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PDBsum entry 2fby

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protein metals links
Transferase PDB id
2fby
Jmol
Contents
Protein chain
196 a.a. *
Metals
EU3 ×2
Waters ×128
* Residue conservation analysis
PDB id:
2fby
Name: Transferase
Title: Wrn exonuclease, eu complex
Structure: Werner syndrome helicase. Chain: a. Fragment: exonuclease domain. Engineered: yes
Source: Homo sapiens. Human. Organism_taxid: 9606. Gene: wrn, recq3, recql2. Expressed in: escherichia coli. Expression_system_taxid: 562.
Resolution:
2.00Å     R-factor:   0.221     R-free:   0.245
Authors: J.J.Perry
Key ref:
J.J.Perry et al. (2006). WRN exonuclease structure and molecular mechanism imply an editing role in DNA end processing. Nat Struct Mol Biol, 13, 414-422. PubMed id: 16622405 DOI: 10.1038/nsmb1088
Date:
10-Dec-05     Release date:   25-Apr-06    
PROCHECK
Go to PROCHECK summary
 Headers
 References

Protein chain
Pfam   ArchSchema ?
Q14191  (WRN_HUMAN) -  Werner syndrome ATP-dependent helicase
Seq:
Struc:
 
Seq:
Struc:
 
Seq:
Struc:
1432 a.a.
196 a.a.*
Key:    PfamA domain  PfamB domain  Secondary structure  CATH domain
* PDB and UniProt seqs differ at 2 residue positions (black crosses)

 Gene Ontology (GO) functional annotation 
  GO annot!
  Biological process     nucleobase-containing compound metabolic process   1 term 
  Biochemical function     nucleic acid binding     2 terms  

 

 
DOI no: 10.1038/nsmb1088 Nat Struct Mol Biol 13:414-422 (2006)
PubMed id: 16622405  
 
 
WRN exonuclease structure and molecular mechanism imply an editing role in DNA end processing.
J.J.Perry, S.M.Yannone, L.G.Holden, C.Hitomi, A.Asaithamby, S.Han, P.K.Cooper, D.J.Chen, J.A.Tainer.
 
  ABSTRACT  
 
WRN is unique among the five human RecQ DNA helicases in having a functional exonuclease domain (WRN-exo) and being defective in the premature aging and cancer-related disorder Werner syndrome. Here, we characterize WRN-exo crystal structures, biochemical activity and participation in DNA end joining. Metal-ion complex structures, active site mutations and activity assays reveal a nuclease mechanism mediated by two metal ions. The DNA end-binding Ku70/80 complex specifically stimulates WRN-exo activity, and structure-based mutational inactivation of WRN-exo alters DNA end joining in human cells. We furthermore establish structural and biochemical similarities of WRN-exo to DnaQ-family replicative proofreading exonucleases, describing WRN-specific adaptations consistent with double-stranded DNA specificity and functionally important conformational changes. These results indicate WRN-exo is a human DnaQ family member and support DnaQ-like proofreading activities stimulated by Ku70/80, with implications for WRN functions in age-related pathologies and maintenance of genomic integrity.
 
  Selected figure(s)  
 
Figure 2.
Figure 2. WRN-exo metal-ion dependence and structural analyses. (a) Nuclease activity assays containing WRN-exo (50 pmol) and radiolabeled DNA substrate were incubated for 30 min with either no metal (control; lane 1) or the noted divalent cation(s). WRN-exo 3' arrow 5' dsDNA nuclease activity is supported by Mg^2+ or Mn^2+ ions, but not by Eu^2+ or in the absence of divalent cations. Addition of Eu^2+ inhibits nuclease activity in the presence of equimolar Mg^2+ or Mn^2+ ions. The DNA digestion pattern with equimolar Mg^2+ and Mn^2+ is indistinguishable from that of Mn^2+ alone. (b) Two Mn^2+ ions (purple) are chelated in the WRN active site in the absence of DNA; dashed magenta lines denote metal-ion bonds, with distances labeled. The inner metal ion, M[A], is directly coordinated by Asp82, Glu84 and Asp216, and the outer metal ion, M[B], directly ligates one side chain, Asp82, that bridges the two metal ions. Asp143 has an indirect interaction with the M[B] metal ion via two water molecules. (c) The WRN active site also accommodates two of the larger lanthanide Eu^3+ ions (blue) in the absence of substrate. Dashed blue lines denote metal-ion bonds. (d) Overlay of WRN Mn^2+ and Eu^3+ metal-ion complex structures, colored as in b and c. Incorporation of Eu^3+ metal ions at sites M[A] and M[B] does not cause appreciable changes in the WRN active site.
Figure 6.
Figure 6. WRN-exo hexameric ring model and dGMP-binding site, and altered processing by the W145A mutant. (a) The WRN ring homology model, with differently colored WRN-exo subunits, was built by structural superimposition with the A. thaliana homolog (PDB entry 1VK0). The active site of the exonuclease (with gray spheres denoting metal ions) faces the center of the ring. The central cavity of the WRN ring is large enough (about 30 Å in diameter by 35 Å deep) to accommodate dsDNA and is similar to that observed in Ku70/80 (ref. 49). (b) DNA processing is altered in a WRN-exo W145A mutant. Control reactions with DNA alone or with 10 pmol of Ku70/80 are indicated. WRN-exo and W145A reactions contained 20 fmol of radiolabeled dsDNA substrate, approximately 200 pmol of each WRN nuclease variant and increasing amounts of Ku70/80 (0.06, 0.6 and 6 pmol), denoted by triangles. (c) F[o] - F[c] electron density map of WRN-exo dGMP soak (blue, 3 ; red, 5 ). dGMP stacks against Trp145, consistent with this region interacting with DNA substrate at the center of the ring. (d) Similar internal and external dimensions of the WRN-exo hexamer model (right) and Ku70/80 bound to DNA (left) suggest a possible interaction mode, which would place the protruding 2- 3 loop (left face) adjacent to the Ku dimer and/or allow Ku to provide a suitable DNA orientation.
 
  The above figures are reprinted by permission from Macmillan Publishers Ltd: Nat Struct Mol Biol (2006, 13, 414-422) copyright 2006.  
  Figures were selected by an automated process.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
21220316 M.Aggarwal, J.A.Sommers, R.H.Shoemaker, and R.M.Brosh (2011).
Inhibition of helicase activity by a small molecule impairs Werner syndrome helicase (WRN) function in the cellular response to DNA damage or replication stress.
  Proc Natl Acad Sci U S A, 108, 1525-1530.  
20854710 W.Yang (2011).
Nucleases: diversity of structure, function and mechanism.
  Q Rev Biophys, 44, 1.  
21317904 Y.Y.Hsiao, C.C.Yang, C.L.Lin, J.L.Lin, Y.Duh, and H.S.Yuan (2011).
Structural basis for RNA trimming by RNase T in stable RNA 3'-end maturation.
  Nat Chem Biol, 7, 236-243.
PDB codes: 3ngy 3ngz 3nh0 3nh1 3nh2
20603073 A.Smogorzewska, R.Desetty, T.T.Saito, M.Schlabach, F.P.Lach, M.E.Sowa, A.B.Clark, T.A.Kunkel, J.W.Harper, M.P.Colaiácovo, and S.J.Elledge (2010).
A genetic screen identifies FAN1, a Fanconi anemia-associated nuclease necessary for DNA interstrand crosslink repair.
  Mol Cell, 39, 36-47.  
  20703329 J.E.Deweese, and N.Osheroff (2010).
The use of divalent metal ions by type II topoisomerases.
  Metallomics, 2, 450-459.  
20159459 K.A.Hoadley, and J.L.Keck (2010).
Werner helicase wings DNA binding.
  Structure, 18, 149-151.  
19896421 K.K.Dhillon, J.M.Sidorova, T.M.Albertson, J.B.Anderson, W.C.Ladiges, P.S.Rabinovitch, B.D.Preston, and R.J.Monnat (2010).
Divergent cellular phenotypes of human and mouse cells lacking the Werner syndrome RecQ helicase.
  DNA Repair (Amst), 9, 11-22.  
19949442 A.Vindigni, and I.D.Hickson (2009).
RecQ helicases: multiple structures for multiple functions?
  HFSP J, 3, 153-164.  
18956248 I.Boubriak, P.A.Mason, D.J.Clancy, J.Dockray, R.D.Saunders, and L.S.Cox (2009).
DmWRNexo is a 3'-5' exonuclease: phenotypic and biochemical characterization of mutants of the Drosophila orthologue of human WRN exonuclease.
  Biogerontology, 10, 267-277.  
19283071 P.L.Opresko, G.Sowd, and H.Wang (2009).
The Werner syndrome helicase/exonuclease processes mobile D-loops through branch migration and degradation.
  PLoS ONE, 4, e4825.  
18524993 A.Sallmyr, A.E.Tomkinson, and F.V.Rassool (2008).
Up-regulation of WRN and DNA ligase IIIalpha in chronic myeloid leukemia: consequences for the repair of DNA double-strand breaks.
  Blood, 112, 1413-1423.  
18408731 E.D.Garcin, D.J.Hosfield, S.A.Desai, B.J.Haas, M.Björas, R.P.Cunningham, and J.A.Tainer (2008).
DNA apurinic-apyrimidinic site binding and excision by endonuclease IV.
  Nat Struct Mol Biol, 15, 515-522.
PDB codes: 2nq9 2nqh 2nqj
18653531 J.E.Deweese, A.B.Burgin, and N.Osheroff (2008).
Human topoisomerase IIalpha uses a two-metal-ion mechanism for DNA cleavage.
  Nucleic Acids Res, 36, 4883-4893.  
18346216 R.D.Saunders, I.Boubriak, D.J.Clancy, and L.S.Cox (2008).
Identification and characterization of a Drosophila ortholog of WRN exonuclease that is required to maintain genome integrity.
  Aging Cell, 7, 418-425.  
  18473724 R.Gupta, and R.M.Brosh (2008).
Helicases as prospective targets for anti-cancer therapy.
  Anticancer Agents Med Chem, 8, 390-401.  
18558713 R.Kusumoto, L.Dawut, C.Marchetti, J.Wan Lee, A.Vindigni, D.Ramsden, and V.A.Bohr (2008).
Werner protein cooperates with the XRCC4-DNA ligase IV complex in end-processing.
  Biochemistry, 47, 7548-7556.  
18658245 Z.Bukowy, J.A.Harrigan, D.A.Ramsden, B.Tudek, V.A.Bohr, and T.Stevnsner (2008).
WRN Exonuclease activity is blocked by specific oxidatively induced base lesions positioned in either DNA strand.
  Nucleic Acids Res, 36, 4975-4987.  
18078545 C.D.Putnam, M.Hammel, G.L.Hura, and J.A.Tainer (2007).
X-ray solution scattering (SAXS) combined with crystallography and computation: defining accurate macromolecular structures, conformations and assemblies in solution.
  Q Rev Biophys, 40, 191-285.  
17911100 J.A.Harrigan, J.Piotrowski, L.Di Noto, R.L.Levine, and V.A.Bohr (2007).
Metal-catalyzed oxidation of the Werner syndrome protein causes loss of catalytic activities and impaired protein-protein interactions.
  J Biol Chem, 282, 36403-36411.  
17174478 J.J.Perry, L.Fan, and J.A.Tainer (2007).
Developing master keys to brain pathology, cancer and aging from the structural biology of proteins controlling reactive oxygen species and DNA repair.
  Neuroscience, 145, 1280-1299.  
17148451 K.Kitano, N.Yoshihara, and T.Hakoshima (2007).
Crystal structure of the HRDC domain of human Werner syndrome protein, WRN.
  J Biol Chem, 282, 2717-2728.
PDB codes: 2e1e 2e1f
17660942 L.S.Cox, and R.G.Faragher (2007).
From old organisms to new molecules: integrative biology and therapeutic targets in accelerated human ageing.
  Cell Mol Life Sci, 64, 2620-2641.  
17293595 U.de Silva, S.Choudhury, S.L.Bailey, S.Harvey, F.W.Perrino, and T.Hollis (2007).
The crystal structure of TREX1 explains the 3' nucleotide specificity and reveals a polyproline II helix for protein partnering.
  J Biol Chem, 282, 10537-10543.
PDB codes: 2ioc 2oa8
16856806 L.Wu, and I.D.Hickson (2006).
DNA helicases required for homologous recombination and repair of damaged replication forks.
  Annu Rev Genet, 40, 279-306.  
16935877 M.P.Killoran, and J.L.Keck (2006).
Sit down, relax and unwind: structural insights into RecQ helicase mechanisms.
  Nucleic Acids Res, 34, 4098-4105.  
The most recent references are shown first. Citation data come partly from CiteXplore and partly from an automated harvesting procedure. Note that this is likely to be only a partial list as not all journals are covered by either method. However, we are continually building up the citation data so more and more references will be included with time. Where a reference describes a PDB structure, the PDB codes are shown on the right.