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PDBsum entry 1m4l
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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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Refined structure of bovine carboxypeptidase a at 1.25 a resolution.
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Authors
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A.Kilshtain-Vardi,
M.Glick,
H.M.Greenblatt,
A.Goldblum,
G.Shoham.
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Ref.
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Acta Crystallogr D Biol Crystallogr, 2003,
59,
323-333.
[DOI no: ]
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PubMed id
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Abstract
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The crystal structure of the bovine zinc metalloproteinase carboxypeptidase A
(CPA) has been refined to 1.25 A resolution based on room-temperature X-ray
synchrotron data. The significantly improved structure of CPA at this resolution
(anisotropic temperature factors, R factor = 10.4%, R(free) = 14.5%) allowed the
modelling of conformational disorders of side chains, improved the description
of the protein solvent network (375 water molecules) and provided a more
accurate picture of the interactions between the active-site zinc and its
ligands. The calculation of standard uncertainties in individual atom positions
of the refined model of CPA allowed the deduction of the protonation state of
some key residues in the active site and confirmed that Glu72 and Glu270 are
negatively charged in the resting state of the enzyme at pH 7.5. These results
were further validated by theoretical calculations that showed significant
reduction of the pK(a) of these side chains relative to solution values. The
distance between the zinc-bound solvent molecule and the metal ion is strongly
suggestive of a neutral water molecule and not a hydroxide ion in the resting
state of the enzyme. These findings could support both the general acid/general
base mechanism, as well as the anhydride mechanism suggested for CPA.
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Figure 1.
Figure 1 Electron-density `omit' map around the active site of
native CPA at 1.25 Å resolution. Electron-density contour
levels are at +4.5  (cyan).
Superimposed on the density is the final 1.25 Å model of CPA
(ball-and-stick representation, common atom colour codes),
featuring the zinc and its ligands, as well as the catalytic
residues Arg127 and Glu270.
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Figure 5.
Figure 5 A stereoview of the active site of native CPA (stick
model) following the proton-addition algorithm. Atoms are
coloured according to conventional codes (C atoms, yellow; N
atoms, blue; O atoms, red; H atoms, white; Zn atoms, purple).
Zinc-ligand interactions are indicated (dashed lines) and their
distances are displayed (Å). The zinc-bound water is indicated
by `W'.
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The above figures are
reprinted
by permission from the IUCr:
Acta Crystallogr D Biol Crystallogr
(2003,
59,
323-333)
copyright 2003.
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