PDBsum entry 1v13

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Hydrolase PDB id
Jmol PyMol
Protein chains
120 a.a. *
126 a.a. *
_ZN ×2
Waters ×93
* Residue conservation analysis
PDB id:
Name: Hydrolase
Title: Crystal structure of the mutant his103ala of the colicin e9 dnase domain in complex with zn+2 (2.0 angstroms)
Structure: Colicin e9. Chain: a, b. Engineered: yes. Mutation: yes
Source: Escherichia coli. Organism_taxid: 562. Expressed in: escherichia coli. Expression_system_taxid: 469008.
Biol. unit: Dimer (from PDB file)
2.00Å     R-factor:   0.234     R-free:   0.307
Authors: M.J.Mate,C.Kleanthous
Key ref:
M.J.Maté and C.Kleanthous (2004). Structure-based analysis of the metal-dependent mechanism of H-N-H endonucleases. J Biol Chem, 279, 34763-34769. PubMed id: 15190054 DOI: 10.1074/jbc.M403719200
06-Apr-04     Release date:   23-Jun-04    
Go to PROCHECK summary

Protein chain
Pfam   ArchSchema ?
P09883  (CEA9_ECOLX) -  Colicin-E9
582 a.a.
120 a.a.*
Protein chain
Pfam   ArchSchema ?
P09883  (CEA9_ECOLX) -  Colicin-E9
582 a.a.
126 a.a.*
Key:    PfamA domain  Secondary structure  CATH domain
* PDB and UniProt seqs differ at 2 residue positions (black crosses)

 Enzyme reactions 
   Enzyme class: Chains A, B: E.C.3.1.-.-
[IntEnz]   [ExPASy]   [KEGG]   [BRENDA]
 Gene Ontology (GO) functional annotation 
  GO annot!
  Biological process     cytolysis   3 terms 
  Biochemical function     receptor binding     2 terms  


DOI no: 10.1074/jbc.M403719200 J Biol Chem 279:34763-34769 (2004)
PubMed id: 15190054  
Structure-based analysis of the metal-dependent mechanism of H-N-H endonucleases.
M.J.Maté, C.Kleanthous.
Controversy surrounds the metal-dependent mechanism of H-N-H endonucleases, enzymes involved in a variety of biological functions, including intron homing and DNA repair. To address this issue we determined the crystal structures for complexes of the H-N-H motif containing bacterial toxin colicin E9 with Zn(2+), Zn(2+).DNA, and Mg(2+).DNA. The structures show that the rigid V-shaped architecture of the active site does not undergo any major conformational changes on binding to the minor groove of DNA and that the same interactions are made to the nucleic acid regardless of which metal ion is bound to the enzyme. The scissile phosphate contacts the single metal ion of the motif through distortion of the DNA brought about by the insertion of the Arg-96-Glu-100 salt bridge into the minor groove and a network of contacts to the DNA phosphate backbone that straddle the metal site. The Mg(2+)-bound structure reveals an unusual coordination scheme involving two H-N-H histidine residues, His-102 and His-127. The mechanism of DNA cleavage is likely related to that of other single metal ion-dependent endonucleases, such as I-PpoI and Vvn, although in these enzymes the single alkaline earth metal ion is coordinated by oxygen-bearing amino acids. The structures also provide a rationale as to why H-N-H endonucleases are inactive in the presence of Zn(2+) but active with other transition metal ions such as Ni(2+). This is because of coordination of the Zn(2+) ion through a third histidine, His-131. "Active" transition metal ions are those that bind more weakly to the H-N-H motif because of the disengagement of His-131, which we suggest allows a water molecule to complete the catalytic cycle.
  Selected figure(s)  
Figure 3.
FIG. 3. A hydrogen-bonding network connects DNA distortion to approach of the scissile bond to the H-N-H motif metal center. Stereo representation of the main hydrogen-bonding interactions between the E9 DNase and the DNA minor groove in the Zn2+ complex, with identical interactions observed in the Mg2+-bound complex. The DNA and protein residues are cream and green, respectively, the Zn2+ ion is magenta, and hydrogen bonds are plotted as dotted lines. The scissile bond of the DNA can only approach the metal ion if the DNA is distorted. This is accomplished by the insertion of the Arg-96-Glu-100 salt bridge into the minor groove and stabilization of this configuration through a number of hydrogen-bonding interactions to the DNA phosphates through residues Arg-5, Asp-51, and Arg-54.
Figure 4.
FIG. 4. Simulated annealing omit maps around the active site for the complexes of H103A E9 DNase·dsDNA bound to Zn2+ (A) and Mg2+ (B). The metal ion, the scissile phosphate, and histidines 102, 127, and 131 were omitted for the calculation, and the resulting maps were plotted at a contour level of 2.0 . The coordination of both metal ions is indicated with dotted lines. The figure highlights how the H-N-H motif is an adaptable metal binding center. It is able to accommodate both the tetrahedral coordination chemistry of a Zn2+ ion (A) and then, through subtle reorientations of the H-N-H histidine residues and the inclusion of DNA phosphodiester oxygen atoms and a water molecule, the octahedral geometry required for Mg2+ ion binding (B). Only five of the possible six coordination sites are discernible for the Mg2+ ion at the current level of resolution.
  The above figures are reprinted by permission from the ASBMB: J Biol Chem (2004, 279, 34763-34769) copyright 2004.  
  Figures were selected by the author.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
20846957 M.Midon, P.Schäfer, A.Pingoud, M.Ghosh, A.F.Moon, M.J.Cuneo, R.E.London, and G.Meiss (2011).
Mutational and biochemical analysis of the DNA-entry nuclease EndA from Streptococcus pneumoniae.
  Nucleic Acids Res, 39, 623-634.  
20854710 W.Yang (2011).
Nucleases: diversity of structure, function and mechanism.
  Q Rev Biophys, 44, 1.  
20140205 S.H.Chan, L.Opitz, L.Higgins, D.O'loane, and S.Y.Xu (2010).
Cofactor requirement of HpyAV restriction endonuclease.
  PLoS One, 5, e9071.  
19053714 C.Chen, K.Krause, and B.M.Pettitt (2009).
Advantage of being a dimer for Serratia marcescens endonuclease.
  J Phys Chem B, 113, 511-521.  
19749752 K.B.Levin, O.Dym, S.Albeck, S.Magdassi, A.H.Keeble, C.Kleanthous, and D.S.Tawfik (2009).
Following evolutionary paths to protein-protein interactions with high affinity and selectivity.
  Nat Struct Mol Biol, 16, 1049-1055.
PDB code: 3gjn
19651876 L.E.Corina, W.Qiu, A.Desai, and D.L.Herrin (2009).
Biochemical and mutagenic analysis of I-CreII reveals distinct but important roles for both the H-N-H and GIY-YIG motifs.
  Nucleic Acids Res, 37, 5810-5821.  
19380375 M.Sokolowska, H.Czapinska, and M.Bochtler (2009).
Crystal structure of the beta beta alpha-Me type II restriction endonuclease Hpy99I with target DNA.
  Nucleic Acids Res, 37, 3799-3810.
PDB codes: 3fc3 3gox
19272175 S.L.Wu, C.C.Li, J.C.Chen, Y.J.Chen, C.T.Lin, T.Y.Ho, and C.Y.Hsiang (2009).
Mutagenesis identifies the critical amino acid residues of human endonuclease G involved in catalysis, magnesium coordination, and substrate specificity.
  J Biomed Sci, 16, 6.  
18261473 C.M.Dupureur (2008).
Roles of metal ions in nucleases.
  Curr Opin Chem Biol, 12, 250-255.  
18283539 J.Hanus, M.Kalinowska-Herok, and P.Widlak (2008).
The major apoptotic endonuclease DFF40/CAD is a deoxyribose-specific and double-strand-specific enzyme.
  Apoptosis, 13, 377-382.  
18953336 W.Yang (2008).
An equivalent metal ion in one- and two-metal-ion catalysis.
  Nat Struct Mol Biol, 15, 1228-1231.  
17499273 A.Jakubauskas, J.Giedriene, J.M.Bujnicki, and A.Janulaitis (2007).
Identification of a single HNH active site in type IIS restriction endonuclease Eco31I.
  J Mol Biol, 370, 157-169.  
17347522 E.Cascales, S.K.Buchanan, D.Duché, C.Kleanthous, R.Lloubès, K.Postle, M.Riley, S.Slatin, and D.Cavard (2007).
Colicin biology.
  Microbiol Mol Biol Rev, 71, 158-229.  
17138564 M.Ghosh, G.Meiss, A.M.Pingoud, R.E.London, and L.C.Pedersen (2007).
The nuclease a-inhibitor complex is characterized by a novel metal ion bridge.
  J Biol Chem, 282, 5682-5690.
PDB code: 2o3b
17430971 M.Saravanan, K.Vasu, R.Kanakaraj, D.N.Rao, and V.Nagaraja (2007).
R.KpnI, an HNH superfamily REase, exhibits differential discrimination at non-canonical sequences in the presence of Ca2+ and Mg2+.
  Nucleic Acids Res, 35, 2777-2786.  
17029241 I.A.Cymerman, A.Obarska, K.J.Skowronek, A.Lubys, and J.M.Bujnicki (2006).
Identification of a new subfamily of HNH nucleases and experimental characterization of a representative member, HphI restriction endonuclease.
  Proteins, 65, 867-876.  
16434744 L.G.Doudeva, H.Huang, K.C.Hsia, Z.Shi, C.L.Li, Y.Shen, Y.S.Cheng, and H.S.Yuan (2006).
Crystal structural analysis and metal-dependent stability and activity studies of the ColE7 endonuclease domain in complex with DNA/Zn2+ or inhibitor/Ni2+.
  Protein Sci, 15, 269-280.
PDB codes: 1zns 1znv
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 code is shown on the right.