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PDBsum entry 1na6

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protein Protein-protein interface(s) links
Hydrolase PDB id
1na6
Jmol
Contents
Protein chains
365 a.a. *
395 a.a. *
Waters ×252
* Residue conservation analysis
PDB id:
1na6
Name: Hydrolase
Title: Crystal structure of restriction endonuclease ecorii mutant r88a
Structure: Restriction endonuclease ecorii. Chain: a, b. Synonym: ecorii. Type ii restriction enzyme ecorii. Endonuclease ecorii. R.Ecorii. Engineered: yes. Mutation: yes
Source: Escherichia coli. Organism_taxid: 562. Gene: ecorii. Expressed in: escherichia coli. Expression_system_taxid: 562.
Biol. unit: Dimer (from PQS)
Resolution:
2.10Å     R-factor:   0.230     R-free:   0.284
Authors: X.E.Zhou,Y.Wang,M.Reuter,M.Mucke,D.H.Kruger,E.J.Meehan, L.Chen
Key ref:
X.E.Zhou et al. (2004). Crystal structure of type IIE restriction endonuclease EcoRII reveals an autoinhibition mechanism by a novel effector-binding fold. J Mol Biol, 335, 307-319. PubMed id: 14659759 DOI: 10.1016/j.jmb.2003.10.030
Date:
26-Nov-02     Release date:   16-Dec-03    
PROCHECK
Go to PROCHECK summary
 Headers
 References

Protein chain
Pfam   ArchSchema ?
P14633  (T2E2_ECOLX) -  Type-2 restriction enzyme EcoRII
Seq:
Struc:
404 a.a.
365 a.a.*
Protein chain
Pfam   ArchSchema ?
P14633  (T2E2_ECOLX) -  Type-2 restriction enzyme EcoRII
Seq:
Struc:
404 a.a.
395 a.a.*
Key:    PfamA domain  Secondary structure  CATH domain
* PDB and UniProt seqs differ at 24 residue positions (black crosses)

 Enzyme reactions 
   Enzyme class: Chains A, B: E.C.3.1.21.4  - Type Ii site-specific deoxyribonuclease.
[IntEnz]   [ExPASy]   [KEGG]   [BRENDA]
      Reaction: Endonucleolytic cleavage of DNA to give specific double-stranded fragments with terminal 5'-phosphates.
      Cofactor: Mg(2+)
 Gene Ontology (GO) functional annotation 
  GO annot!
  Biological process     nucleic acid phosphodiester bond hydrolysis   3 terms 
  Biochemical function     hydrolase activity     5 terms  

 

 
DOI no: 10.1016/j.jmb.2003.10.030 J Mol Biol 335:307-319 (2004)
PubMed id: 14659759  
 
 
Crystal structure of type IIE restriction endonuclease EcoRII reveals an autoinhibition mechanism by a novel effector-binding fold.
X.E.Zhou, Y.Wang, M.Reuter, M.Mücke, D.H.Krüger, E.J.Meehan, L.Chen.
 
  ABSTRACT  
 
EcoRII is a type IIE restriction endonuclease that interacts with two copies of the DNA recognition sequence 5'CCWGG, one being the actual target of cleavage, the other serving as the allosteric effector. The mode of enzyme activation by effector binding is unknown. To investigate the molecular basis of activation and cleavage mechanisms by EcoRII, the crystal structure of EcoRII mutant R88A has been solved at 2.1A resolution. The EcoRII monomer has two domains linked through a hinge loop. The N-terminal effector-binding domain has a novel DNA recognition fold with a prominent cleft. The C-terminal catalytic domain has a restriction endonuclease-like fold. Structure-based sequence alignment identified the putative catalytic site of EcoRII that is spatially blocked by the N-terminal domain. The structure together with the earlier characterized EcoRII enzyme activity enhancement in the absence of its N-terminal domain reveal an autoinhibition/activation mechanism of enzyme activity mediated by a novel effector-binding fold. This is the first case of autoinhibition, a mechanism described for many transcription factors and signal transducing proteins, of a restriction endonuclease.
 
  Selected figure(s)  
 
Figure 6.
Figure 6. The N-domain structure and the effector-binding cleft. The secondary structural elements are labeled and so are some of the key amino acid residues in the proposed effector-binding cleft. Putative DNA-binding residues H36, Y41, K92, R94, E96, K97 and R98 are shown in ball-and-stick.
Figure 9.
Figure 9. The inactive dimer structure of EcoRII and a model of autoinhibition. The ribbon diagram is a back view of Figure 5 and related to Figure 5 by a 180° rotation about the vertical axis. The locations of the two effector clefts and the two catalytic half-sites are indicated. The catalytic half-sites are blocked by the N-domains and inaccessible by a substrate DNA. The two key residues Y41 and E96 in the effector clefts are shown in ball-and-stick and labeled. The a-helix H6 is also labeled.
 
  The above figures are reprinted by permission from Elsevier: J Mol Biol (2004, 335, 307-319) copyright 2004.  
  Figures were selected by the author.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
20961958 M.Laganeckas, M.Margelevicius, and C.Venclovas (2011).
Identification of new homologs of PD-(D/E)XK nucleases by support vector machines trained on data derived from profile-profile alignments.
  Nucleic Acids Res, 39, 1187-1196.  
19729506 D.Golovenko, E.Manakova, G.Tamulaitiene, S.Grazulis, and V.Siksnys (2009).
Structural mechanisms for the 5'-CCWGG sequence recognition by the N- and C-terminal domains of EcoRII.
  Nucleic Acids Res, 37, 6613-6624.
PDB codes: 3hqf 3hqg
19788335 J.L.Gilmore, Y.Suzuki, G.Tamulaitis, V.Siksnys, K.Takeyasu, and Y.L.Lyubchenko (2009).
Single-molecule dynamics of the DNA-EcoRII protein complexes revealed with high-speed atomic force microscopy.
  Biochemistry, 48, 10492-10498.  
19400796 M.Szczepek, P.Mackeldanz, E.Möncke-Buchner, J.Alves, D.H.Krüger, and M.Reuter (2009).
Molecular analysis of restriction endonuclease EcoRII from Escherichia coli reveals precise regulation of its enzymatic activity by autoinhibition.
  Mol Microbiol, 72, 1011-1021.  
18400177 A.R.Lambert, D.Sussman, B.Shen, R.Maunus, J.Nix, J.Samuelson, S.Y.Xu, and B.L.Stoddard (2008).
Structures of the rare-cutting restriction endonuclease NotI reveal a unique metal binding fold involved in DNA binding.
  Structure, 16, 558-569.
PDB codes: 3bvq 3c25
18820295 G.Tamulaitis, M.Zaremba, R.H.Szczepanowski, M.Bochtler, and V.Siksnys (2008).
How PspGI, catalytic domain of EcoRII and Ecl18kI acquire specificities for different DNA targets.
  Nucleic Acids Res, 36, 6101-6108.  
18456708 J.Orlowski, and J.M.Bujnicki (2008).
Structural and evolutionary classification of Type II restriction enzymes based on theoretical and experimental analyses.
  Nucleic Acids Res, 36, 3552-3569.  
  18687114 M.Wayengera, H.Kajumbula, and W.Byarugaba (2008).
Identification of restriction endonuclease with potential ability to cleave the HSV-2 genome: inherent potential for biosynthetic versus live recombinant microbicides.
  Theor Biol Med Model, 5, 18.  
18829716 R.H.Szczepanowski, M.A.Carpenter, H.Czapinska, M.Zaremba, G.Tamulaitis, V.Siksnys, A.S.Bhagwat, and M.Bochtler (2008).
Central base pair flipping and discrimination by PspGI.
  Nucleic Acids Res, 36, 6109-6117.  
18367450 S.Takahashi, H.Matsuno, H.Furusawa, and Y.Okahata (2008).
Direct monitoring of allosteric recognition of type IIE restriction endonuclease EcoRII.
  J Biol Chem, 283, 15023-15030.  
17617640 G.Tamulaitis, M.Zaremba, R.H.Szczepanowski, M.Bochtler, and V.Siksnys (2007).
Nucleotide flipping by restriction enzymes analyzed by 2-aminopurine steady-state fluorescence.
  Nucleic Acids Res, 35, 4792-4799.  
17584917 L.Knizewski, L.N.Kinch, N.V.Grishin, L.Rychlewski, and K.Ginalski (2007).
Realm of PD-(D/E)XK nuclease superfamily revisited: detection of novel families with modified transitive meta profile searches.
  BMC Struct Biol, 7, 40.  
17344322 M.Kaus-Drobek, H.Czapinska, M.SokoĊ‚owska, G.Tamulaitis, R.H.Szczepanowski, C.Urbanke, V.Siksnys, and M.Bochtler (2007).
Restriction endonuclease MvaI is a monomer that recognizes its target sequence asymmetrically.
  Nucleic Acids Res, 35, 2035-2046.
PDB codes: 2oa9 2oaa
16723432 G.J.Gemmen, R.Millin, and D.E.Smith (2006).
DNA looping by two-site restriction endonucleases: heterogeneous probability distributions for loop size and unbinding force.
  Nucleic Acids Res, 34, 2864-2877.  
16962970 G.Tamulaitiene, A.Jakubauskas, C.Urbanke, R.Huber, S.Grazulis, and V.Siksnys (2006).
The crystal structure of the rare-cutting restriction enzyme SdaI reveals unexpected domain architecture.
  Structure, 14, 1389-1400.
PDB code: 2ixs
16628220 M.Bochtler, R.H.Szczepanowski, G.Tamulaitis, S.Grazulis, H.Czapinska, E.Manakova, and V.Siksnys (2006).
Nucleotide flips determine the specificity of the Ecl18kI restriction endonuclease.
  EMBO J, 25, 2219-2229.
PDB codes: 2fqz 2gb7
16893959 M.Carpenter, P.Divvela, V.Pingoud, J.Bujnicki, and A.S.Bhagwat (2006).
Sequence-dependent enhancement of hydrolytic deamination of cytosines in DNA by the restriction enzyme PspGI.
  Nucleic Acids Res, 34, 3762-3770.  
15667656 A.A.Chmiel, J.M.Bujnicki, and K.J.Skowronek (2005).
A homology model of restriction endonuclease SfiI in complex with DNA.
  BMC Struct Biol, 5, 2.  
15972856 L.N.Kinch, K.Ginalski, L.Rychlewski, and N.V.Grishin (2005).
Identification of novel restriction endonuclease-like fold families among hypothetical proteins.
  Nucleic Acids Res, 33, 3598-3605.  
16195548 Q.S.Xu, R.J.Roberts, and H.C.Guo (2005).
Two crystal forms of the restriction enzyme MspI-DNA complex show the same novel structure.
  Protein Sci, 14, 2590-2600.
PDB code: 1yfi
16247004 S.Grazulis, E.Manakova, M.Roessle, M.Bochtler, G.Tamulaitiene, R.Huber, and V.Siksnys (2005).
Structure of the metal-independent restriction enzyme BfiI reveals fusion of a specific DNA-binding domain with a nonspecific nuclease.
  Proc Natl Acad Sci U S A, 102, 15797-15802.
PDB code: 2c1l
15563460 V.Pingoud, A.Sudina, H.Geyer, J.M.Bujnicki, R.Lurz, G.Lüder, R.Morgan, E.Kubareva, and A.Pingoud (2005).
Specificity changes in the evolution of type II restriction endonucleases: a biochemical and bioinformatic analysis of restriction enzymes that recognize unrelated sequences.
  J Biol Chem, 280, 4289-4298.  
15383647 H.Geyer, R.Geyer, and V.Pingoud (2004).
A novel strategy for the identification of protein-DNA contacts by photocrosslinking and mass spectrometry.
  Nucleic Acids Res, 32, e132.  
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.