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Open data
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Basic information
Entry | Database: EMDB / ID: EMD-20447 | |||||||||
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Title | Cryo-EM structure of AdnAB-AMPPNP-DNA complex | |||||||||
![]() | AdnAB-AMPPNP-DNA complex | |||||||||
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Function / homology | ![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() Similarity search - Function | |||||||||
Biological species | ![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() | |||||||||
Method | ![]() ![]() | |||||||||
![]() | Jia N / Unciuleac M / Shuman S / Patel DJ | |||||||||
![]() | ![]() Title: Structures and single-molecule analysis of bacterial motor nuclease AdnAB illuminate the mechanism of DNA double-strand break resection. Authors: Ning Jia / Mihaela C Unciuleac / Chaoyou Xue / Eric C Greene / Dinshaw J Patel / Stewart Shuman / ![]() Abstract: Mycobacterial AdnAB is a heterodimeric helicase-nuclease that initiates homologous recombination by resecting DNA double-strand breaks (DSBs). The AdnA and AdnB subunits are each composed of an N- ...Mycobacterial AdnAB is a heterodimeric helicase-nuclease that initiates homologous recombination by resecting DNA double-strand breaks (DSBs). The AdnA and AdnB subunits are each composed of an N-terminal motor domain and a C-terminal nuclease domain. Here we report cryoelectron microscopy (cryo-EM) structures of AdnAB in three functional states: in the absence of DNA and in complex with forked duplex DNAs before and after cleavage of the 5' single-strand DNA (ssDNA) tail by the AdnA nuclease. The structures reveal the path of the 5' ssDNA through the AdnA nuclease domain and the mechanism of 5' strand cleavage; the path of the 3' tracking strand through the AdnB motor and the DNA contacts that couple ATP hydrolysis to mechanical work; the position of the AdnA iron-sulfur cluster subdomain at the Y junction and its likely role in maintaining the split trajectories of the unwound 5' and 3' strands. Single-molecule DNA curtain analysis of DSB resection reveals that AdnAB is highly processive but prone to spontaneous pausing at random sites on duplex DNA. A striking property of AdnAB is that the velocity of DSB resection slows after the enzyme experiences a spontaneous pause. Our results highlight shared as well as distinctive properties of AdnAB vis-à-vis the RecBCD and AddAB clades of bacterial DSB-resecting motor nucleases. | |||||||||
History |
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Structure visualization
Movie |
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Structure viewer | EM map: ![]() ![]() ![]() |
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 6.5 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 15.6 KB 15.6 KB | Display Display | ![]() |
FSC (resolution estimation) | ![]() | 10 KB | Display | ![]() |
Images | ![]() | 70.8 KB | ||
Filedesc metadata | ![]() | 6.8 KB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 6ppuMC ![]() 6ppjC ![]() 6pprC C: citing same article ( M: atomic model generated by this map |
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Similar structure data |
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Links
EMDB pages | ![]() ![]() |
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Related items in Molecule of the Month |
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Map
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Annotation | AdnAB-AMPPNP-DNA complex | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 0.8613 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
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Sample components
-Entire : AdnAB-AMPPNP-DNA complex
Entire | Name: AdnAB-AMPPNP-DNA complex |
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Components |
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-Supramolecule #1: AdnAB-AMPPNP-DNA complex
Supramolecule | Name: AdnAB-AMPPNP-DNA complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#3 |
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Molecular weight | Theoretical: 200 KDa |
-Supramolecule #2: UvrD/REP helicase
Supramolecule | Name: UvrD/REP helicase / type: complex / ID: 2 / Parent: 1 |
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Source (natural) | Organism: ![]() ![]() |
-Supramolecule #3: ATP-dependent DNA helicase (UvrD/REP)
Supramolecule | Name: ATP-dependent DNA helicase (UvrD/REP) / type: complex / ID: 3 / Parent: 1 |
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Source (natural) | Organism: ![]() ![]() |
-Supramolecule #4: DNA
Supramolecule | Name: DNA / type: complex / ID: 4 / Parent: 1 |
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Source (natural) | Organism: ![]() ![]() ![]() |
-Macromolecule #1: UvrD/REP helicase
Macromolecule | Name: UvrD/REP helicase / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() Strain: ATCC 700084 / mc(2)155 |
Molecular weight | Theoretical: 118.128547 KDa |
Recombinant expression | Organism: ![]() ![]() ![]() |
Sequence | String: MTQVASPVVQ ARYSPVELSA ALGLFPPTDE QAAVIAAPPG PLVVIAGAGA GKTETMAARV VWLVANGFAT PSQVLGLTFT RKAAGQLLR RVRTRLARLA GAGLAPGSGA SDESATVSTY HAFAGTLLRE HGLLLPVEPD TRLLSETELW QLAYDVVCAH P GHLDTEKT ...String: MTQVASPVVQ ARYSPVELSA ALGLFPPTDE QAAVIAAPPG PLVVIAGAGA GKTETMAARV VWLVANGFAT PSQVLGLTFT RKAAGQLLR RVRTRLARLA GAGLAPGSGA SDESATVSTY HAFAGTLLRE HGLLLPVEPD TRLLSETELW QLAYDVVCAH P GHLDTEKT PAAVTAMVLR LSGALAEHLV DTDQLRDTHV ELERLVHTLP AGPYQRDRGP SQWLLRMLAT QTERTELVPL ID ALHQRMR AEKVMDFGMQ MAAAARLAAR FPQVGEQLRQ RFRVVLLDEY QDTGHAQRIA LSSLFGGGAD DGLALTAVGD PIQ SIYGWR GASATNLPRF TTDFPYSDGT PAPTLELRTS WRNPPSTLHV ANAVSEEARR RSVAVRALRP RPDAEPGTIR CALL NNVAA ERDWVADHLA RAYHGAIGRG EAAPTAAVLV RRNADAAPMA EALTARGVPV EVVGVAGLLA VPEVADLVAM LRLIA DPTA GSAVMRILTG PRWRFGARDI AALWRRAVEL DDRPKGELGT ADIVAQAAPD ADTACVADAI CDPGDAERYS PAGYER IVA LGRELTMLRA HLGHPLPELV AEVRRVLGLD AEARAARPVA AGWAGTENLD RFSDLVSDFA GHAGASVSAL LAYLDAA VE VENGLAPAEL TVSHDRVQIL TVHAAKGLEW QVVAVPHLSA RVFPSTTQAR TWLTDASDLP PLLRGDRATE SEIGVPVL D TSDIYDRKIL SDKISDHKKS LDQRRVDEER RLLYVAITRA EDTLLLSGHH WGATESKPRG PSEFLCELKT ILEEATAAG TPCGEIEHWA PDPAPGETNP LRDQVVEALW PPVASADDHV HRGAQLVAAA MAGEVSAEAD QEGWAADVDA LLAERERPPQ QEDTELPGQ LSVSTLVELS RDPKAALTRL RRRLPQRPDP HALLGTTFHE WVQRYFHAER LFDLDDLPGA VDSDSGRAVE E SLAELQDA FVKSPWAART PVEVEVPFDM VLGETVVRGR IDAVFAEPDG TTMVLDWKTG DPPETPEAKE HAAVQLAVYR LA WAAMRGC PPESVRAAFH YVRSGQTVIP ETLPGAEELV KLLAAAPTET AEEADRIT UniProtKB: DNA 3'-5' helicase |
-Macromolecule #2: ATP-dependent DNA helicase (UvrD/REP)
Macromolecule | Name: ATP-dependent DNA helicase (UvrD/REP) / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO / EC number: ![]() |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 76.049312 KDa |
Recombinant expression | Organism: ![]() ![]() ![]() |
Sequence | String: (UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK) (UNK)(UNK)(UNK)(UNK)(UNK)(UNK) (UNK)(UNK)(UNK) (UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK) (UNK)(UNK)(UNK) (UNK)(UNK)(UNK)(UNK)(UNK) ...String: (UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK) (UNK)(UNK)(UNK)(UNK)(UNK)(UNK) (UNK)(UNK)(UNK) (UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK) (UNK)(UNK)(UNK) (UNK)(UNK)(UNK)(UNK)(UNK)(UNK) GTVTVRLAAS THAEGTMIAD ALRRAHLVDG IPWSQMAVIV RSVPRVGTAL ARALTAAGV PVQDNGTDVP VGRQPAAAAL LTVLDVTATG HLDADSAVAL LTGPIGRVDP VTLRQLRRAL RRADGSQPPR D FGDLLVDA IEREPKGLSA EHARTLRRLR AVLTAARRSD ASGADPRYTL WQAWHASGLQ RRWLAASERG GSVGAQADRD LD AVTTLFD VADQYVNRTA GASLRGLVDH VTRLGAAVAR TEPETAAEAV AVLSVHGALA GEWDFVVIAG VQEGLWPNMI PRG GVLGTQ HLVDVLDGVA DMTDRTVSTR APLVAEERRL LMAAMGRART RVMITAVDS(UNK) (UNK)(UNK)(UNK)(UNK) (UNK)(UNK)(UNK)(UNK)(UNK) (UNK)(UNK)(UNK)PPLVAPR VLAPSALVGR LRAVVCAPDG AVDDDARACA AAQ LARLAA AGVPGADPSQ WHAMTSLTTE EPLWSEPGHV VTLSPSTLQM LTDCPLRWLL ERHGGDDGRD VRSTVGSLVH ALVS EPGKT ESQLVNELEK VWDDLPYDAK WYSDNELARH RAMLETFTRW REDTRRQLTE VATEIPVEGI VVEPGENTPG VRVRG RLDR LERDEAGRLV VVDLKTGKSP VTKDDAQNHA QLAMYQLAVA AGLLDDGDEP GGGKLVYLGK AGAAGATERE QDPLTP DKR AEWLETVGEA AAATAGPRFV ARVNNGCANC PVRSSCPAQA NGDRP UniProtKB: ![]() ![]() |
-Macromolecule #3: DNA (29-MER)
Macromolecule | Name: DNA (29-MER) / type: dna / ID: 3 / Number of copies: 1 / Classification: DNA |
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Source (natural) | Organism: ![]() ![]() ![]() |
Molecular weight | Theoretical: 21.477703 KDa |
Sequence | String: (DT)(DT)(DT)(DT)(DT)(DT)(DT)(DC)(DT)(DA) (DA)(DT)(DG)(DC)(DG)(DA)(DG)(DC)(DA)(DC) (DT)(DG)(DC)(DT)(DA)(DT)(DT)(DC)(DC) (DC)(DT)(DA)(DG)(DC)(DA)(DG)(DT)(DG)(DC) (DT) (DC)(DG)(DC)(DA)(DT)(DT) ...String: (DT)(DT)(DT)(DT)(DT)(DT)(DT)(DC)(DT)(DA) (DA)(DT)(DG)(DC)(DG)(DA)(DG)(DC)(DA)(DC) (DT)(DG)(DC)(DT)(DA)(DT)(DT)(DC)(DC) (DC)(DT)(DA)(DG)(DC)(DA)(DG)(DT)(DG)(DC) (DT) (DC)(DG)(DC)(DA)(DT)(DT)(DA)(DG) (DA)(DT)(DT)(DT)(DT)(DG)(DT)(DT)(DT)(DT) (DT)(DT) (DT)(DA)(DG)(DC)(DG)(DG)(DT) (DT)(DT)(DT) |
-Macromolecule #4: MAGNESIUM ION
Macromolecule | Name: MAGNESIUM ION / type: ligand / ID: 4 / Number of copies: 1 / Formula: MG |
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Molecular weight | Theoretical: 24.305 Da |
-Macromolecule #5: IRON/SULFUR CLUSTER
Macromolecule | Name: IRON/SULFUR CLUSTER / type: ligand / ID: 5 / Number of copies: 1 / Formula: SF4 |
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Molecular weight | Theoretical: 351.64 Da |
Chemical component information | ![]() ChemComp-FS1: |
-Experimental details
-Structure determination
Method | ![]() |
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Aggregation state | particle |
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Sample preparation
Concentration | 1.5 mg/mL |
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Buffer | pH: 7.5 / Component - Formula: Tris![]() |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Instrument: FEI VITROBOT MARK IV |
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Electron microscopy
Microscope | FEI TITAN KRIOS |
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Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD![]() |
Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Average electron dose: 2.16 e/Å2 |
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |