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PDBsum entry 2iz7
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Metal transport
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PDB id
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2iz7
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References listed in PDB file
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Key reference
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Title
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Function and structure of the molybdenum cofactor carrier protein from chlamydomonas reinhardtii.
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Authors
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K.Fischer,
A.Llamas,
M.Tejada-Jimenez,
N.Schrader,
J.Kuper,
F.S.Ataya,
A.Galvan,
R.R.Mendel,
E.Fernandez,
G.Schwarz.
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Ref.
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J Biol Chem, 2006,
281,
30186-30194.
[DOI no: ]
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PubMed id
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Abstract
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The molybdenum cofactor (Moco) forms the catalytic site in all eukaryotic
molybdenum enzymes and is synthesized by a multistep biosynthetic pathway. The
mechanism of transfer, storage, and insertion of Moco into the appropriate
apo-enzyme is poorly understood. In Chlamydomonas reinhardtii, a Moco carrier
protein (MCP) has been identified and characterized recently. Here we show
biochemical evidence that MCP binds Moco as well as the tungstate-substituted
form of the cofactor (Wco) with high affinity, whereas molybdopterin, the
ultimate cofactor precursor, is not bound. This binding selectivity points to a
specific metal-mediated interaction with MCP, which protects Moco and Wco from
oxidation with t((1/2)) of 24 and 96 h, respectively. UV-visible spectroscopy
showed defined absorption bands at 393, 470, and 570 nm pointing to
ene-diothiolate and protein side-chain charge transfer bonds with molybdenum. We
have determined the crystal structure of MCP at 1.6 Angstrom resolution using
seleno-methionated and native protein. The monomer constitutes a Rossmann fold
with two homodimers forming a symmetrical tetramer in solution. Based on
conserved surface residues, charge distribution, shape, in silico docking
studies, structural comparisons, and identification of an anionbinding site, a
prominent surface depression was proposed as a Moco-binding site, which was
confirmed by structure-guided mutagenesis coupled to substrate binding studies.
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Figure 3.
FIGURE 3. Structure of C. reinhardtii MCP. A, ribbon
presentation of the MCP monomer (MCP2 chain A) with  -strands
colored in gold and  -helices shown in
green. Residues Gly-70 and Ile-76 that border the stretch of
delocalized residues are highlighted. B and C, top view (B) and
side view (C)ofthe tetramer in ribbon presentation (MCP2). the
color code of the monomers is as follows: A, red;B, green;C,
blue;D, yellow. The monomers C and D were generated by applying
crystallographic symmetry operations. Interface I depicts the
hydrophobic contacts between A and B and C and D, and Interface
II describes the more hydrophilic contacts of B and C and A-D.
D, superposition of MCP2 (chain B, green) with T. thermophilus
HB8 hypothetical protein TT1465 (Protein Data Bank code 1WEK
chain F, gray), B. subtilis putative lysine decarboxylase
(Protein Data Bank code 1T35 chain B, light pink), and A.
thaliana lysine decarboxylase-like protein from gene AT2G37210
(Protein Data Bank code 2A33 chain A, lilac). Figures were
generated with MOLSCRIPT (52) and rendered with POVRAY.
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Figure 5.
FIGURE 5. The Moco-binding site. A ribbon presentation of
MCP (MCP2 chain B, green) and homologous structures is shown, as
shown (compare Fig. 3D). Cocrystallized phosphate (Protein Data
Bank code 1WEK) and sulfate molecules (Protein Data Bank codes
2A33, 1T35) are depicted in stick mode and color-coded according
to the protein structure (compare Fig. 3). The top-scored Moco
model from GOLD docking studies is shown in green. Three
structures have disordered residues in a similar region (MCP,
70-77; Protein Data Bank code 1WEK, 100-101-107; and Protein
Data Bank code 2A33, 78-87). Figures were prepared as in Fig. 3.
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The above figures are
reprinted
by permission from the ASBMB:
J Biol Chem
(2006,
281,
30186-30194)
copyright 2006.
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