chain RNA (64-MER)


Mass: 20545.252 Da / Num. of mol.: 1 / Source method: obtained synthetically / Details: 26S-GU / Source: (synth.) synthetic construct (others)
#2: Protein Endoribonuclease Dicer / Helicase with RNase motif / Helicase MOI


Mass: 217849.875 Da / Num. of mol.: 1 / Mutation: D991G, H992G
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: DICER1, DICER, HERNA, KIAA0928 / Production host: Homo sapiens (human) / References: UniProt: Q9UPY3, ribonuclease III
#3: Chemical ChemComp-CA / CALCIUM ION


Mass: 40.078 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Ca
Has ligand of interestN
Has protein modificationN

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Complex of DICER-D991G-H992G/26S-GU in dicing state / Type: COMPLEX
Details: Recombinant DICER with mutations D991G -H992G and chemically synthesis shRNA forming complex in dicing-competent state
Entity ID: #1-#2 / Source: MULTIPLE SOURCES
Molecular weightExperimental value: NO
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Homo sapiens (human)
Buffer solutionpH: 8
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: OTHER / Nominal magnification: 81000 X / Calibrated magnification: 81000 X / Nominal defocus max: 2400 nm / Nominal defocus min: 1000 nm
Image recordingAverage exposure time: 3.1 sec. / Electron dose: 50 e/Å2 / Film or detector model: TFS FALCON 4i (4k x 4k) / Num. of grids imaged: 2 / Num. of real images: 16972

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Processing

EM software
IDNameVersionCategory
1cryoSPARC4.6.2particle selection
2EPUimage acquisition
4cryoSPARC4.6.2CTF correction
7Coot0.9.8.96model fitting
11cryoSPARC4.6.2classification
12cryoSPARC4.6.23D reconstruction
13PHENIX1.20.1model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.29 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 787381 / Symmetry type: POINT
Atomic model buildingProtocol: OTHER
RefinementHighest resolution: 3.29 Å
Stereochemistry target values: REAL-SPACE (WEIGHTED MAP SUM AT ATOM CENTERS)
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0057053
ELECTRON MICROSCOPYf_angle_d0.7699814
ELECTRON MICROSCOPYf_dihedral_angle_d14.4921417
ELECTRON MICROSCOPYf_chiral_restr0.0421138
ELECTRON MICROSCOPYf_plane_restr0.0061053

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