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PDBsum entry 2bnf
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References listed in PDB file
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Key reference
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Title
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Structure of escherichia coli ump kinase differs from that of other nucleoside monophosphate kinases and sheds new light on enzyme regulation.
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Authors
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P.Briozzo,
C.Evrin,
P.Meyer,
L.Assairi,
N.Joly,
O.Barzu,
A.M.Gilles.
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Ref.
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J Biol Chem, 2005,
280,
25533-25540.
[DOI no: ]
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PubMed id
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Abstract
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Bacterial UMP kinases are essential enzymes involved in the multistep synthesis
of nucleoside triphosphates. They are hexamers regulated by the allosteric
activator GTP and inhibited by UTP. We solved the crystal structure of
Escherichia coli UMP kinase bound to the UMP substrate (2.3 A resolution), the
UDP product (2.6 A), or UTP (2.45 A). The monomer fold, unrelated to that of
other nucleoside monophosphate kinases, belongs to the carbamate kinase-like
superfamily. However, the phosphate acceptor binding cleft and subunit assembly
are characteristic of UMP kinase. Interactions with UMP explain the high
specificity for this natural substrate. UTP, previously described as an
allosteric inhibitor, was unexpectedly found in the phosphate acceptor site,
suggesting that it acts as a competitive inhibitor. Site-directed mutagenesis of
residues Thr-138 and Asn-140, involved in both uracil recognition and active
site interaction within the hexamer, decreased the activation by GTP and
inhibition by UTP. These experiments suggest a cross-talk mechanism between
enzyme subunits involved in cooperative binding at the phosphate acceptor site
and in allosteric regulation by GTP. As bacterial UMP kinases have no
counterpart in eukaryotes, the information provided here could help the design
of new antibiotics.
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Figure 2.
FIG. 2. Overall fold and quaternary structure. A, ribbon
representation of the UMPKeco monomer fold (using the
UMP-containing complex).  -strands (yellow) and
helices (blue, except  3, pink) are numbered.
A stick model of UMP is shown in magenta. The cross-talk loop
(green) is labeled CT. The loops are smoothed for clarity. A
gray line connects residues that delimit a segment in which no
clear density can be seen. B, the dimer constituted by the two
molecules of an asymmetric unit. View along the
non-crystallographic two-fold axis (indicated by a black ellipse
symbol). The blue subunit orientation is close to that in panel
A. C, ribbon representation of the hexamer viewed along the
three-fold crystallographic axis (indicated by a black
triangle). A particular color is used for each subunit. For the
blue and green dimer, 3 helices are pink.
Corey-Pauling-Koltun space-filling models of UMP are magenta.
The three non-crystallographic two-fold axes are shown by dotted
lines; they are perpendicular to the three-fold axis. D,
magnification of the dimer-dimer interface emphasizing the 2
residues (shown in sticks and labeled) from the cross-talk loop
that interact both with their homologues from the facing dimer
and with UMP (stick model with carbon atoms in magenta).
Hydrogen bonds are shown as red dots. -helices are
transparized for clarity. The figure was drawn with PyMOL,
Version 0.97 (38).
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Figure 3.
FIG. 3. Comparison with NAGK structure. A, superposition of
the ribbon representations of the monomers of UMPKeco (blue,
with a stick model of UMP in cyan) and NAGK (yellow, with
N-acetyl glutamine and ADPNP in red; the helix homologous to
3
is pink). The cross-talk loop of UMPKeco is green, the flexible
loops close to ADPNP are magenta, and the extra -hairpins
of NAGK are orange. The orientation is close to that in Fig. 2A
but slightly modified to better see all ligands. B, the dimer of
NAGK. The orientation is slightly different from that in Fig. 2B
to give a better view of the interface.
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The above figures are
reprinted
by permission from the ASBMB:
J Biol Chem
(2005,
280,
25533-25540)
copyright 2005.
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