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PDBsum entry 2dg1
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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 and mutational analyses of drp35 from staphylococcus aureus: a possible mechanism for its lactonase activity.
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Authors
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Y.Tanaka,
K.Morikawa,
Y.Ohki,
M.Yao,
K.Tsumoto,
N.Watanabe,
T.Ohta,
I.Tanaka.
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Ref.
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J Biol Chem, 2007,
282,
5770-5780.
[DOI no: ]
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PubMed id
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Abstract
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Drp35 is a protein induced by cell wall-affecting antibiotics or detergents; it
possesses calcium-dependent lactonase activity. To determine the molecular basis
of the lactonase activity, we first solved the crystal structures of Drp35 with
and without Ca(2+); these showed that the molecule has a six-bladed
beta-propeller structure with two calcium ions bound at the center of the
beta-propeller and surface region. Mutational analyses of evolutionarily
conserved residues revealed that the central calcium-binding site is essential
for the enzymatic activity of Drp35. Substitution of some other amino acid
residues for the calcium-binding residues demonstrated the critical
contributions of Glu(48), Asp(138), and Asp(236) to the enzymatic activity.
Differential scanning calorimetric analysis revealed that the loss of activity
of E48Q and D236N, but not D138N, was attributed to their inability to hold the
calcium ion. Further structural analysis of the D138N mutant indicates that it
lacks a water molecule bound to the calcium ion rather than the calcium ion
itself. Based on these observations and structural information, a possible
catalytic mechanism in which the calcium ion and its binding residues play
direct roles was proposed for the lactonase activity of Drp35.
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Figure 5.
FIGURE 5. Differential scanning calorimetry of Drp35, E48Q,
D138N, and D236N. Shown are heat capacity curves in 50 mM
acetate, pH 5.6, and 1 mM EDTA (A) and in 50 mM acetate, pH 5.6,
and 1 mM CaCl[2] (B). Solid lines, wild type; dotted lines,
E48Q; dashed lines, D138N; dashed and dotted lines, D236N.
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Figure 8.
FIGURE 8. Schematic representation of proposed mechanism
for lactonase activity of Drp35. A water molecule bound to Ca1
and Asp^138 is activated by Asp^138 and Ca1 (left). The
generated hydroxyl group attacks the carbon atom in the carbonyl
group of the substrate, and the oxygen atom whose covalent bond
is broken is protonated by Asp^236 (center).
3-(2-Hydroxyphenyl)propionic acid is generated.
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The above figures are
reprinted
by permission from the ASBMB:
J Biol Chem
(2007,
282,
5770-5780)
copyright 2007.
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