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PDBsum entry 2civ
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Oxidoreductase
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PDB id
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2civ
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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 structures of chloroperoxidase with its bound substrates and complexed with formate, Acetate, And nitrate.
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Authors
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K.Kühnel,
W.Blankenfeldt,
J.Terner,
I.Schlichting.
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Ref.
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J Biol Chem, 2006,
281,
23990-23998.
[DOI no: ]
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PubMed id
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Abstract
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Chloroperoxidase (CPO) is a heme-thiolate enzyme that catalyzes hydrogen
peroxide-dependent halogenation reactions. Structural data on substrate binding
have not been available so far. CPO was therefore crystallized in the presence
of iodide or bromide. One halide binding site was identified at the surface near
a narrow channel that connects the surface with the heme. Two other halide
binding sites were identified within and at the other end of this channel.
Together, these sites suggest a pathway for access of halide anions to the
active site. The structure of CPO complexed with its natural substrate
cyclopentanedione was determined at a resolution of 1.8 A. This is the first
example of a CPO structure with a bound organic substrate. In addition,
structures of CPO bound with nitrate, acetate, and formate and of a ternary
complex with dimethylsulfoxide (Me2SO) and cyanide were determined. These
structures have implications for the mechanism of compound I formation. Before
binding to the heme, the incoming hydrogen peroxide first interacts with
Glu-183. The deprotonated Glu-183 abstracts a proton from hydrogen peroxide. The
hydroperoxo-anion then binds at the heme, yielding compound 0. Glu-183
protonates the distal oxygen of compound 0, water is released, and compound I is
formed.
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Figure 3.
FIGURE 3. Complexes of CPO with acetate, nitrate, and
formate. A and B, acetate binds in two conformations at the
active site (shown in light and dark gray). A second acetate
molecule binds in the solvent channel near the iodide binding
site. Two orientations rotated by 90° are shown. Final
sigmaA-weighted 2mF[o] - DF[c] maps are shown with a contour
level of 1  . C, CPO complexed with
nitrate. An ethylene glycol molecule used as cryoprotectant is
located above the nitrate. The ethylene glycol binds at the
iodide specific binding site 3 and forms a hydrogen bond with
Asn-74. D, complex of CPO with formate in the presence of
ethylene glycol. A formate molecule is bound at the active site
and forms hydrogen bonds with an ethylene glycol molecule. E,
CPO-formate complex with a xylitol and sucrose-containing
cryoprotectant. The formate binds in two orientations (shown in
light and dark gray). F, solution spectra of CPO measured in the
presence or absence of sodium formate and/or ethylene glycol.
Measurements were done with solutions containing 0.06 mM CPO in
0.1 M sodium citrate, pH 3.6. Spectra were recorded at 20 °C
with a path length of 1 mm using a ND-1000 spectrophotometer
(NanoDrop Technologies Inc., Philadelphia, PA).
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Figure 4.
FIGURE 4. Proposed mechanism of compound I formation
catalyzed by chloroperoxidase.
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The above figures are
reprinted
by permission from the ASBMB:
J Biol Chem
(2006,
281,
23990-23998)
copyright 2006.
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