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PDBsum entry 2v8h
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References listed in PDB file
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Key reference
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Title
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Crystal structures of yeast beta-Alanine synthase complexes reveal the mode of substrate binding and large scale domain closure movements.
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Authors
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S.Lundgren,
B.Andersen,
J.Piskur,
D.Dobritzsch.
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Ref.
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J Biol Chem, 2007,
282,
36037-36047.
[DOI no: ]
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PubMed id
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Abstract
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Beta-alanine synthase is the final enzyme of the reductive pyrimidine catabolic
pathway, which is responsible for the breakdown of uracil and thymine in higher
organisms. The fold of the homodimeric enzyme from the yeast Saccharomyces
kluyveri identifies it as a member of the AcyI/M20 family of metallopeptidases.
Its subunit consists of a catalytic domain harboring a di-zinc center and a
smaller dimerization domain. The present site-directed mutagenesis studies
identify Glu(159) and Arg(322) as crucial for catalysis and His(262) and
His(397) as functionally important but not essential. We determined the crystal
structures of wild-type beta-alanine synthase in complex with the reaction
product beta-alanine, and of the mutant E159A with the substrate
N-carbamyl-beta-alanine, revealing the closed state of a dimeric AcyI/M20
metallopeptidase-like enzyme. Subunit closure is achieved by a approximately 30
degrees rigid body domain rotation, which completes the active site by
integration of substrate binding residues that belong to the dimerization domain
of the same or the partner subunit. Substrate binding is achieved via a salt
bridge, a number of hydrogen bonds, and coordination to one of the zinc ions of
the di-metal center.
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Figure 1.
FIGURE 1. Open and closed conformation of SkβAS. a,
stereoview of the homodimer of SkβAS-E159A (E159A_NCβA). The
two subunits are colored differently. Substrate molecules bound
in the respective active sites and Bicine molecules bound close
to the hinge region are shown as ball-and-stick models with
white and yellow carbon atoms, respectively. Zinc ions are
represented as black spheres. b, stereoview of the extended,
open enzyme conformation as observed for SkβAS-R322A (R322A).
The distance separating the C[  ]-atom positions of
D192 from both monomers in this enzyme state is given. Zinc ions
are shown as black spheres. c, schematic view of the subunit
backbones of E159A_NCβA (magenta), WT-βAla (white), E159A
(yellow), and R322A (blue) after superposition of the respective
catalytic domains. The rigid body hinge movement of the
dimerization domain from the open to the closed state is
indicated by an arrow and the approximate rotation angle is
given. d, stereoview of the domain interface in the open state
(R322A). The two subunits per homodimer are shown in white (A)
and yellow (B) cartoon representation, respectively. Residue
stretches interacting with each other in the closed but not in
the open state are colored identically (red: amino acids 118,
120, 160, 162 from subunit A and 305-310 from subunit B; green:
189, 190, 192 (A) and 192, 306 (B); bright green: 229, 231, 235
(A) and 262, 265, 269 (B); blue: 247, 249, 322, 365, 393-396,
403 (A) and 264 (B); cyan: 419-422 (A) and 259-261, 309-310 (B);
orange: 166, 359-360 (A); magenta: 161, 167, 249, 251 (A)).
Residues H262, R322, N309, and D192 are shown as stick models.
Black spheres represent zinc ions. e, stereoview of the domain
interface in the closed state (E159A_NCβA). The orientation of
the catalytic domain in monomer A is the same as in d. All
representations and color codes correspond to those used in
d.NCβA is shown as stick model with oxygen atoms in red,
nitrogen atoms in blue, and carbon atoms in yellow.
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Figure 3.
FIGURE 3. Comparison of SkβAS with PepV. a, stereoview of
the superimposed crystal structures of monomeric PepV (gray) and
dimeric E159A_NCβA. From the latter only one complete subunit
(yellow) and the dimerization domain of the partner subunit
(green) are shown. Stick models of NCβA (red), and a transition
state mimic bound to PepV (cyan) are included in the figure.
Zinc ions are represented as spheres, for SkβAS in orange and
for PepV in blue. b, stereoview of the superimposed active sites
of SkβAS (E159A_NCβA) and PepV. The dimensions of the
substrate binding cavities are outlined by the white surface for
PepV and by the orange surface for E159A_NCβA. The latter was
calculated after modeling of a glutamate at position 159 to
correspond to the active site of the wild-type enzyme. Zinc ions
are represented as orange (E159A_NCβA) and green spheres
(PepV). Stick models of substrate- and zinc-binding residues as
well as some other residues lining the cavity walls are shown
with green carbon atoms for PepV and yellow carbon atoms for the
SkβAS complex. For clarity, only the latter are labeled. The
transition state mimic Asp  [PO[2]CH[2]]AlaOH bound
to PepV is depicted with thicker sticks and carbon atoms in
cyan, while those of NCβA are shown in orange.
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The above figures are
reprinted
by permission from the ASBMB:
J Biol Chem
(2007,
282,
36037-36047)
copyright 2007.
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Secondary reference #1
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Title
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Yeast beta-Alanine synthase shares a structural scaffold and origin with dizinc-Dependent exopeptidases.
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Authors
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S.Lundgren,
Z.Gojković,
J.Piskur,
D.Dobritzsch.
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Ref.
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J Biol Chem, 2003,
278,
51851-51862.
[DOI no: ]
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PubMed id
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Figure 2.
FIG. 2. Comparison of the subunit structures of
dizinc-dependent exopeptidases and Sk  AS. The color scheme for
the latter is the same as described for Fig. 1a. For the
exopeptidases, the secondary structure elements are shown in
blue ( -strands) and red ( - and
3[10]-helices). The zinc ions are depicted as purple (for Sk
AS)
and blue (for exopeptidases) spheres.
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Figure 4.
FIG. 4. Structure-based alignment of the sequences of Sk
AS
and dizinc-dependent exopeptidases. Row 1, Sk AS; row
2, SGAP; row 3; APAP; row 4, CPG2; row 5, PepV; row 6, PepT. The
secondary structure elements (labeled as described for Fig. 1a)
are indicated by arrows for -strands and boxes for
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and 3[10]-helices, which are white for the catalytic domain and
light blue for the dimerization domain of Sk AS. For the
exopeptidases, only the sequences of the structurally aligned
parts are shown (uppercase letters). Pink number signs indicate
the zinc-coordinating residues. Amino acids labeled by boldface
letters indicate sequence conservation in all structurally
aligned sequence parts of exopeptidases. If the amino acid is
also conserved in the sequence of Sk AS, the residue is
marked additionally by pink background shading. Gray background
shading indicates sequence conservation between Sk AS and
any of the exopeptidases. For residues that are conserved and
aligned only in some of the six proteins, the corresponding
amino acid in the non-aligned proteins is given as a lowercase
italic letter if its spatial position only barely does not
fulfill the requirements defining structural alignment. A hyphen
is used when the residue is not present at the same position due
to either considerable differences in structure or deletions in
the sequence. Dotted lines indicate the absence of the
dimerization domain in SGAP and APAP.
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The above figures are
reproduced from the cited reference
with permission from the ASBMB
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Secondary reference #2
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Title
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Crystallization and preliminary X-Ray analysis of beta-Alanine synthase from the yeast saccharomyces kluyveri.
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Authors
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D.Dobritzsch,
Z.Gojković,
B.Andersen,
J.Piskur.
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Ref.
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Acta Crystallogr D Biol Crystallogr, 2003,
59,
1267-1269.
[DOI no: ]
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PubMed id
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Figure 1.
Figure 1 A crystal of recombinant S. kluyveri  AS
as grown by the hanging-drop method. The largest dimension is
[110]~ 0.2 mm.
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The above figure is
reproduced from the cited reference
with permission from the IUCr
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