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PDBsum entry 1vfs
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Contents |
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* Residue conservation analysis
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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 evidence that alanine racemase from a d-Cycloserine-Producing microorganism exhibits resistance to its own product.
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Authors
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M.Noda,
Y.Matoba,
T.Kumagai,
M.Sugiyama.
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Ref.
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J Biol Chem, 2004,
279,
46153-46161.
[DOI no: ]
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PubMed id
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Abstract
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Alanine racemase (ALR), an enzyme that catalyzes the interconversion of Ala
enantiomers, is essential for the synthesis of the bacterial cell wall. We have
shown that it is harder to inhibit the catalytic activity of ALR from
D-cycloserine (DCS)-producing Streptomyces lavendulae than that from Escherichia
coli by DCS. To obtain structural evidence for the fact that Streptomyces ALR
displays resistance to DCS, we determined the precise nature of the x-ray
crystal structures of the cycloserine-free and cycloserine enantiomer-bound
forms of Streptomyces ALR at high resolutions. Streptomyces ALR takes a dimer
structure, which is formed by interactions between the N-terminal domain of one
monomer with the C-terminal domain of its partner. Each of the two active sites
of ALR, which is generated as a result of the formation of the dimer structure,
is composed of pyridoxal 5'-phosphate (PLP), the PLP-binding residue Lys(38),
and the amino acids in the immediate environment of the pyridoxal cofactor. The
current model suggests that each active site of Streptomyces ALR maintains a
larger space and takes a more rigid conformation than that of Bacillus
stearothermophilus ALR determined previously. Furthermore, we show that
Streptomyces ALR results in a slow conversion to a final form of a pyridoxal
derivative arising from either isomer of cycloserine, which inhibits the
catalytic activity noncompetitively. In fact, the slow conversion is confirmed
by the fact that each enzyme bound cycloserine derivative, which is bound to
PLP, takes an asymmetric structure.
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Figure 6.
FIG. 6. Active site around the DCS- or LCS-bound PLP
derivative. DCS-bound derivatives at sites A and B are shown in
a and b, respectively. The LCS-bound derivatives at sites A and
B are shown in c and d, respectively. The carbon atoms of the
residues from each monomer are shown in orange and cyan. The
carbon atoms of the residues from the PLP derivative, designated
SCP, are shown in purple.
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Figure 7.
FIG. 7. Proposed mechanism of inactivation for ALR by DCS
or LCS. DCS and LCS are converted to an identical final aromatic
adduct through transamination, transamination, and
tautomerization steps. The DCS- or LSC-bound PLP intermediate
following the transamination step can be racemized to each
counterpart by the same reaction as used for the true substrates.
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The above figures are
reprinted
by permission from the ASBMB:
J Biol Chem
(2004,
279,
46153-46161)
copyright 2004.
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Secondary reference #1
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Title
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Self-Protection mechanism in d-Cycloserine-Producing streptomyces lavendulae. Gene cloning, Characterization, And kinetics of its alanine racemase and d-Alanyl-D-Alanine ligase, Which are target enzymes of d-Cycloserine.
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Authors
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M.Noda,
Y.Kawahara,
A.Ichikawa,
Y.Matoba,
H.Matsuo,
D.G.Lee,
T.Kumagai,
M.Sugiyama.
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Ref.
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J Biol Chem, 2004,
279,
46143-46152.
[DOI no: ]
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PubMed id
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Figure 3.
FIG. 3. Frame analysis of the 3.3-kb DNA fragment from
DCS-producing S. lavendulae. Three complete open reading frames
were designated orf1, orf2, and orf3. orf1 encodes Ala racemase.
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Figure 7.
FIG. 7. Resistance to DCS in E. coli carrying ALR and/or
DDL. The survival (%) by a given concentration of DCS was
expressed as a ratio of E. coli harboring each plasmid grown in
the presence of DCS to the same cells grown in the absence of
DCS. The cell growth was monitored as A[600 nm]. A, plasmids
carrying alrS and K12alr are shown as open and closed triangles,
respectively. B, plasmids carrying ddlS, ddlA, and ddlB are
shown as open squares, closed squares, and closed diamonds,
respectively. C, plasmids carrying alrS-ddlS, K12alr-ddlA, and
K12alr-ddlB are shown as open diamonds, closed squares, and
closed diamonds, respectively. E. coli harboring pET-21a(+) was
used as a control strain (A-C, closed circles).
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The above figures are
reproduced from the cited reference
with permission from the ASBMB
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