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- EMDB-49593: Cryo-EM structure of Csm/AcrIIIA2/enolase 3:2 complex -

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Basic information

Entry
Database: EMDB / ID: EMD-49593
TitleCryo-EM structure of Csm/AcrIIIA2/enolase 3:2 complex
Map dataCryo-EM map of Csm-AcrIIIA2-enolase complex from Streptococcus thermophilus
Sample
  • Complex: Csm/AcrIIIA2/enolase complex
    • Protein or peptide: CRISPR system single-strand-specific deoxyribonuclease Cas10/Csm1 (subtype III-A)
    • Protein or peptide: CRISPR system Cms protein Csm4
    • Protein or peptide: CRISPR system Cms endoribonuclease Csm3
    • Protein or peptide: CRISPR system Cms endoribonuclease Csm3
    • RNA: crRNA RNA (35-MER)
    • Protein or peptide: CRISPR system Cms protein Csm2
    • Protein or peptide: CRISPR system Cms protein Csm5
    • Protein or peptide: Enolase
    • Protein or peptide: AcrIIIA2
  • Ligand: MAGNESIUM ION
KeywordsANTI-CRISPR / TYPE III CRISPR / CRYO-EM / STRUCTURAL BIOLOY / ANTI-BACTERIAL / RNA BINDING PROTEIN / RNA BINDING PROTEIN-RNA complex
Function / homology
Function and homology information


phosphopyruvate hydratase / phosphopyruvate hydratase complex / phosphopyruvate hydratase activity / exonuclease activity / peptidoglycan-based cell wall / glycolytic process / Transferases; Transferring phosphorus-containing groups; Nucleotidyltransferases / transferase activity / endonuclease activity / defense response to virus ...phosphopyruvate hydratase / phosphopyruvate hydratase complex / phosphopyruvate hydratase activity / exonuclease activity / peptidoglycan-based cell wall / glycolytic process / Transferases; Transferring phosphorus-containing groups; Nucleotidyltransferases / transferase activity / endonuclease activity / defense response to virus / Hydrolases; Acting on ester bonds / hydrolase activity / magnesium ion binding / cell surface / RNA binding / extracellular region / ATP binding
Similarity search - Function
: / CRISPR RNA silencing complex Cmr2 subunit, second helical domain / Csm4, C-terminal / CRISPR Csm4 C-terminal domain / CRISPR-associated protein Csm5 / CRISPR-associated protein, Csm2 Type III-A / Csm2 Type III-A / : / CRISPR-associated RAMP Csm3 / CRISPR type III-associated RAMP protein Csm4 ...: / CRISPR RNA silencing complex Cmr2 subunit, second helical domain / Csm4, C-terminal / CRISPR Csm4 C-terminal domain / CRISPR-associated protein Csm5 / CRISPR-associated protein, Csm2 Type III-A / Csm2 Type III-A / : / CRISPR-associated RAMP Csm3 / CRISPR type III-associated RAMP protein Csm4 / CRISPR system single-strand-specific deoxyribonuclease Cas10/Csm1 / Csm1, subunit domain B / Csm1 subunit domain B / : / Cas10/Cmr2, second palm domain / Enolase / Enolase, conserved site / Enolase, C-terminal TIM barrel domain / Enolase, N-terminal / Enolase, C-terminal TIM barrel domain / Enolase, N-terminal domain / Enolase signature. / Enolase, C-terminal TIM barrel domain / Enolase, N-terminal domain / : / CRISPR type III-associated protein / RAMP superfamily / HD domain / GGDEF domain profile. / GGDEF domain / HD domain / Enolase-like, N-terminal / Enolase-like, C-terminal domain superfamily / Reverse transcriptase/Diguanylate cyclase domain
Similarity search - Domain/homology
CRISPR system Cms protein Csm5 / CRISPR system single-strand-specific deoxyribonuclease Cas10/Csm1 (subtype III-A) / Uncharacterized protein / CRISPR system Cms protein Csm4 / CRISPR system Cms protein Csm2 / CRISPR system Cms endoribonuclease Csm3 / Enolase
Similarity search - Component
Biological speciesStreptococcus thermophilus (bacteria) / Streptococcus phage 2972 (virus)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.67 Å
AuthorsGoswami HN / Li H
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35 GM152081 United States
CitationJournal: To Be Published
Title: Cryo-EM structure of Csm/AcrIIIA2/enolase 3:2 complex
Authors: Goswami HN / Li H
History
DepositionMar 7, 2025-
Header (metadata) releaseNov 12, 2025-
Map releaseNov 12, 2025-
UpdateNov 12, 2025-
Current statusNov 12, 2025Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_49593.png

Downloads & links

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Map

FileDownload / File: emd_49593.map.gz / Format: CCP4 / Size: 669.9 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationCryo-EM map of Csm-AcrIIIA2-enolase complex from Streptococcus thermophilus
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.83 Å/pix.
x 560 pix.
= 463.68 Å		0.83 Å/pix.
x 560 pix.
= 463.68 Å		0.83 Å/pix.
x 560 pix.
= 463.68 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.828 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.216
Minimum - Maximum-0.56451434 - 1.5861754
Average (Standard dev.)0.008268404 (±0.03078575)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions560560560
Spacing560560560
CellA=B=C: 463.68 Å
α=β=γ: 90.0 °

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Supplemental data

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Sample components

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Entire : Csm/AcrIIIA2/enolase complex

EntireName: Csm/AcrIIIA2/enolase complex
Components
  • Complex: Csm/AcrIIIA2/enolase complex
    • Protein or peptide: CRISPR system single-strand-specific deoxyribonuclease Cas10/Csm1 (subtype III-A)
    • Protein or peptide: CRISPR system Cms protein Csm4
    • Protein or peptide: CRISPR system Cms endoribonuclease Csm3
    • Protein or peptide: CRISPR system Cms endoribonuclease Csm3
    • RNA: crRNA RNA (35-MER)
    • Protein or peptide: CRISPR system Cms protein Csm2
    • Protein or peptide: CRISPR system Cms protein Csm5
    • Protein or peptide: Enolase
    • Protein or peptide: AcrIIIA2
  • Ligand: MAGNESIUM ION

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Supramolecule #1: Csm/AcrIIIA2/enolase complex

SupramoleculeName: Csm/AcrIIIA2/enolase complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #9, #1-#8
Details: AcrIIIA2 forms a stable ternary complex with the type III-A (Csm) crRNP (CRISPR RNA-Protein complex) and host enolase
Source (natural)Organism: Streptococcus thermophilus (bacteria)
Molecular weightTheoretical: 680 KDa

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Macromolecule #1: CRISPR system Cms protein Csm4

MacromoleculeName: CRISPR system Cms protein Csm4 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Streptococcus thermophilus (bacteria)
Molecular weightTheoretical: 33.828984 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MTYKLYIMTF QNAHFGSGTL DSSKLTFSAD RIFSALVLEA LKMGKLDAFL AEANQDKFTL TDAFPFQFGP FLPKPIGYPK HDQIDQSVD VKEVRRQAKL SKKLQFLALE NVDDYLNGEL FENEEHAVID TVTKNQPHKD DNLYQVATTR FSNDTSLYVI A NESDLLNE ...String:
MTYKLYIMTF QNAHFGSGTL DSSKLTFSAD RIFSALVLEA LKMGKLDAFL AEANQDKFTL TDAFPFQFGP FLPKPIGYPK HDQIDQSVD VKEVRRQAKL SKKLQFLALE NVDDYLNGEL FENEEHAVID TVTKNQPHKD DNLYQVATTR FSNDTSLYVI A NESDLLNE LMSSLQYSGL GGKRSSGFGR FELDIQNIPL ELSDRLTKNH SDKVMSLTTA LPVDADLEEA MEDGHYLLTK SS GFAFSHA TNENYRKQDL YKFASGSTFS KTFEGQIVDV RPLDFPHAVL NYAKPLFFKL EV

UniProtKB: CRISPR system Cms protein Csm4

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Macromolecule #2: CRISPR system Cms endoribonuclease Csm3

MacromoleculeName: CRISPR system Cms endoribonuclease Csm3 / type: protein_or_peptide / ID: 2 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Streptococcus thermophilus (bacteria)
Molecular weightTheoretical: 24.58584 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MTFAKIKFSA QIRLETGLHI GGSDAFAAIG AIDSPVIKDP ITNLPIIPGS SLKGKMRTLL AKVYNEKVAE KPSDDSDILS RLFGNSKDK RFKMGRLIFR DAFLSNADEL DSLGVRSYTE VKFENTIDRI TAEANPRQIE RAIRNSTFDF ELIYEITDEN E NQVEEDFK ...String:
MTFAKIKFSA QIRLETGLHI GGSDAFAAIG AIDSPVIKDP ITNLPIIPGS SLKGKMRTLL AKVYNEKVAE KPSDDSDILS RLFGNSKDK RFKMGRLIFR DAFLSNADEL DSLGVRSYTE VKFENTIDRI TAEANPRQIE RAIRNSTFDF ELIYEITDEN E NQVEEDFK VIRDGLKLLE LDYLGGSGSR GYGKVAFENL KATTVFGNYD VKTLNELLTA EV

UniProtKB: CRISPR system Cms endoribonuclease Csm3

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Macromolecule #3: CRISPR system Cms endoribonuclease Csm3

MacromoleculeName: CRISPR system Cms endoribonuclease Csm3 / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Streptococcus thermophilus (bacteria)
Molecular weightTheoretical: 24.555857 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MTFAKIKFSA QIRLETGLHI GGSDAFAAIG AIDSPVIKDP ITNLPIIPGS SLKGKMRTLL AKVYNEKVAE KPSDDSDILS RLFGNSKDK RFKMGRLIFR DAFLSNADEL DSLGVRSYTE VKFENTIDRI TAEANPRQIE RAIRNSTFDF ELIYEITDEN E NQVEEDFK ...String:
MTFAKIKFSA QIRLETGLHI GGSDAFAAIG AIDSPVIKDP ITNLPIIPGS SLKGKMRTLL AKVYNEKVAE KPSDDSDILS RLFGNSKDK RFKMGRLIFR DAFLSNADEL DSLGVRSYTE VKFENTIDRI TAEANPRQIE RAIRNSTFDF ELIYEITDEN E NQVEEDFK VIRDGLKLLE LDYLGGSGSR GYGKVAFENL KATTVFGNYD VKTLNELLTA VV

UniProtKB: CRISPR system Cms endoribonuclease Csm3

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Macromolecule #5: CRISPR system Cms protein Csm2

MacromoleculeName: CRISPR system Cms protein Csm2 / type: protein_or_peptide / ID: 5 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Streptococcus thermophilus (bacteria)
Molecular weightTheoretical: 15.361617 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
MAILTDENYV DKAERAISLL EKDNKGNYLL TTSQIRKLLS LCSSLYDRSK ERKFDELIND VSYLRVQFVY QSGRNSVRVN RQTFFPVKD LVEKGQILEA LKEIKDRETL QRFCRYMEAL VAYFKFYGGK D

UniProtKB: CRISPR system Cms protein Csm2

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Macromolecule #6: CRISPR system Cms protein Csm5

MacromoleculeName: CRISPR system Cms protein Csm5 / type: protein_or_peptide / ID: 6 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Streptococcus thermophilus (bacteria)
Molecular weightTheoretical: 41.226324 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MKNDYRTFKL SLLTLAPIHI GNGEKYTSRE FIYENKKFYF PDMGKFYNKM VEKRLAEKFE AFLIQTRPNA RNNRLISFLN DNRIAERSF GGYSISETGL ESDRNPNSAG AINEVNKFIR DAFGNPYIPG SSLKGAIRTI LMNTTPKWNN ENAVNDFGRF P KENKNLIP ...String:
MKNDYRTFKL SLLTLAPIHI GNGEKYTSRE FIYENKKFYF PDMGKFYNKM VEKRLAEKFE AFLIQTRPNA RNNRLISFLN DNRIAERSF GGYSISETGL ESDRNPNSAG AINEVNKFIR DAFGNPYIPG SSLKGAIRTI LMNTTPKWNN ENAVNDFGRF P KENKNLIP WGPKKGKEYD DLFNAIRVSD SKPFDNKRLI LVQKWDYSAK TNKAKPLPLY RESISPLTKI EFEITTTTDE AG RLIEELG KRAQAFYKDY KAFFLSEFPD DKIQANLQYP IYLGAGSGAW TKTLFKQADG ILQRRYSRMK TKMVKKGVLK LTK APLKIV KIPSGNHSLI KNHESFYEMG KANFMIKEID K

UniProtKB: CRISPR system Cms protein Csm5

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Macromolecule #7: Enolase

MacromoleculeName: Enolase / type: protein_or_peptide / ID: 7 / Number of copies: 8 / Enantiomer: LEVO / EC number: phosphopyruvate hydratase
Source (natural)Organism: Streptococcus thermophilus (bacteria)
Molecular weightTheoretical: 46.948484 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MSIITDVYAR EVLDSRGNPT LEVEVYTESG AFGRGMVPSG ASTGEHEAVE LRDGDKARYG GLGTQKAVDN VNNVIAEHII GFDVRDQQG IDRAMIALDG TPNKGKLGAN AILGVSIAVA RAAADYLEVP LYSYLGGFNT KVLPTPMMNI INGGSHSDAP I AFQEFMIV ...String:
MSIITDVYAR EVLDSRGNPT LEVEVYTESG AFGRGMVPSG ASTGEHEAVE LRDGDKARYG GLGTQKAVDN VNNVIAEHII GFDVRDQQG IDRAMIALDG TPNKGKLGAN AILGVSIAVA RAAADYLEVP LYSYLGGFNT KVLPTPMMNI INGGSHSDAP I AFQEFMIV PAGAPTFKEA LRWGAEIFHA LKKILKERGL ETAVGDEGGF APRFNGTEDG VETIIKAIEA AGYVPGKDVF IG LDCASSE FYDAEHKVYG YTKFEGEGAA VRTAAEQIDY LEELVNKYPI ITIEDGMDEN DWDGWKALTE RLGGKVQLVG DDF FVTNTA YLEKGIAEHA ANSILIKVNQ IGTLTETFDA IEMAKEAGYT AVVSHRSGET EDSTIADIAV ATNAGQIKTG SLSR TDRIA KYNQLLRIED QLGEVAEYRG LKSFYNLKK

UniProtKB: Enolase

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Macromolecule #8: AcrIIIA2

MacromoleculeName: AcrIIIA2 / type: protein_or_peptide / ID: 8 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Streptococcus phage 2972 (virus)
Molecular weightTheoretical: 12.226886 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
MKNFKISSTY RAARKQQKTA NRKSFYNDEG YMISPSEWAD GVIKGLINPK NSWSNDHVKG YLPRVSPRSH WTKNGYREYL GIGKSRDIP EKEPEVIEMM DLELVP

UniProtKB: Uncharacterized protein

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Macromolecule #9: CRISPR system single-strand-specific deoxyribonuclease Cas10/Csm1...

MacromoleculeName: CRISPR system single-strand-specific deoxyribonuclease Cas10/Csm1 (subtype III-A)
type: protein_or_peptide / ID: 9 / Details: Cas10 / Number of copies: 1 / Enantiomer: LEVO / EC number: Hydrolases; Acting on ester bonds
Source (natural)Organism: Streptococcus thermophilus (bacteria)
Molecular weightTheoretical: 86.930672 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MKKEKIDLFY GALLHDIGKV IQRATGERKK HALVGADWFD EIADNQVISD QIRYHMANYQ SDKLGNDHLA YITYIADNIA SGVDRRQSN EESDEDASAK IWDTYTNQAD IFNVFGAQTD KRYFKPTVLN LKSKPNFASA TYEPFSKGDY AAIATRIKNE L AEFEFNQA ...String:
MKKEKIDLFY GALLHDIGKV IQRATGERKK HALVGADWFD EIADNQVISD QIRYHMANYQ SDKLGNDHLA YITYIADNIA SGVDRRQSN EESDEDASAK IWDTYTNQAD IFNVFGAQTD KRYFKPTVLN LKSKPNFASA TYEPFSKGDY AAIATRIKNE L AEFEFNQA QIDSLLNLFE AILSFVPSST NSKEIADISL AEHSRLTAAF ALAIYDYLED KGRHNYKEDL FTKASAFYEE EA FLLASFD LSGIQDFIYN IATSGAAKQL KARSLYLDFM SEYIADSLLD KLGLNRANLL YVGGGHAYFV LANTEKTVET LVQ FEKDFN QFLLANFQTR LYVAFGWGSF AAKDIMSELN SPESYRQIYQ KASRMISEKK ISRYDYRTLM LLNRGGKSSE RECE ICHSV ENLVSYHDQK VCDICRGLYQ FSKEIAHDHF IITENEGLPI GPNACLKGVA FEKLSQESFS RVYVKNDYKA GTIKA THVF VGDYQCDEIH KYAALSKNED GLGIKRLAVV RLDVDDLGAA FMAGFSRQGN GQYSTLSRSA TFSRSMSLFF KVYINQ FAS DKKLSIIYAG GDDVFAIGSW QDIIAFTVEL RQNFIKWTNG KLTLSAGIGL FADKTPISLM AHQTGELEEA AKGNEKD SI SLFSSDYTFK FDRFITNVYD DKLEQIRYFF NHQDERGKNF IYKLIELLRN YESEEKMNVA RLAYYLTRLE ELTDKDER D KFKQFKKLFF KWYTNNESDR KEAELALLLY VYEIRKD

UniProtKB: CRISPR system single-strand-specific deoxyribonuclease Cas10/Csm1 (subtype III-A)

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Macromolecule #4: crRNA RNA (35-MER)

MacromoleculeName: crRNA RNA (35-MER) / type: rna / ID: 4 / Number of copies: 1
Source (natural)Organism: Streptococcus thermophilus (bacteria)
Molecular weightTheoretical: 11.118652 KDa
SequenceString:
ACGGAAACCU UGAUUCUAAC GCUACUUCUA AAUAA

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Macromolecule #10: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 10 / Number of copies: 8 / 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

Concentration3 mg/mL
BufferpH: 7.5 / Component - Concentration: 250.0 mM / Component - Formula: NaCl / Component - Name: Sodium Chloride / Details: 250mM NaCl, 30mM HEPES, 10mM Beta-mercaptoethanol
GridModel: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec.
VitrificationCryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: GATAN K3 BIOCONTINUUM (6k x 4k) / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: OTHER / Nominal defocus max: 2.0 µm / Nominal defocus min: 1.0 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionType: PHASE FLIPPING ONLY
Startup modelType of model: INSILICO MODEL
Final reconstructionResolution.type: BY AUTHOR / Resolution: 2.67 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION / Number images used: 311708
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: NOT APPLICABLE

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