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PDBsum entry 2flc
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Hydrolase/DNA
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PDB id
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2flc
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References listed in PDB file
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Key reference
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Title
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Dna nicking by hinp1i endonuclease: bending, Base flipping and minor groove expansion.
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Authors
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J.R.Horton,
X.Zhang,
R.Maunus,
Z.Yang,
G.G.Wilson,
R.J.Roberts,
X.Cheng.
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Ref.
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Nucleic Acids Res, 2006,
34,
939-948.
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PubMed id
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Abstract
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HinP1I recognizes and cleaves the palindromic tetranucleotide sequence G
downward arrowCGC in DNA. We report three structures of HinP1I-DNA complexes: in
the presence of Ca(2+) (pre-reactive complex), in the absence of metal ion
(binary complex) and in the presence of Mg(2+) (post-reactive complex). HinP1I
forms a back-to-back dimer with two active sites and two DNA duplexes bound on
the outer surfaces of the dimer facing away from each other. The 10 bp DNA
duplexes undergo protein-induced distortions exhibiting features of A-, B- and
Z-conformations: bending on one side (by intercalation of a phenylalanine side
chain into the major groove), base flipping on the other side of the recognition
site (by expanding the step rise distance of the local base pair to Z-form) and
a local A-form conformation between the two central C:G base pairs of the
recognition site (by binding of the N-terminal helix in the minor groove). In
the pre- and post-reactive complexes, two metals (Ca(2+) or Mg(2+)) are found in
the active site. The enzyme appears to cleave DNA sequentially, hydrolyzing
first one DNA strand, as seen in the post-reactive complex in the crystalline
state, and then the other, as supported by the observation that, in solution, a
nicked DNA intermediate accumulates before linearization.
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Figure 2.
HinP1I-DNA interactions. (A) Summary of the protein-DNA contacts of HinP1I
(green). Backbone mediated interactions are indicated with main chain amine (N) or
carbonyl (O). For simplicity, only single water (w) molecule mediated interactions are
shown. The A(3) base can be in either an intrahelical or extrahelical location (see text).
(B) HinP1I side chains (green) in the major groove of the DNA duplex with a central GCGC
site. The side chain of F91 intercalates DNA immediately outside of the recognition
sequence, between the outer base pair of the recognition site (G:C) and the first base
pair of the flanking DNA (A:T). (C-F) Detailed interactions with the recognition sequence
GCGC. All interactions occur from the major groove side, except for the side chain of F15,
which approaches the DNA from the minor groove.
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Figure 3.
Base flipping outside of the recognition sequence. (A) Superimposition of the
two DNA duplexes, bound with molecule A (colored in grey) or molecule B (colored with
yellow for carbon atoms, blue for nitrogen atoms, red for oxygen atoms and magenta for
phosphate atoms). Base pairs are numbered from 1 to 10, as shown in Figure 2A. The
distances between the stacked bases on strand F are indicated. For comparison, the flipped
Ade, A(3) of strand E, and its corresponding A(3) of strand C are enlarged and boxed. (B)
The flipped Ade, A(3) of strand E, lies against the HinP1I protein surface of molecule B.
The double arrows indicate van der Waals contacts. (C) A view of DNA structure containing
a junction between Z-DNA and B-DNA [PDB 2ACJ ([213]17)]. The two bases of a A:T base pair
at the junction are extruded. Note the van der Waals interaction (double arrow) between
the extruded Thy and an intrahelical base.
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Secondary reference #1
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Title
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Structure of hinp1i endonuclease reveals a striking similarity to the monomeric restriction enzyme mspi.
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Authors
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Z.Yang,
J.R.Horton,
R.Maunus,
G.G.Wilson,
R.J.Roberts,
X.Cheng.
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Ref.
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Nucleic Acids Res, 2005,
33,
1892-1901.
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PubMed id
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Figure 4.
Structural similarity between HinP1I and MspI (A) Schematic diagram of MspI-DNA
interactions, reproduced and modified from Xu et al. (5). Solid lines indicate direct
hydrogen bonds, dotted lines indicate water-mediated hydrogen bonds and `mc' indicates
interaction involving main chain atom. (B) Residues potentially important for DNA base
specific recognition: superimposition of residues of HinP1I (green) and MspI (cyan). (C)
Residues potentially important for catalysis: superimposition of active site residues of
HinP1I (green), MspI (cyan) and EcoRV (grey). Four residues belong to a common motif of
E......PDX[15]DXK (EcoRV), E......SDX[18]QXK (HinP1I) and E......TDX[17]NXK (MspI).
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Figure 7.
A hypothetical model of a HinP1I dimer bound with a single copy of DNA. The two
monomers of Hinp1I are shown in green and magenta. (A) The presentation of green monomer
of Hinp1I is similar to that shown in Figure 2D. (B) A view looking into the DNA
major groove.
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