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PDBsum entry 1vsd
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Endoribonuclease
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PDB id
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1vsd
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References listed in PDB file
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Key reference
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Title
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The catalytic domain of avian sarcoma virus integrase: conformation of the active-Site residues in the presence of divalent cations.
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Authors
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G.Bujacz,
M.Jaskólski,
J.Alexandratos,
A.Wlodawer,
G.Merkel,
R.A.Katz,
A.M.Skalka.
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Ref.
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Structure, 1996,
4,
89-96.
[DOI no: ]
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PubMed id
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Abstract
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BACKGROUND: Members of the structurally-related superfamily of enzymes that
includes RNase H, RuvC resolvase, MuA transposase, and retroviral integrase
require divalent cations for enzymatic activity. So far, cation positions are
reported in the X-ray crystal structures of only two of these proteins, E. coli
and human immunodeficiency virus 1 (HIV-1) RNase H. Details of the placement of
metal ions in the active site of retroviral integrases are necessary for the
understanding of the catalytic mechanism of these enzymes. RESULTS: The
structure of the enzymatically active catalytic domain (residues 52-207) of
avian sarcoma virus integrase (ASV IN) has been solved in the presence of
divalent cations (Mn2+ or Mg2+), at 1.7-2.2 A resolution. A single ion of either
type interacts with the carboxylate groups of the active site aspartates and
uses four water molecules to complete its octahedral coordination. The placement
of the aspartate side chains and metal ions is very similar to that observed in
the RNase H members of this superfamily; however, the conformation of the
catalytic aspartates in the active site of ASV IN differs significantly from
that reported for the analogous residues in HIV-1 IN. CONCLUSIONS: Binding of
the required metal ions does not lead to significant structural modifications in
the active site of the catalytic domain of ASV IN. This indicates that at least
one metal-binding site is preformed in the structure, and suggests that the
observed constellation of the acidic residues represents a catalytically
competent active site. Only a single divalent cation was observed even at
extremely high concentrations of the metals. We conclude that either only one
metal ion is needed for catalysis, or that a second metal-binding site can only
exist in the presence of substrate and/or other domains of the protein. The
unexpected differences between the active sites of ASV IN and HIV-1 IN remain
unexplained; they may reflect the effects of crystal contacts on the active site
of HIV-1 IN, or a tendency for structural polymorphism.
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Figure 1.
Figure 1. Chain tracing of the catalytic domain of ASV IN,
showing the secondary structure elements and the location of the
active site. Figure 1. Chain tracing of the catalytic domain
of ASV IN, showing the secondary structure elements and the
location of the active site. (Figure prepared using the program
RIBBONS [[3]33].)
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Figure 2.
Figure 2. Active site of the catalytic domain of ASV IN. (a)
Stereoview of the electron-density map (generated using O [34])
for the Mg^2+ complex (500 mM MgCl[2], see text). This
F[o]–F[c] map, contoured at 5σ level, was calculated at 1.8
å resolution after refinement of a model which excluded
the Mg^2+ cation and its coordinated water molecules. The
density corresponding to the cluster of an
octahedrally-coordinated metal ion and four waters is
exceedingly clear. (b) Stereoview of the active site of ASV IN
generated using MOLSCRIPT [35]. Shown is part of the active site
displaying the coordination of Mn^2+ with four water molecules,
as well as with the carboxylates of Asp121 and Asp64. The water
molecule marked W324 is found in the same location in all ASV
IN structures. The putative hydrogen bonds made by this
molecule (red dashed lines), identified by an analysis of
distances and angles, form a distorted tetrahedron (also
including a bond to Nε2 of Gln153, not marked). Figure 2.
Active site of the catalytic domain of ASV IN. (a) Stereoview of
the electron-density map (generated using O [[4]34]) for the
Mg^2+ complex (500 mM MgCl[2], see text). This F[o]–F[c] map,
contoured at 5σ level, was calculated at 1.8 å resolution
after refinement of a model which excluded the Mg^2+ cation and
its coordinated water molecules. The density corresponding to
the cluster of an octahedrally-coordinated metal ion and four
waters is exceedingly clear. (b) Stereoview of the active site
of ASV IN generated using MOLSCRIPT [[5]35]. Shown is part of
the active site displaying the coordination of Mn^2+ with four
water molecules, as well as with the carboxylates of Asp121 and
Asp64. The water molecule marked W324 is found in the same
location in all ASV IN structures. The putative hydrogen bonds
made by this molecule (red dashed lines), identified by an
analysis of distances and angles, form a distorted tetrahedron
(also including a bond to Nε2 of Gln153, not marked).
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The above figures are
reprinted
by permission from Cell Press:
Structure
(1996,
4,
89-96)
copyright 1996.
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