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PDBsum entry 2zxc
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References listed in PDB file
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Key reference
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Title
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Mechanistic insights into the hydrolysis and synthesis of ceramide by neutral ceramidase.
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Authors
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T.Inoue,
N.Okino,
Y.Kakuta,
A.Hijikata,
H.Okano,
H.M.Goda,
M.Tani,
N.Sueyoshi,
K.Kambayashi,
H.Matsumura,
Y.Kai,
M.Ito.
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Ref.
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J Biol Chem, 2009,
284,
9566-9577.
[DOI no: ]
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PubMed id
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Abstract
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Ceramidase (CDase; EC 3.5.1.23) hydrolyzes ceramide to generate sphingosine and
fatty acid. The enzyme plays a regulatory role in a variety of physiological
events in eukaryotes and also functions as an exotoxin in particular bacteria.
The crystal structures of neutral CDase from Pseudomonas aeruginosa (PaCD) in
the C2-ceramide-bound and -unbound forms were determined at 2.2 and 1.4 A
resolutions, respectively. PaCD consists of two domains, and the Zn(2+)- and
Mg(2+)/Ca(2+)-binding sites are found within the center of the N-terminal domain
and the interface between the domains, respectively. The structural comparison
between the C2-ceramide-bound and unbound forms revealed an open-closed
conformational change occurring to loop I upon binding of C2-ceramide. In the
closed state, this loop sits above the Zn(2+) coordination site and over the
opening to the substrate binding site. Mutational analyses of residues
surrounding the Zn(2+) of PaCD and rat neutral CDase revealed that the cleavage
or creation of the N-acyl linkage of ceramide follows a similar mechanism as
observed for the Zn(2+)-dependent carboxypeptidases. The results provide an
understanding of the molecular mechanism of hydrolysis and synthesis of ceramide
by the enzyme. Furthermore, insights into the actions of PaCD and eukaryotic
neutral CDases as an exotoxin and mediators of sphingolipid signaling are also
revealed, respectively.
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Figure 3.
C2-Cer bound in the putative active site of PaCD and a
proposed reactionmechanism.A, the structural comparison between
substrate-unbound and C2 Cer-bound forms of PaCD. The
substrate-unbound form is shown in red, whereas the C2 Cer-bound
form is shown in green. B, the binding model of C2-Cer is shown
as a stick representation with oxygen atoms in red, nitrogen
atoms in blue, and carbon atoms in yellow. The electron density
map was calculated at the 1.0 σ level. C, schematic drawing of
the binding model of C2-Cer. The hydrogen bonds and the metal
coordinate ion structure was shown by dotted lines. D, a
proposed reaction mechanism of PaCD based on the structural data.
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Figure 5.
The proposed conformations of nCDase acting on membrane-bound
Cer (B and C) and free Cer (A and D). nCDase is shown as a
ribbon model. The type II integral membrane (A and B) and
soluble (C and D) forms of nCDase are shown.
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The above figures are
reprinted
by permission from the ASBMB:
J Biol Chem
(2009,
284,
9566-9577)
copyright 2009.
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