+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-15022 | |||||||||
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Title | CryoEM structure of Ku heterodimer bound to DNA, PAXX and XLF | |||||||||
Map data | ||||||||||
Sample |
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Keywords | NHEJ / Ku70 / Ku80 / DNA damage / DNA BINDING PROTEIN / XLF | |||||||||
Function / homology | Function and homology information positive regulation of ligase activity / DNA ligase IV complex / DNA ligation involved in DNA repair / Ku70:Ku80 complex / negative regulation of t-circle formation / DNA end binding / small-subunit processome assembly / positive regulation of lymphocyte differentiation / DNA-dependent protein kinase complex / DNA-dependent protein kinase-DNA ligase 4 complex ...positive regulation of ligase activity / DNA ligase IV complex / DNA ligation involved in DNA repair / Ku70:Ku80 complex / negative regulation of t-circle formation / DNA end binding / small-subunit processome assembly / positive regulation of lymphocyte differentiation / DNA-dependent protein kinase complex / DNA-dependent protein kinase-DNA ligase 4 complex / cellular response to X-ray / immunoglobulin V(D)J recombination / nonhomologous end joining complex / DNA ligation / regulation of smooth muscle cell proliferation / nuclear telomere cap complex / Cytosolic sensors of pathogen-associated DNA / double-strand break repair via classical nonhomologous end joining / IRF3-mediated induction of type I IFN / recombinational repair / regulation of telomere maintenance / U3 snoRNA binding / positive regulation of neurogenesis / cellular response to fatty acid / hematopoietic stem cell proliferation / protein localization to chromosome, telomeric region / cellular hyperosmotic salinity response / response to ionizing radiation / telomeric DNA binding / positive regulation of catalytic activity / 2-LTR circle formation / site of DNA damage / Lyases; Carbon-oxygen lyases; Other carbon-oxygen lyases / T cell differentiation / 5'-deoxyribose-5-phosphate lyase activity / hematopoietic stem cell differentiation / positive regulation of protein kinase activity / ATP-dependent activity, acting on DNA / DNA polymerase binding / DNA helicase activity / : / enzyme activator activity / positive regulation of telomere maintenance via telomerase / activation of innate immune response / telomere maintenance / cyclin binding / B cell differentiation / neurogenesis / cellular response to leukemia inhibitory factor / central nervous system development / protein-DNA complex / small-subunit processome / Nonhomologous End-Joining (NHEJ) / Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement / cellular response to gamma radiation / fibrillar center / double-strand break repair via nonhomologous end joining / double-strand break repair / site of double-strand break / scaffold protein binding / double-stranded DNA binding / secretory granule lumen / DNA recombination / ficolin-1-rich granule lumen / transcription regulator complex / chromosome, telomeric region / damaged DNA binding / molecular adaptor activity / transcription cis-regulatory region binding / ribonucleoprotein complex / response to xenobiotic stimulus / innate immune response / negative regulation of DNA-templated transcription / ubiquitin protein ligase binding / DNA damage response / Neutrophil degranulation / protein-containing complex binding / nucleolus / positive regulation of DNA-templated transcription / protein homodimerization activity / ATP hydrolysis activity / positive regulation of transcription by RNA polymerase II / protein-containing complex / DNA binding / RNA binding / extracellular region / nucleoplasm / ATP binding / identical protein binding / membrane / nucleus / plasma membrane / cytoplasm / cytosol Similarity search - Function | |||||||||
Biological species | Homo sapiens (human) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 2.68 Å | |||||||||
Authors | Hardwick SW / Kefala-Stavridi A / Chirgadze DY / Blundell TL / Chaplin AK | |||||||||
Funding support | United Kingdom, 1 items
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Citation | Journal: Sci Adv / Year: 2023 Title: PAXX binding to the NHEJ machinery explains functional redundancy with XLF. Authors: Murielle Seif-El-Dahan / Antonia Kefala-Stavridi / Philippe Frit / Steven W Hardwick / Dima Y Chirgadze / Taiana Maia De Oliviera / Jessica Andreani / Sébastien Britton / Nadia Barboule / ...Authors: Murielle Seif-El-Dahan / Antonia Kefala-Stavridi / Philippe Frit / Steven W Hardwick / Dima Y Chirgadze / Taiana Maia De Oliviera / Jessica Andreani / Sébastien Britton / Nadia Barboule / Madeleine Bossaert / Arun Prasad Pandurangan / Katheryn Meek / Tom L Blundell / Virginie Ropars / Patrick Calsou / Jean-Baptiste Charbonnier / Amanda K Chaplin / Abstract: Nonhomologous end joining is a critical mechanism that repairs DNA double-strand breaks in human cells. In this work, we address the structural and functional role of the accessory protein PAXX ...Nonhomologous end joining is a critical mechanism that repairs DNA double-strand breaks in human cells. In this work, we address the structural and functional role of the accessory protein PAXX [paralog of x-ray repair cross-complementing protein 4 (XRCC4) and XRCC4-like factor (XLF)] in this mechanism. Here, we report high-resolution cryo-electron microscopy (cryo-EM) and x-ray crystallography structures of the PAXX C-terminal Ku-binding motif bound to Ku70/80 and cryo-EM structures of PAXX bound to two alternate DNA-dependent protein kinase (DNA-PK) end-bridging dimers, mediated by either Ku80 or XLF. We identify residues critical for the Ku70/PAXX interaction in vitro and in cells. We demonstrate that PAXX and XLF can bind simultaneously to the Ku heterodimer and act as structural bridges in alternate forms of DNA-PK dimers. Last, we show that engagement of both proteins provides a complementary advantage for DNA end synapsis and end joining in cells. | |||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_15022.map.gz | 226.9 MB | EMDB map data format | |
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Header (meta data) | emd-15022-v30.xml emd-15022.xml | 22.5 KB 22.5 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_15022_fsc.xml | 13.2 KB | Display | FSC data file |
Images | emd_15022.png | 60.1 KB | ||
Filedesc metadata | emd-15022.cif.gz | 7.4 KB | ||
Others | emd_15022_half_map_1.map.gz emd_15022_half_map_2.map.gz | 226.5 MB 226.5 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-15022 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-15022 | HTTPS FTP |
-Validation report
Summary document | emd_15022_validation.pdf.gz | 1.1 MB | Display | EMDB validaton report |
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Full document | emd_15022_full_validation.pdf.gz | 1.1 MB | Display | |
Data in XML | emd_15022_validation.xml.gz | 21 KB | Display | |
Data in CIF | emd_15022_validation.cif.gz | 26.8 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-15022 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-15022 | HTTPS FTP |
-Related structure data
Related structure data | 7zygMC 7zwaC 8ascC 8bh3C 8bhvC 8bhyC M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_15022.map.gz / Format: CCP4 / Size: 244.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||
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Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 0.65 Å | ||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Half map: #1
File | emd_15022_half_map_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: #2
File | emd_15022_half_map_2.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Sample components
-Entire : Ku70/80 hetero dimer bound to DNA, PAXX and XLF
Entire | Name: Ku70/80 hetero dimer bound to DNA, PAXX and XLF |
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Components |
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-Supramolecule #1: Ku70/80 hetero dimer bound to DNA, PAXX and XLF
Supramolecule | Name: Ku70/80 hetero dimer bound to DNA, PAXX and XLF / type: organelle_or_cellular_component / ID: 1 / Parent: 0 / Macromolecule list: #1-#4, #6 |
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Source (natural) | Organism: Homo sapiens (human) |
Molecular weight | Theoretical: 160 KDa |
-Macromolecule #1: X-ray repair cross-complementing protein 6
Macromolecule | Name: X-ray repair cross-complementing protein 6 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO EC number: Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement |
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Source (natural) | Organism: Homo sapiens (human) |
Molecular weight | Theoretical: 69.945039 KDa |
Recombinant expression | Organism: Spodoptera frugiperda (fall armyworm) |
Sequence | String: MSGWESYYKT EGDEEAEEEQ EENLEASGDY KYSGRDSLIF LVDASKAMFE SQSEDELTPF DMSIQCIQSV YISKIISSDR DLLAVVFYG TEKDKNSVNF KNIYVLQELD NPGAKRILEL DQFKGQQGQK RFQDMMGHGS DYSLSEVLWV CANLFSDVQF K MSHKRIML ...String: MSGWESYYKT EGDEEAEEEQ EENLEASGDY KYSGRDSLIF LVDASKAMFE SQSEDELTPF DMSIQCIQSV YISKIISSDR DLLAVVFYG TEKDKNSVNF KNIYVLQELD NPGAKRILEL DQFKGQQGQK RFQDMMGHGS DYSLSEVLWV CANLFSDVQF K MSHKRIML FTNEDNPHGN DSAKASRART KAGDLRDTGI FLDLMHLKKP GGFDISLFYR DIISIAEDED LRVHFEESSK LE DLLRKVR AKETRKRALS RLKLKLNKDI VISVGIYNLV QKALKPPPIK LYRETNEPVK TKTRTFNTST GGLLLPSDTK RSQ IYGSRQ IILEKEETEE LKRFDDPGLM LMGFKPLVLL KKHHYLRPSL FVYPEESLVI GSSTLFSALL IKCLEKEVAA LCRY TPRRN IPPYFVALVP QEEELDDQKI QVTPPGFQLV FLPFADDKRK MPFTEKIMAT PEQVGKMKAI VEKLRFTYRS DSFEN PVLQ QHFRNLEALA LDLMEPEQAV DLTLPKVEAM NKRLGSLVDE FKELVYPPDY NPEGKVTKRK HDNEGSGSKR PKVEYS EEE LKTHISKGTL GKFTVPMLKE ACRAYGLKSG LKKQELLEAL TKHFQD UniProtKB: X-ray repair cross-complementing protein 6 |
-Macromolecule #2: X-ray repair cross-complementing protein 5
Macromolecule | Name: X-ray repair cross-complementing protein 5 / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO EC number: Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement |
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Source (natural) | Organism: Homo sapiens (human) |
Molecular weight | Theoretical: 82.812438 KDa |
Recombinant expression | Organism: Spodoptera frugiperda (fall armyworm) |
Sequence | String: MVRSGNKAAV VLCMDVGFTM SNSIPGIESP FEQAKKVITM FVQRQVFAEN KDEIALVLFG TDGTDNPLSG GDQYQNITVH RHLMLPDFD LLEDIESKIQ PGSQQADFLD ALIVSMDVIQ HETIGKKFEK RHIEIFTDLS SRFSKSQLDI IIHSLKKCDI S LQFFLPFS ...String: MVRSGNKAAV VLCMDVGFTM SNSIPGIESP FEQAKKVITM FVQRQVFAEN KDEIALVLFG TDGTDNPLSG GDQYQNITVH RHLMLPDFD LLEDIESKIQ PGSQQADFLD ALIVSMDVIQ HETIGKKFEK RHIEIFTDLS SRFSKSQLDI IIHSLKKCDI S LQFFLPFS LGKEDGSGDR GDGPFRLGGH GPSFPLKGIT EQQKEGLEIV KMVMISLEGE DGLDEIYSFS ESLRKLCVFK KI ERHSIHW PCRLTIGSNL SIRIAAYKSI LQERVKKTWT VVDAKTLKKE DIQKETVYCL NDDDETEVLK EDIIQGFRYG SDI VPFSKV DEEQMKYKSE GKCFSVLGFC KSSQVQRRFF MGNQVLKVFA ARDDEAAAVA LSSLIHALDD LDMVAIVRYA YDKR ANPQV GVAFPHIKHN YECLVYVQLP FMEDLRQYMF SSLKNSKKYA PTEAQLNAVD ALIDSMSLAK KDEKTDTLED LFPTT KIPN PRFQRLFQCL LHRALHPREP LPPIQQHIWN MLNPPAEVTT KSQIPLSKIK TLFPLIEAKK KDQVTAQEIF QDNHED GPT AKKLKTEQGG AHFSVSSLAE GSVTSVGSVN PAENFRVLVK QKKASFEEAS NQLINHIEQF LDTNETPYFM KSIDCIR AF REEAIKFSEE QRFNNFLKAL QEKVEIKQLN HFWEIVVQDG ITLITKEEAS GSSVTAEEAK KFLAPKDKPS GDTAAVFE E GGDVDDLLDM I UniProtKB: X-ray repair cross-complementing protein 5 |
-Macromolecule #3: Protein PAXX
Macromolecule | Name: Protein PAXX / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: Homo sapiens (human) |
Molecular weight | Theoretical: 21.663498 KDa |
Recombinant expression | Organism: Escherichia coli BL21(DE3) (bacteria) |
Sequence | String: MDPLSPPLCT LPPGPEPPRF VCYCEGEESG EGDRGGFNLY VTDAAELWST CFTPDSLAAL KARFGLSAAE DITPRFRAAC EQQAVALTL QEDRASLTLS GGPSALAFDL SKVPGPEAAP RLRALTLGLA KRVWSLERRL AAAEETAVSP RKSPRPAGPQ L FLPDPDPQ ...String: MDPLSPPLCT LPPGPEPPRF VCYCEGEESG EGDRGGFNLY VTDAAELWST CFTPDSLAAL KARFGLSAAE DITPRFRAAC EQQAVALTL QEDRASLTLS GGPSALAFDL SKVPGPEAAP RLRALTLGLA KRVWSLERRL AAAEETAVSP RKSPRPAGPQ L FLPDPDPQ RGGPGPGVRR RCPGESLINP GFKSKKPAGG VDFDET UniProtKB: Protein PAXX |
-Macromolecule #4: Non-homologous end-joining factor 1
Macromolecule | Name: Non-homologous end-joining factor 1 / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: Homo sapiens (human) |
Molecular weight | Theoretical: 33.372234 KDa |
Recombinant expression | Organism: Escherichia coli BL21(DE3) (bacteria) |
Sequence | String: MEELEQGLLM QPWAWLQLAE NSLLAKVFIT KQGYALLVSD LQQVWHEQVD TSVVSQRAKE LNKRLTAPPA AFLCHLDNLL RPLLKDAAH PSEATFSCDC VADALILRVR SELSGLPFYW NFHCMLASPS LVSQHLIRPL MGMSLALQCQ VRELATLLHM K DLEIQDYQ ...String: MEELEQGLLM QPWAWLQLAE NSLLAKVFIT KQGYALLVSD LQQVWHEQVD TSVVSQRAKE LNKRLTAPPA AFLCHLDNLL RPLLKDAAH PSEATFSCDC VADALILRVR SELSGLPFYW NFHCMLASPS LVSQHLIRPL MGMSLALQCQ VRELATLLHM K DLEIQDYQ ESGATLIRDR LKTEPFEENS FLEQFMIEKL PEACSIGDGK PFVMNLQDLY MAVTTQEVQV GQKHQGAGDP HT SNSASLQ GIDSQCVNQP EQLVSSAPTL SAPEKESTGT SGPLQRPQLS KVKRKKPRGL FS UniProtKB: Non-homologous end-joining factor 1 |
-Macromolecule #5: DNA
Macromolecule | Name: DNA / type: dna / ID: 5 / Number of copies: 1 / Classification: DNA |
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Source (natural) | Organism: Homo sapiens (human) |
Molecular weight | Theoretical: 4.503948 KDa |
Sequence | String: (DA)(DT)(DC)(DC)(DC)(DT)(DC)(DT)(DA)(DG) (DA)(DT)(DA)(DT)(DC) |
-Macromolecule #6: DNA
Macromolecule | Name: DNA / type: dna / ID: 6 / Number of copies: 1 / Classification: DNA |
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Source (natural) | Organism: Homo sapiens (human) |
Molecular weight | Theoretical: 4.673059 KDa |
Sequence | String: (DG)(DA)(DT)(DA)(DT)(DC)(DT)(DA)(DG)(DA) (DG)(DG)(DG)(DA)(DT) |
-Macromolecule #7: PHOSPHATE ION
Macromolecule | Name: PHOSPHATE ION / type: ligand / ID: 7 / Number of copies: 1 / Formula: PO4 |
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Molecular weight | Theoretical: 94.971 Da |
Chemical component information | ChemComp-PO4: |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 2 mg/mL |
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Buffer | pH: 8 |
Vitrification | Cryogen name: ETHANE |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Number grids imaged: 1 / Number real images: 13024 / Average exposure time: 1.37 sec. / Average electron dose: 52.1 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.2 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 130000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |