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PDBsum entry 2gie

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protein dna_rna metals Protein-protein interface(s) links
Hydrolase/DNA PDB id
2gie
Jmol
Contents
Protein chains
257 a.a.
DNA/RNA
Metals
_NA
Waters ×508
PDB id:
2gie
Name: Hydrolase/DNA
Title: Hincii bound to cognate DNA gttaac
Structure: 5'-d( Gp Cp Cp Gp Gp Tp Tp Ap Ap Cp Cp Gp G)-3'. Chain: e, f, g, h. Engineered: yes. Type ii restriction enzyme hincii. Chain: a, b, c, d. Synonym: endonuclease hincii, r.Hincii. Engineered: yes
Source: Synthetic: yes. Haemophilus influenzae. Organism_taxid: 727. Gene: hinciir. Expressed in: escherichia coli. Expression_system_taxid: 562.
Biol. unit: Tetramer (from PQS)
Resolution:
2.60Å     R-factor:   0.238     R-free:   0.306
Authors: N.C.Horton,H.K.Joshi,C.Etzkorn,L.Chatwell,J.Bitinaite
Key ref:
H.K.Joshi et al. (2006). Alteration of sequence specificity of the type II restriction endonuclease HincII through an indirect readout mechanism. J Biol Chem, 281, 23852-23869. PubMed id: 16675462 DOI: 10.1074/jbc.M512339200
Date:
28-Mar-06     Release date:   18-Jul-06    
PROCHECK
Go to PROCHECK summary
 Headers
 References

Protein chains
Pfam   ArchSchema ?
P17743  (T2C2_HAEIF) -  Type-2 restriction enzyme HincII
Seq:
Struc:
258 a.a.
257 a.a.*
Key:    PfamA domain  Secondary structure  CATH domain
* PDB and UniProt seqs differ at 2 residue positions (black crosses)

 Enzyme reactions 
   Enzyme class: 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: 10.1074/jbc.M512339200 J Biol Chem 281:23852-23869 (2006)
PubMed id: 16675462  
 
 
Alteration of sequence specificity of the type II restriction endonuclease HincII through an indirect readout mechanism.
H.K.Joshi, C.Etzkorn, L.Chatwell, J.Bitinaite, N.C.Horton.
 
  ABSTRACT  
 
The functional and structural consequences of a mutation of the DNA intercalating residue of HincII, Q138F, are presented. Modeling has suggested that the DNA intercalation by Gln-138 results in DNA distortions potentially used by HincII in indirect readout of its cognate DNA, GTYRAC (Y = C or T, R = A or G) (Horton, N. C., Dorner, L. F., and Perona, J. J. (2002) Nat. Struct. Biol. 9, 42-47). Kinetic data presented here indicate that the mutation of glutamine 138 to phenylalanine (Q138F) results in a change in sequence specificity at the center two base pairs of the cognate recognition site. We show that the preference of HincII for cutting, but not binding, the three cognate sites differing in the center two base pairs has been altered by the mutation Q138F. Five new crystal structures are presented including Q138F HincII bound to GTTAAC and GTCGAC both with and without Ca2+ as well as the structure of wild type HincII bound to GTTAAC. The Q138F HincII/DNA structures show conformational changes in the protein, bound DNA, and at the protein-DNA interface, consistent with the formation of adaptive complexes. Analysis of these structures and the effect of Ca2+ binding on the protein-DNA interface illuminates the origin of the altered specificity by the mutation Q138F in the HincII enzyme.
 
  Selected figure(s)  
 
Figure 6.
FIGURE 6. Stereo diagrams of superpositions using base atoms of purine 8 of wild type HincII/CG/Ca^2+ (black) showing the central two base pairs of the recognition sequence GTYRAC (underlined) of both strands. A, Q138F/TA (red). B, Q138F/TA/Ca^2+ (pink). C, Q138F/CG (blue). D, Q138F/CG/Ca^2+ (cyan). E, Q138F/CG/Ca^2+ (cyan), Q138F/TA/Ca^2+ (pink), and wild type HincII/CG/Ca^2+ (gray) showing the propagation of differences in stacking with the center step purines to the phosphate position of Ade 9. A residue of the active site, Asp-127, as well as the adjacent residue implicated in potential steric conflicts with the DNA, Thr-130, are also shown. Py, a pyrimidine; Pur, a purine.
Figure 9.
FIGURE 9. Stereo diagrams of active site superpositions, wild type HincII/CG/Ca^2+ (gray) and Q138F/DNA structures (colored). A, subunit A of Q138F/TA (red) and Q138F/CG (blue). B, subunit B of Q138F/TA (red) and Q138F/CG (blue). C, subunit A of Q138F/TA/Ca^2+ (pink) and Q138F/CG/Ca^2+ (cyan). D, subunit B of Q138F/TA/Ca^2+ (pink) and Q138F/CG/Ca^2+ (cyan).
 
  The above figures are reprinted by permission from the ASBMB: J Biol Chem (2006, 281, 23852-23869) copyright 2006.  
  Figures were selected by an automated process.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
22961381 J.H.Chang, X.Jiao, K.Chiba, C.Oh, C.E.Martin, M.Kiledjian, and L.Tong (2012).
Dxo1 is a new type of eukaryotic enzyme with both decapping and 5'-3' exoribonuclease activity.
  Nat Struct Mol Biol, 19, 1011-1017.
PDB codes: 4gps 4gpu
20861000 M.Firczuk, M.Wojciechowski, H.Czapinska, and M.Bochtler (2011).
DNA intercalation without flipping in the specific ThaI-DNA complex.
  Nucleic Acids Res, 39, 744-754.
PDB code: 3ndh
20668693 S.Guan, A.Blanchard, P.Zhang, and Z.Zhu (2010).
Alteration of sequence specificity of the type IIS restriction endonuclease BtsI.
  PLoS One, 5, e11787.  
20375162 S.K.Menon, B.J.Eilers, M.J.Young, and C.M.Lawrence (2010).
The crystal structure of D212 from sulfolobus spindle-shaped virus ragged hills reveals a new member of the PD-(D/E)XK nuclease superfamily.
  J Virol, 84, 5890-5897.
PDB code: 2w8m
19419959 S.R.Coffin, and N.O.Reich (2009).
Escherichia coli DNA adenine methyltransferase: the structural basis of processive catalysis and indirect read-out.
  J Biol Chem, 284, 18390-18400.  
18762194 A.C.Babic, E.J.Little, V.M.Manohar, J.Bitinaite, and N.C.Horton (2008).
DNA distortion and specificity in a sequence-specific endonuclease.
  J Mol Biol, 383, 186-204.
PDB codes: 3e3y 3e40 3e41 3e42 3e43 3e44 3e45
19081059 E.J.Little, A.C.Babic, and N.C.Horton (2008).
Early interrogation and recognition of DNA sequence by indirect readout.
  Structure, 16, 1828-1837.
PDB code: 3ebc
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.  
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.