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PDBsum entry 1kqp
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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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Nh3-Dependent NAD+ synthetase from bacillus subtilis at 1 a resolution.
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Authors
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J.Symersky,
Y.Devedjiev,
K.Moore,
C.Brouillette,
L.Delucas.
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Ref.
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Acta Crystallogr D Biol Crystallogr, 2002,
58,
1138-1146.
[DOI no: ]
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PubMed id
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Abstract
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The final step of NAD+ biosynthesis includes an amide transfer to nicotinic acid
adenine dinucleotide (NaAD) catalyzed by NAD+ synthetase. This enzyme was
co-crystallized in microgravity with natural substrates NaAD and ATP at pH 8.5.
The crystal was exposed to ammonium ions, synchrotron diffraction data were
collected and the atomic model was refined anisotropically at 1 A resolution to
R = 11.63%. Both binding sites are occupied by the NAD-adenylate intermediate,
pyrophosphate and two magnesium ions. The atomic resolution of the structure
allows better definition of non-planar peptide groups, reveals a low mean
anisotropy of protein and substrate atoms and indicates the H-atom positions of
the phosphoester group of the reaction intermediate. The phosphoester group is
protonated at the carbonyl O atom O7N, suggesting a carbenium-ion structure
stabilized by interactions with two solvent sites presumably occupied by ammonia
and a water molecule. A mechanism is proposed for the second catalytic step,
which includes a nucleophilic attack by the ammonia molecule on the intermediate.
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Figure 5.
Figure 5 Spatial relationship between the intermediate
protonated at O7N and cation-binding sites M+(I) and M+(II).
Distances are shown in Å for subunit A. C atoms are in green, N
atoms in blue, O atoms in red, P atoms in magenta and H atoms in
cyan. Hydrophobic residues are all in gray, magnesium ions in
black and the monovalent cation-binding sites M+(I) and M+(II)
are shown as golden spheres. Rendered using RIBBONS (Carson,
1997[Carson, M. (1997). Methods Enzymol. 277, 493-505.]).
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Figure 6.
Figure 6 (a) Stabilization of the protonated intermediate by the
hydrogen bond with ammonia positioned in the cation-binding site
M+(I) and a hypothetical hydrogen bond with water molecule in
the cation-binding site M+(II) releasing the ammonia molecule
for nucleophilic attack. (b) Reaction scheme for the second
catalytic step of NAD^+ synthetase. The NAD-adenylate
intermediate reacts with ammonia to form NAD^+ and AMP and two
protons are released. R[1] represents the ADP-nicotinosyl moiety
and R[2] represents the AMP moiety.
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The above figures are
reprinted
by permission from the IUCr:
Acta Crystallogr D Biol Crystallogr
(2002,
58,
1138-1146)
copyright 2002.
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Secondary reference #1
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Title
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Stabilization of active-Site loops in nh3-Dependent NAD+ synthetase from bacillus subtilis.
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Authors
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Y.Devedjiev,
J.Symersky,
R.Singh,
M.Jedrzejas,
C.Brouillette,
W.Brouillette,
D.Muccio,
D.Chattopadhyay,
L.Delucas.
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Ref.
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Acta Crystallogr D Biol Crystallogr, 2001,
57,
806-812.
[DOI no: ]
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PubMed id
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Figure 1.
Figure 1 A scheme of the reaction catalyzed by NAD^+ synthetase.
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Figure 6.
Figure 6 Coordination of Mg2+ in the ATP-binding site of NAD^+
synthetase. The nicotinosyl moiety of NaAD is shown in violet,
AMP in gold and PP[i] in red; Mg2+ with coordinated O atoms are
shown in silver, relevant amino-acid residues are in green and
the loop 204-225 is indicated by thin brown lines. Positions
Mg(I), Mg(II) and Mg(III) are explained in the text.
Coordination of the new Mg(III) position is indicated by thin
silver lines. The new conformation of Glu162 at pH 7.5 is shown
in cyan. ATP and AMP-CPP are not shown for clarity. Prepared
with RIBBONS (Carson, 1997[Carson, M. (1997). Methods Enzymol.
277, 493-505.]).
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The above figures are
reproduced from the cited reference
with permission from the IUCr
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Secondary reference #2
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Title
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A novel deamido-Nad+-Binding site revealed by the trapped NAD-Adenylate intermediate in the NAD+ synthetase structure.
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Authors
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M.Rizzi,
M.Bolognesi,
A.Coda.
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Ref.
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Structure, 1998,
6,
1129-1140.
[DOI no: ]
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PubMed id
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Figure 1.
Figure 1. A scheme of the two-step reaction catalyzed by
NAD^+ synthetase.
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The above figure is
reproduced from the cited reference
with permission from Cell Press
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Secondary reference #3
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Title
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Crystal structure of nh3-Dependent NAD+ synthetase from bacillus subtilis.
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Authors
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M.Rizzi,
C.Nessi,
A.Mattevi,
A.Coda,
M.Bolognesi,
A.Galizzi.
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Ref.
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Embo J, 1996,
15,
5125-5134.
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PubMed id
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