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PDBsum entry 3bvq
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References listed in PDB file
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Key reference
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Title
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Structures of the rare-Cutting restriction endonuclease noti reveal a unique metal binding fold involved in DNA binding.
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Authors
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A.R.Lambert,
D.Sussman,
B.Shen,
R.Maunus,
J.Nix,
J.Samuelson,
S.Y.Xu,
B.L.Stoddard.
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Ref.
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Structure, 2008,
16,
558-569.
[DOI no: ]
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PubMed id
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Abstract
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The structure of the rare-cutting restriction endonuclease NotI, which
recognizes the 8 bp target 5'-GCGGCCGC-3', has been solved with and without
bound DNA. Because of its specificity (recognizing a site that occurs once per
65 kb), NotI is used to generate large genomic fragments and to map DNA
methylation status. NotI contains a unique metal binding fold, found in a
variety of putative endonucleases, occupied by an iron atom coordinated within a
tetrahedral Cys4 motif. This domain positions nearby protein elements for DNA
recognition, and serves a structural role. While recognition of the central six
base pairs of the target is accomplished via a saturated hydrogen bond network
typical of restriction enzymes, the most peripheral base pairs are engaged in a
single direct contact in the major groove, reflecting reduced pressure to
recognize those positions. NotI may represent an evolutionary intermediate
between mobile endonucleases (which recognize longer target sites) and canonical
restriction endonucleases.
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Figure 2.
Figure 2. Homodimeric NotI REase
Domains are colored as
described in Figure 1. Select secondary structural elements are
numbered for reference. The NotI dimer interface is formed by
the domain-swapped α helices (cyan) and the DNA recognition
helices (red).
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Figure 6.
Figure 6. The NotI Active Site
(A) Active site residues
Glu145, Asp160, Glu182, and Gln184 are colored yellow. The bound
DNA is colored white, with the scissile phosphate designated
orange. The nucleophilic water molecule is shown as a blue
sphere and two calcium ions are depicted as magenta spheres.
Dashed lines represent interactions between atoms in the active
site and numbers indicate distances in angstroms (as average
values calculated between the two active sites of the NotI
homodimer).
(B) Superposition of active site residues of
the P(D)…(D/E)xK active site motif from NotI (green) and BglII
(orange). Both REases have a glutamine in the general base
position typically occupied by a lysine residue.
(C)
Superposition of the two-metal ion active sites of NotI (green)
and BamHI (purple). Asp160/Asp94, Glu182/Glu111, and
Gln184/Glu113 are residues belonging to the canonical
PD…(D/E)xK nuclease motif, while the third acidic residue,
Glu145/Glu77, helps coordinate a second metal ion in the active
site. The position of this third acidic residue is conserved
between the NotI and BamHI enzymes.
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The above figures are
reprinted
from an Open Access publication published by Cell Press:
Structure
(2008,
16,
558-569)
copyright 2008.
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