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Yorodumi- EMDB-23301: The negative stain EM structure of full-length DNA Ligase III in ... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-23301 | ||||||||||||||||||
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Title | The negative stain EM structure of full-length DNA Ligase III in complex with TDP1 | ||||||||||||||||||
Map data | Full-length DNA LigIII in complex with TDP1 | ||||||||||||||||||
Sample |
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Keywords | DNA repair / protein-protein interactions / DNA BINDING PROTEIN | ||||||||||||||||||
Function / homology | Function and homology information DNA ligase III-XRCC1 complex / negative regulation of mitochondrial DNA replication / 3'-tyrosyl-DNA phosphodiesterase activity / base-excision repair, DNA ligation / DNA ligase activity / DNA ligase (ATP) / DNA ligase (ATP) activity / single strand break repair / Hydrolases; Acting on ester bonds; Phosphoric-diester hydrolases / DNA ligation ...DNA ligase III-XRCC1 complex / negative regulation of mitochondrial DNA replication / 3'-tyrosyl-DNA phosphodiesterase activity / base-excision repair, DNA ligation / DNA ligase activity / DNA ligase (ATP) / DNA ligase (ATP) activity / single strand break repair / Hydrolases; Acting on ester bonds; Phosphoric-diester hydrolases / DNA ligation / HDR through MMEJ (alt-NHEJ) / lagging strand elongation / exonuclease activity / mitochondrial DNA repair / Resolution of AP sites via the single-nucleotide replacement pathway / DNA biosynthetic process / double-strand break repair via alternative nonhomologous end joining / APEX1-Independent Resolution of AP Sites via the Single Nucleotide Replacement Pathway / mitochondrion organization / Gap-filling DNA repair synthesis and ligation in GG-NER / base-excision repair, gap-filling / Nonhomologous End-Joining (NHEJ) / double-strand break repair via homologous recombination / Gap-filling DNA repair synthesis and ligation in TC-NER / double-strand break repair / single-stranded DNA binding / double-stranded DNA binding / cell cycle / cell division / intracellular membrane-bounded organelle / DNA repair / mitochondrion / DNA binding / zinc ion binding / nucleoplasm / ATP binding / nucleus / plasma membrane / cytoplasm Similarity search - Function | ||||||||||||||||||
Biological species | Homo sapiens (human) | ||||||||||||||||||
Method | single particle reconstruction / negative staining / Resolution: 28.0 Å | ||||||||||||||||||
Authors | Sverzhinsky A / Pascal JM | ||||||||||||||||||
Funding support | United States, Canada, 5 items
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Citation | Journal: J Biol Chem / Year: 2021 Title: Direct interaction of DNA repair protein tyrosyl DNA phosphodiesterase 1 and the DNA ligase III catalytic domain is regulated by phosphorylation of its flexible N-terminus. Authors: Ishtiaque Rashid / Michal Hammel / Aleksandr Sverzhinsky / Miaw-Sheue Tsai / John M Pascal / John A Tainer / Alan E Tomkinson / Abstract: Tyrosyl DNA phosphodiesterase 1 (TDP1) and DNA Ligase IIIα (LigIIIα) are key enzymes in single-strand break (SSB) repair. TDP1 removes 3'-tyrosine residues remaining after degradation of DNA ...Tyrosyl DNA phosphodiesterase 1 (TDP1) and DNA Ligase IIIα (LigIIIα) are key enzymes in single-strand break (SSB) repair. TDP1 removes 3'-tyrosine residues remaining after degradation of DNA topoisomerase (TOP) 1 cleavage complexes trapped by either DNA lesions or TOP1 inhibitors. It is not known how TDP1 is linked to subsequent processing and LigIIIα-catalyzed joining of the SSB. Here we define a direct interaction between the TDP1 catalytic domain and the LigIII DNA-binding domain (DBD) regulated by conformational changes in the unstructured TDP1 N-terminal region induced by phosphorylation and/or alterations in amino acid sequence. Full-length and N-terminally truncated TDP1 are more effective at correcting SSB repair defects in TDP1 null cells compared with full-length TDP1 with amino acid substitutions of an N-terminal serine residue phosphorylated in response to DNA damage. TDP1 forms a stable complex with LigIII, as well as full-length LigIIIα alone or in complex with the DNA repair scaffold protein XRCC1. Small-angle X-ray scattering and negative stain electron microscopy combined with mapping of the interacting regions identified a TDP1/LigIIIα compact dimer of heterodimers in which the two LigIII catalytic cores are positioned in the center, whereas the two TDP1 molecules are located at the edges of the core complex flanked by highly flexible regions that can interact with other repair proteins and SSBs. As TDP1and LigIIIα together repair adducts caused by TOP1 cancer chemotherapy inhibitors, the defined interaction architecture and regulation of this enzyme complex provide insights into a key repair pathway in nonmalignant and cancer cells. | ||||||||||||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_23301.map.gz | 1.4 MB | EMDB map data format | |
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Header (meta data) | emd-23301-v30.xml emd-23301.xml | 15.1 KB 15.1 KB | Display Display | EMDB header |
Images | emd_23301.png | 34.4 KB | ||
Filedesc metadata | emd-23301.cif.gz | 5.8 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-23301 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-23301 | HTTPS FTP |
-Validation report
Summary document | emd_23301_validation.pdf.gz | 393.7 KB | Display | EMDB validaton report |
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Full document | emd_23301_full_validation.pdf.gz | 393.2 KB | Display | |
Data in XML | emd_23301_validation.xml.gz | 5 KB | Display | |
Data in CIF | emd_23301_validation.cif.gz | 5.7 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-23301 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-23301 | HTTPS FTP |
-Related structure data
Related structure data | C: citing same article (ref.) |
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Similar structure data |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_23301.map.gz / Format: CCP4 / Size: 2 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Full-length DNA LigIII in complex with TDP1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 3.3 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Sample components
-Entire : Full-length DNA LigIII in complex with TDP1
Entire | Name: Full-length DNA LigIII in complex with TDP1 |
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Components |
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-Supramolecule #1: Full-length DNA LigIII in complex with TDP1
Supramolecule | Name: Full-length DNA LigIII in complex with TDP1 / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all |
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Source (natural) | Organism: Homo sapiens (human) |
Molecular weight | Theoretical: 261 KDa |
-Macromolecule #1: LigIII-alpha nuclear
Macromolecule | Name: LigIII-alpha nuclear / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: Homo sapiens (human) |
Recombinant expression | Organism: Spodoptera frugiperda (fall armyworm) |
Sequence | String: MAEQRFCVDY AKRGTAGCKK CKEKIVKGVC RIGKVVPNPF SESGGDMKEW YHIKCMFEKL ERARATTKK IEDLTELEGW EELEDNEKEQ ITQHIADLSS KAAGTPKKKA VVQAKLTTTG Q VTSPVKGA SFVTSTNPRK FSGFSAKPNN SGEAPSSPTP KRSLSSSKCD ...String: MAEQRFCVDY AKRGTAGCKK CKEKIVKGVC RIGKVVPNPF SESGGDMKEW YHIKCMFEKL ERARATTKK IEDLTELEGW EELEDNEKEQ ITQHIADLSS KAAGTPKKKA VVQAKLTTTG Q VTSPVKGA SFVTSTNPRK FSGFSAKPNN SGEAPSSPTP KRSLSSSKCD PRHKDCLLRE FR KLCAMVA DNPSYNTKTQ IIQDFLRKGS AGDGFHGDVY LTVKLLLPGV IKTVYNLNDK QIV KLFSRI FNCNPDDMAR DLEQGDVSET IRVFFEQSKS FPPAAKSLLT IQEVDEFLLR LSKL TKEDE QQQALQDIAS RCTANDLKCI IRLIKHDLKM NSGAKHVLDA LDPNAYEAFK ASRNL QDVV ERVLHNAQEV EKEPGQRRAL SVQASLMTPV QPMLAEACKS VEYAMKKCPN GMFSEI KYD GERVQVHKNG DHFSYFSRSL KPVLPHKVAH FKDYIPQAFP GGHSMILDSE VLLIDNK TG KPLPFGTLGV HKKAAFQDAN VCLFVFDCIY FNDVSLMDRP LCERRKFLHD NMVEIPNR I MFSEMKRVTK ALDLADMITR VIQEGLEGLV LKDVKGTYEP GKRHWLKVKK DYLNEGAMA DTADLVVLGA FYGQGSKGGM MSIFLMGCYD PGSQKWCTVT KCAGGHDDAT LARLQNELDM VKISKDPSK IPSWLKVNKI YYPDFIVPDP KKAAVWEITG AEFSKSEAHT ADGISIRFPR C TRIRDDKD WKSATNLPQL KELYQLSKEK ADFTVVAGDE GSSTTGGSSE ENKGPSGSAV SR KAPSKPS ASTKKAEGKL SNSNSKDGNM QTAKPSAMKV GEKLATKSSP VKVGEKRKAA DET LCQTKV LLDIFTGVRL YLPPSTPDFS RLRRYFVAFD GDLVQEFDMT SATHVLGSRD KNPA AQQVS PEWIWACIRK RRLVAPC |
-Macromolecule #2: Tyrosyl-DNA phosphodiesterase 1
Macromolecule | Name: Tyrosyl-DNA phosphodiesterase 1 / type: protein_or_peptide / ID: 2 / Enantiomer: LEVO EC number: Hydrolases; Acting on ester bonds; Phosphoric-diester hydrolases |
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Source (natural) | Organism: Homo sapiens (human) |
Recombinant expression | Organism: Spodoptera frugiperda (fall armyworm) |
Sequence | String: MDYKDDDDKE FTMSQEGDYG RWTISSSDES EEEKPKPDKP STSSLLCARQ GAANEPRYTC SEAQKAAHKR KISPVKFSNT DSVLPPKRQK SGSQEDLGWC LSSSDDELQP EMPQKQAEKV VIKKEKDISA PNDGTAQRTE NHGAPACHRL KEEEDEYETS GEGQDIWDML ...String: MDYKDDDDKE FTMSQEGDYG RWTISSSDES EEEKPKPDKP STSSLLCARQ GAANEPRYTC SEAQKAAHKR KISPVKFSNT DSVLPPKRQK SGSQEDLGWC LSSSDDELQP EMPQKQAEKV VIKKEKDISA PNDGTAQRTE NHGAPACHRL KEEEDEYETS GEGQDIWDML DKGNPFQFYL TRVSGVKPKY NSGALHIKDI LSPLFGTLVS SAQFNYCFDV DWLVKQYPPE FRKKPILLVH GDKREAKAHL HAQAKPYENI SLCQAKLDIA FGTHHTKMML LLYEEGLRVV IHTSNLIHAD WHQKTQGIWL SPLYPRIADG THKSGESPTH FKADLISYLM AYNAPSLKEW IDVIHKHDLS ETNVYLIGST PGRFQGSQKD NWGHFRLKKL LKDHASSMPN AESWPVVGQF SSVGSLGADE SKWLCSEFKE SMLTLGKESK TPGKSSVPLY LIYPSVENVR TSLEGYPAGG SLPYSIQTAE KQNWLHSYFH KWSAETSGRS NAMPHIKTYM RPSPDFSKIA WFLVTSANLS KAAWGALEKN GTQLMIRSYE LGVLFLPSAF GLDSFKVKQK FFAGSQEPMA TFPVPYDLPP ELYGSKDRPW IWNIPYVKAP DTHGNMWVPS |
-Experimental details
-Structure determination
Method | negative staining |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 0.0052 mg/mL |
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Buffer | pH: 7.5 |
Staining | Type: NEGATIVE / Material: Uranyl Formate |
Details | Co-purified proteins were crosslinked with glutaraldehyde, then buffer exchanged to remove the crosslinker before negative staining. |
-Electron microscopy
Microscope | FEI TECNAI 12 |
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Image recording | Film or detector model: FEI EAGLE (4k x 4k) / Average exposure time: 1.0 sec. / Average electron dose: 70.0 e/Å2 |
Electron beam | Acceleration voltage: 120 kV / Electron source: LAB6 |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal magnification: 67000 |