PDBsum entry 3e40

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protein dna_rna metals Protein-protein interface(s) links
Hydrolase/DNA PDB id
Protein chains
247 a.a.
_NA ×3
_CA ×2
Waters ×501
PDB id:
Name: Hydrolase/DNA
Title: Q138f hincii bound to gttaac and cocrystallized with 5 mm ca2+
Structure: Type-2 restriction enzyme hindii. Chain: a, b. Synonym: r.Hindii, type ii restriction enzyme hindii, endonuclease hindii. Engineered: yes. Mutation: yes. 5'- d( Dgp Dcp Dcp Dgp Dgp Dtp Dp Dap Dap Dcp Dcp Dgp Dgp Dc)- 3').
Source: Haemophilus influenzae. Organism_taxid: 727. Gene: hindiir, hi0512. Expressed in: escherichia coli. Expression_system_taxid: 562. Synthetic: yes
2.10Å     R-factor:   0.172     R-free:   0.232
Authors: N.C.Horton,A.C.Babic,E.J.Little,V.M.Manohar
Key ref:
A.C.Babic et al. (2008). DNA distortion and specificity in a sequence-specific endonuclease. J Mol Biol, 383, 186-204. PubMed id: 18762194 DOI: 10.1016/j.jmb.2008.08.032
08-Aug-08     Release date:   26-Aug-08    
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Protein chains
Pfam   ArchSchema ?
P17743  (T2C2_HAEIF) -  Type-2 restriction enzyme HincII
258 a.a.
247 a.a.*
Key:    PfamA domain  Secondary structure  CATH domain
* PDB and UniProt seqs differ at 3 residue positions (black crosses)

 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.2008.08.032 J Mol Biol 383:186-204 (2008)
PubMed id: 18762194  
DNA distortion and specificity in a sequence-specific endonuclease.
A.C.Babic, E.J.Little, V.M.Manohar, J.Bitinaite, N.C.Horton.
Five new structures of the Q138F HincII enzyme bound to a total of three different DNA sequences and three different metal ions (Ca(2+), Mg(2+), and Mn(2+)) are presented. While previous structures were produced from soaking Ca(2+) into preformed Q138F HincII/DNA crystals, the new structures are derived from cocrystallization with Ca(2+), Mg(2+), or Mn(2+). The Mn(2)(+)-bound structure provides the first view of a product complex of Q138F HincII with cleaved DNA. Binding studies and a crystal structure show how Ca(2+) allows trapping of a Q138F HincII complex with noncognate DNA in a catalytically incompetent conformation. Many Q138F HincII/DNA structures show asymmetry, despite the binding of a symmetric substrate by a symmetric enzyme. The various complexes are fit into a model describing the different conformations of the DNA-bound enzyme and show how DNA conformational energetics determine DNA-cleavage rates by the Q138F HincII enzyme.
  Selected figure(s)  
Figure 1.
Fig. 1. (a) Ribbon diagram of HincII dimer (subunits shown in light and dark green) showing bound DNA (pale beige, red, and blue), site of Gln138 (cyan) intercalation, position of Ca^2+ (yellow) in the two active sites, and center step of the recognition sequence (red, blue). (b) The numbering scheme used throughout the paper for the HincII recognition sequence. Y indicates either C or T, R indicates either G or A. (c) Structure of the pyrimidine–purine step as found in the structure of wild-type HincII bound to cognate DNA containing GTCGAC (left) and in B-form DNA (right). The Y7 is unstacked from R8 within the same strand in the HincII bound DNA, and the center step purines (R8 and R8′) have increased stacking across the DNA duplex, forming a cross-strand purine stack (CSPS). (d) Overlay of portions of the structures of wtHincII/CG/Ca^2+ (white) and Q138F/TA/Ca^2+ (cocrystal) (color); nuc, putative water nucleophile of the DNA-cleavage reaction; SP, phosphate at the scissile phosphodiester bond. Arrows emphasize the largest structural differences: the side chain of residue 138 sits differently between the adjacent cytosine bases at the site of intercalation and the main chain around Phe138 is shifted away from the DNA to accommodate this different position (1). As a result, the side chain of Ala137 no longer makes a van der Waals contact to the DNA (2), which may be the reason for the altered pucker of the deoxyribose at Cyt10 from C1′exo to C2′endo (3). The different pucker at Cyt 0 causes a twisting of the sugar–phosphate backbone of the DNA strand (4) propagating to the phosphate between the Pyr7 and Pur8, potentially resulting in the O5′ of Ade9 jutting into the space normally occupied by the side chain of Thr130 (5). To avoid the steric conflict, the phosphate of Ade9 is rotated, blocking the position of the putative nucleophilic water (6).
Figure 7.
Fig. 7. (a) Model of DNA-cleavage mechanism of HincII. M1 and M2 mark the two metal ion binding sites. Dashes indicate direct ligation to a metal ion or hydrogen bond from the 3′P to the nucleophilic water, shown as a pink sphere marked nuc. The scissile bond is colored red. (b) Cartoon of the connection between the CSPS and the distance between the phosphorus of the phosphate at the scissile phosphodiester bond (SP) and the phosphate 3′ from the SP (3′P). Dashes indicate direct ligation to the metal ion, M1, or hydrogen bond from the 3′P to the nucleophile (water or hydroxide molecule) shown as a pink sphere. Left: inactive conformation with the 3′P blocking the nucleophile binding site on the M1 ion. Right: active conformation where disruption of the CSPS separates SP from 3′P on each strand to allow the nucleophile to bind to the M1 ion. The scissile bond is shown in red.
  The above figures are reprinted from an Open Access publication published by Elsevier: J Mol Biol (2008, 383, 186-204) copyright 2008.  
  Figures were selected by an automated process.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
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
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