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PDBsum entry 1uhk
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Luminescent protein
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PDB id
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1uhk
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References listed in PDB file
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Key reference
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Title
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The crystal structures of semi-Synthetic aequorins.
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Authors
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S.Toma,
K.T.Chong,
A.Nakagawa,
K.Teranishi,
S.Inouye,
O.Shimomura.
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Ref.
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Protein Sci, 2005,
14,
409-416.
[DOI no: ]
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PubMed id
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Abstract
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The photoprotein aequorin emits light by an intramolecular reaction in the
presence of a trace amount of Ca(2+). Semi-synthetic aequorins, produced by
replacing the coelenterazine moiety in aequorin with the analogues of
coelenterazine, show widely different sensitivities to Ca(2+). To understand the
structural basis of the Ca(2+)-sensitivity, we determined the crystal structures
of four semi-synthetic aequorins (cp-, i-, br- and n-aequorins) at resolutions
of 1.6-1.8 A. In general, the protein structures of these semi-synthetic
aequorins are almost identical to native aequorin. Of the four EF-hand domains
in the molecule, EF-hand II does not bind Ca(2+), and the loop of EF-hand IV is
clearly deformed. It is most likely that the binding of Ca(2+) with EF-hands I
and III triggers luminescence. Although little difference was found in the
overall structures of aequorins investigated, some significant differences were
found in the interactions between the substituents of coelenterazine moiety and
the amino acid residues in the binding pocket. The coelenterazine moieties in
i-, br-, and n-aequorins have bulky 2-substitutions, which can interfere with
the conformational changes of protein structure that follow the binding of
Ca(2+) to aequorin. In cp-aequorin, the cyclopentylmethyl group that substitutes
for the original 8-benzyl group does not interact hydrophobically with the
protein part, giving the coelenterazine moiety more conformational freedom to
promote the light-emitting reaction. The differences of various semi-synthetic
aequorins in Ca(2+)-sensitivity and reaction rate are explained by the
capability of the involved groups and structures to undergo conformational
changes in response to the Ca(2+)-binding.
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Figure 2.
Figure 2. Structural comparison of native aequorin,
semi-synthetic aequorins, and calmodulin. (A) Superimposed
structures of molecule A of native aequorin and semi-synthetic
aequorins. Native, cp-, br-, i-, and n-aequorin are shown in
yellow, pink, blue, sky blue, and green, respectively.
Apoaequorins are shown as C  trace models.
Coelenterazine moieties are drawn as ball-and-stick models with
same colors as above. The coelenterazine moiety and the loop
region of EF-hands I, III, and IV are surrounded by red, light
blue, green, and orange circles, respectively. W1 in native
aequorin is drawn as a sphere model in cyan. The OH group of the
2-substituent of native coelenterazine is stabilized by a
hydrogen-bonding network mediated by W1. W1 is absent in br-,
i-, and n-aequorin. The cyclopentyl group at the C8 position of
cp-coelenterazine can make neither a stacking interaction with
Lys39 nor a  - interaction with
Trp108. This figure was prepared with MolScript (Kraulis 1991)
and Raster3D (Merrit and Bacon 1997). (B) Superimposed
structures of EF-hands I, III, and IV of aequorin. (1) The
superimposed structures of EF-hands I (shown in sky blue circle
in Fig. 2A: 20-37), III (shown in green circle in Fig. 2A:
113-130), and IV (shown in orange circle in Fig. 2A: 149-166).
EF-hand IV in molecule A is colored cyan and EF-hand IV in
molecule B is colored blue. EF-hands I and III are colored green
and yellow-green, respectively. The blue dashed line, EF-IV(B2),
is the loop structure of EF-hand-IV(B) superimposed onto the
same region of EF-IV(A). (2) The superimposed structures of the
EF-hand I loop (residues 20-37) of aequorin (green) and
Ca^2+-bound (deep pink) and Ca^2+-unbound (cream) calmodulin.
The Ca^2+ is drawn as a sphere. All figures were prepared with
MolScript (Kraulis 1991). (C) Structure comparison between
aequorin and calmodulin. The superimposed structures of aequorin
(yellow-green) and Ca^2+-unbound calmodulin (cream). The traced
region of aequorin is from Thr103 to Pro189. The traced region
of calmodulin is from Ser81 to Lys148. The coelenterazine moiety
(CZH) is drawn as a ball-and-stick model. Dashed lines appeared
in the inside of the red circle are main-chain interactions,
green dotted lines are interactions in aequorin, and cream ones
are interactions in calmodulin.
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The above figure is
reprinted
by permission from the Protein Society:
Protein Sci
(2005,
14,
409-416)
copyright 2005.
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