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PDBsum entry 2a7x
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References listed in PDB file
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Key reference
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Title
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Crystal structure of the pantothenate synthetase from mycobacterium tuberculosis, Snapshots of the enzyme in action.
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Authors
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S.Wang,
D.Eisenberg.
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Ref.
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Biochemistry, 2006,
45,
1554-1561.
[DOI no: ]
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PubMed id
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Abstract
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Pantothenate synthetase (PS) from Mycobacterium tuberculosis represents a
potential target for antituberculosis drugs. PS catalyzes the ATP-dependent
condensation of pantoate and beta-alanine to form pantothenate. Previously, we
determined the crystal structure of PS from M. tuberculosis and its complexes
with AMPCPP, pantoate, and pantoyl adenylate. Here, we describe the crystal
structure of this enzyme complexed with AMP and its last substrate,
beta-alanine, and show that the phosphate group of AMP serves as an anchor for
the binding of beta-alanine. This structure confirms that binding of
beta-alanine in the active site cavity can occur only after formation of the
pantoyl adenylate intermediate. A new crystal form was also obtained; it
displays the flexible wall of the active site cavity in a conformation incapable
of binding pantoate. Soaking of this crystal form with ATP and pantoate gives a
fully occupied complex of PS with ATP. Crystal structures of these complexes
with substrates, the reaction intermediate, and the reaction product AMP provide
a step-by-step view of the PS-catalyzed reaction. A detailed reaction mechanism
and its implications for inhibitor design are discussed.
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Secondary reference #1
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Title
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Crystal structures of a pantothenate synthetase from m. Tuberculosis and its complexes with substrates and a reaction intermediate.
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Authors
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S.Wang,
D.Eisenberg.
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Ref.
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Protein Sci, 2003,
12,
1097-1108.
[DOI no: ]
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PubMed id
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Figure 1.
Figure 1. Ribbon diagram of the M. tuberculosis
pantothenate synthetase dimer. (A) A side view of the dimer
structure showing that it resembles the shape of a butterfly.
(B) An orthogonal view of (A) from top, with the twofold NCS
symmetry axis (labeled with a dot) approximately perpendicular
to the paper plane. Secondary structure elements for the subunit
A (left) are labeled. Those for subunit B are identical except
that the short helix  3' is not
present. The figure was prepared from the coordinates of the
intermediate complex (data set 5), with the program Molscript
(Kraulis 1991) and Raster3D (Merritt and Murphy 1994). The
molecule in the active site of each subunit, shown in
ball-and-stick, is the reaction intermediate, pantoyl adenylate.
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Figure 4.
Figure 4. Active site cavity and the binding of AMPCPP,
pantoate, and pantoyl adenylate. (A) A stereo view of the active
site cavity of subunit A of the complex with both AMPCPP and
pantoate. The substrates (both with partial occupancy) are shown
as ball-and-stick models. The active site cavity is surrounded
by ß2-loop- 2, ß7-loop,
ß6-loop- 6,
3[10]5'-loop- 5, and
ß3-loop-3[10]3- 3'-loop, and
covered by 3[10]7 and the ß-sheet of C-terminal domain. Residues
around helix 3' (shown in
cyan) are disordered in subunit B, which has a fully occupied
AMPCPP and a glycerol molecule in the active site. (B) A section
of the initial difference electron density map (Fo - Fc) in the
active site of subunit B superimposed on the refined model,
calculated at 1.7 Å and contoured at the 2  level. Side
chains of Lys160, Ser196, and Arg198 have moved relative to
those in the apo enzyme to interact with the phosphate groups,
and thus also have positive initial difference electron density.
The electron density figures are prepared with PYMOL (DeLano
2002). (C) Detailed binding interactions between AMPCPP (shown
with carbon atoms in gold) and protein active site residues of
subunit B. The Mg2+ ion is shown as a yellow sphere, and water
molecules are shown as red spheres. Hydrogen bonds between
AMPCPP and protein atoms, and some water-mediated hydrogen bonds
are shown as dashed lines. A glycerol molecule found next to the
-phosphate of
AMPCPP, at the pantoate binding site, is also shown. (D) A
section of the initial difference electron density (Fo - Fc)
around the bound pantoate molecule in the active site of subunit
A of the pantoate-ß-alanine complex (data set 7) shows that
pantoate is very well ordered with full occupancy. The nearby
residues did not move relative to those of the apo enzyme, and
therefore did not have initial difference density. The electron
density was calculated at 1.7 Å and contoured at 2 . (E) The
pantoate molecule (shown in gold for the carbon atoms) is
tightly bound and fits snugly in its binding site. Two glutamine
side chains form hydrogen bonds to the hydroxyl groups and one
carboxyl oxygen of the pantoate. The two methyl groups and the
hydrophobic side of pantoate interact with the side chains of
Pro38, Met40, and Phe157. (F) A section of the initial
difference electron density (Fo - Fc) around the bound pantoyl
adenylate molecule in the active site of subunit B of the
intermediate complex (data set 6) shows that intermediate is
very well ordered with full occupancy. The electron density was
calculated at 1.7 Å and contoured at 2 . (G) The
pantoyl adenylate molecule (shown with carbon atoms in gold) is
tightly bound and fits snugly in the active site cavity. The
adenosine and pantoyl groups are at identical positions as those
in the AMPCPP complex and pantoate complex, respectively, and
have identical interactions with the active site residues.
However, the -phosphate
moved down to have a covalent bond to the pantoate, which allows
the phosphate group to have a hydrogen bond to the amide
nitrogen of Met40.
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The above figures are
reproduced from the cited reference
with permission from the Protein Society
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