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PDBsum entry 2fpx
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References listed in PDB file
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Key reference
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Title
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Structural snapshots of escherichia coli histidinol phosphate phosphatase along the reaction pathway.
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Authors
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E.S.Rangarajan,
A.Proteau,
J.Wagner,
M.N.Hung,
A.Matte,
M.Cygler.
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Ref.
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J Biol Chem, 2006,
281,
37930-37941.
[DOI no: ]
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PubMed id
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Abstract
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HisB from Escherichia coli is a bifunctional enzyme catalyzing the sixth and
eighth steps of l-histidine biosynthesis. The N-terminal domain (HisB-N)
possesses histidinol phosphate phosphatase activity, and its crystal structure
shows a single domain with fold similarity to the haloacid dehalogenase (HAD)
enzyme family. HisB-N forms dimers in the crystal and in solution. The structure
shows the presence of a structural Zn(2+) ion stabilizing the conformation of an
extended loop. Two metal binding sites were also identified in the active site.
Their presence was further confirmed by isothermal titration calorimetry. HisB-N
is active in the presence of Mg(2+), Mn(2+), Co(2+), or Zn(2+), but Ca(2+) has
an inhibitory effect. We have determined structures of several intermediate
states corresponding to snapshots along the reaction pathway, including that of
the phosphoaspartate intermediate. A catalytic mechanism, different from that
described for other HAD enzymes, is proposed requiring the presence of the
second metal ion not found in the active sites of previously characterized HAD
enzymes, to complete the second half-reaction. The proposed mechanism is
reminiscent of two-Mg(2+) ion catalysis utilized by DNA and RNA polymerases and
many nucleases. The structure also provides an explanation for the inhibitory
effect of Ca(2+).
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Figure 5.
FIGURE 5. a, states along the reaction pathway based on
determined structures. I, the initial state with site 1 occupied
by Mg^2+; II, HisB-N with histidinol phosphate substrate modeled
on the structure of histidinol complex; III, phosphoaspartate
intermediate with Mg^2+ occupying sites 1 and 2; IV, release of
phosphate and Mg^2+ from site 2; b, superposition of the active
site residues of HisB-N (gray) with E. coli AphA (orange). Red
spheres represent water molecules; violet spheres show metal
binding sites. The orientations of Asp^12 and its equivalent in
AphA are different. Asp^12 is stabilized by hydrogen bonds
(blue) to Arg^11, site 2, and bridging water W4. Asp^46 of AphA
is hydrogen-bonded (orange) to Arg^114, approaching from the
opposite direction to Arg^11 of HisB-N, and to the of phosphate
oxygen in the position of water W1. Only HisB residues are
labeled.
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Figure 6.
FIGURE 6. Changes in the HisB-N substrate binding region
along the reaction pathway shown in a surface representation.
The residues that undergo conformational changes due to binding
of the substrate and/or metal ions, namely Glu^18, Phe^23, and
Arg^132 are shown as sticks under semitransparent surface. All
structures are shown in the same orientation. a,
HisB-N·Mg; b, HisB-N·Mg/histidinol; c,
HisB-N·Ca/pAsp; and d, HisB-N·Mg/sulfate.
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The above figures are
reprinted
by permission from the ASBMB:
J Biol Chem
(2006,
281,
37930-37941)
copyright 2006.
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