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PDBsum entry 1vj7
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Hydrolase, transferase
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PDB id
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1vj7
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Contents |
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* Residue conservation analysis
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References listed in PDB file
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Key reference
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Title
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Conformational antagonism between opposing active sites in a bifunctional rela/spot homolog modulates (p)ppgpp metabolism during the stringent response [corrected].
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Authors
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T.Hogg,
U.Mechold,
H.Malke,
M.Cashel,
R.Hilgenfeld.
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Ref.
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Cell, 2004,
117,
57-68.
[DOI no: ]
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PubMed id
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Abstract
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Enzymes of the Rel/Spo family enable bacteria to survive prolonged periods of
nutrient limitation by producing an intracellular signaling alarmone, (p)ppGpp,
which triggers the so-called stringent response. Both the synthesis of (p)ppGpp
from ATP and GDP(GTP), and its hydrolysis to GDP(GTP) and pyrophosphate, are
catalyzed by Rel/Spo proteins. The 2.1 A crystal structure of the bifunctional
catalytic fragment of the Rel/Spo homolog from Streptococcus dysgalactiae subsp.
equisimilis, Rel(Seq), reveals two conformations of the enzyme corresponding to
known reciprocal activity states: (p)ppGpp-hydrolase-OFF/(p)ppGpp-synthetase-ON
and hydrolase-ON/synthetase-OFF. The hydrolase and synthetase domains bear
remarkable similarities to the catalytic domains of the cyclic phosphodiesterase
and nucleotidyltransferase superfamilies, respectively. The active sites,
separated by more than 30 A, contain bound nucleotides including an unusual
(p)ppGpp derivative, GDP-2':3'-cyclic monophosphate. Reciprocal regulation of
the antagonistic catalytic activities, suggested by the structure, is supported
by mutagenesis experiments and appears to involve ligand-induced signal
transmission between the two active sites.
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Figure 2.
Figure 2. Similarities between the Catalytic Domains of
Rel[Seq], Human Phosphodiesterase (PDE) and Human DNA Polymerase
Beta (pol β)Structural and topological diagrams highlighting
equivalent folds and active-site residues for: (A) catalytic
domain (residues 152−528) of PDE4; (B) Rel[Seq]1–385; (C)
catalytic domain (residues 10−335) of pol β. Homologous
structural elements are displayed as ribbons; nonequivalent
regions as thin gray lines. Monomer 2 of Rel[Seq]1–385 is
shown, with ppG2′:3′p, and GDP. Dark blue sphere,
catalytic metal ion (Zn^2+ for PDE4; Mn^2+ for Rel[Seq]).
Conserved residues of the H−X[(n)]−HD−X[(n)]−D metal
binding tetrad are labeled in the accompanying topology diagrams
(A and B). Two of the three catalytic carboxylates in pol β
(Asp190 and Asp256, C) are also found in Rel[Seq] (Asp264 and
Glu323, B). Rel/Spo enzymes lack a counterpart for the second
carboxylate of the D-X-D motif in NTases (Asp192 in pol β).
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Figure 4.
Figure 4. Conformations of the Synthetase Site in
Rel[Seq]1–385 and a Superposition with pol β(A) The
synthetase-ON conformation (monomer 1). Coloring is according to
Figure 1; individual structural elements are labeled in red. The
nucleophilic O3′ of GDP is marked. The final 2mFo-DFc
electron density map, shown for GDP (blue mesh), is contoured at
1.0 σ. Selected H-bonds are shown as gray dashed lines. The
catalytic loop (α13/β4) harboring Asp264 is stabilized in a
3[10]-helical conformation through multiple van der Waals
interactions (represented by black double-arrow dashed lines)
with loop α11/α12 and the first two turns of α12 (labeled t1,
t2). Chain traces extending from loops α11/α12, β3/α13, and
α13/β4 are not visible due to image slab restrictions.(B) The
synthetase-OFF conformation (monomer 2). The α11/α12 loop and
the first two turns of α12 are disordered; the resulting
elimination of van der Waals contacts to the catalytic loop
(α13/β4) leads to (1), partial refolding of the latter into an
N-terminal extension of β4, and (2), disorder of residues
254–261 including the remaining residues of the catalytic loop
and the C terminus of α13.(C) Representative electron density
in the synthetase site of monomer 1. GDP is highlighted in
orange. The final 2mFo-DFc electron density map (1.0 σ) is
overlaid as blue mesh.(D) Stereographic superposition between
the synthetase site of Rel[Seq]1–385 (monomer 1) and the
active site of pol β in the (pol β)·(gapped
DNA)·(ddCTP) complex. The latter complex is rendered in
gray shading with the exception of the primer 3′-terminal
nucleotide (orange), ddCTP (cyan), and the two Mg^2+ ions (dark
blue). Rel[Seq]1–385 and its GDP ligand are colored according
to (A). The putative catalytic carboxylates of Rel[Seq], Asp264
and Glu323, are N terminally frameshifted by two residues
relative to their pol β counterparts (indicated by black
arrows).
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The above figures are
reprinted
by permission from Cell Press:
Cell
(2004,
117,
57-68)
copyright 2004.
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