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PDBsum entry 1e8m
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Hydrolase/hydrolase inhibitor
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PDB id
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1e8m
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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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Structures of prolyl oligopeptidase substrate/inhibitor complexes. Use of inhibitor binding for titration of the catalytic histidine residue.
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Authors
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V.Fülöp,
Z.Szeltner,
V.Renner,
L.Polgár.
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Ref.
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J Biol Chem, 2001,
276,
1262-1266.
[DOI no: ]
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PubMed id
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Abstract
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Structure determination of the inactive S554A variant of prolyl oligopeptidase
complexed with an octapeptide has shown that substrate binding is restricted to
the P4-P2' region. In addition, it has revealed a hydrogen bond network of
potential catalytic importance not detected in other serine peptidases. This
involves a unique intramolecular hydrogen bond between the P1' amide and P2
carbonyl groups and another between the P2' amide and Nepsilon2 of the catalytic
histidine 680 residue. It is argued that both hydrogen bonds promote proton
transfer from the imidazolium ion to the leaving group. Another complex formed
with the product-like inhibitor benzyloxycarbonyl-glycyl-proline, indicating
that the carboxyl group of the inhibitor forms a hydrogen bond with the
Nepsilon2 of His(680). Because a protonated histidine makes a stronger
interaction with the carboxyl group, it offers a possibility of the
determination of the real pK(a) of the catalytic histidine residue. This was
found to be 6.25, lower than that of the well studied serine proteases. The new
titration method gave a single pK(a) for prolyl oligopeptidase, whose reaction
exhibited a complex pH dependence for k(cat)/K(m), and indicated that the
observed pK(a) values are apparent. The procedure presented may be applicable
for other serine peptidases.
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Figure 1.
Fig. 1. Stereo view of the peptide/inhibitor binding site
of prolyl oligopeptidase. A, octapeptide binding. B,
Z-Gly-Pro-OH binding to the S554A variant. The bound ligands are
shown darker than the protein residues. The SIGMAA (28) weighted
2mF[o]   F[c]
electron density using phases from the final model is contoured
at 1  level,
where represents
the root-mean-square electron density for the unit cell.
Contours more than 1.4 Å from any of the displayed atoms
have been removed for clarity. C, covalently bound inhibitor
Z-Pro-prolinal to Ser554 of the wild type enzyme (drawn from
Protein Data Bank code 1qfs (14)). Dashed lines indicate
hydrogen bonds (drawn with MolScript (29, 30)).
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Figure 2.
Fig. 2. A, the pH rate profiles for the reaction of
prolyl oligopeptidase with the octapeptide. The reactions were
performed in the presence (  ) and
absence (  circle )
of 0.5 M NaCl. The broken lines calculated from Equation 1 stand
for the two pH-dependent forms in the presence of 0.5 M NaCl. B,
formation of enzyme-inhibitor complex as a function of pH. The
association constants (1/K[i]) were calculated from Equation 3
for prolyl oligopeptidase and Z-Gly-Pro-OH in the presence (
) and
absence ( circle )
of 0.5 M NaCl. First-order rate constants were measured with
2-20 nM enzyme and 0.29 µM Z-Gly-Pro-Nap as substrate.
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The above figures are
reprinted
by permission from the ASBMB:
J Biol Chem
(2001,
276,
1262-1266)
copyright 2001.
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Secondary reference #1
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Title
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Catalysis of serine oligopeptidases is controlled by a gating filter mechanism.
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Authors
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V.Fülöp,
Z.Szeltner,
L.Polgár.
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Ref.
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Embo Rep, 2000,
1,
277-281.
[DOI no: ]
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PubMed id
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Figure 1.
Figure 1 Oxidation of prolyl oligopeptidase variants. Reaction
of the enzymes was carried out with 1.0 mM oxidized glutathione
at pH 9.0 and 28C. Open circles, C78A/C255T variant; filled
circles, C255T/Q397C variant. The curves represent single
exponential decays.
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Figure 2.
Figure 2 Electron density around the disulfide bond of the
C255T/Q397C variant of prolyl oligopeptidase. The SIGMAA (Read,
1986) weighted 2mF[o] - DF[c] electron density using phases from
the final model is contoured at 1  level,
where represents
the r.m.s. electron density for the unit cell. Contours >1.4
from any of the displayed atoms have been removed for clarity.
The position of the Glu397 side chain of the wild-type enzyme is
also shown in thin lines. The picture was drawn with MolScript
(Kraulis, 1991; Esnouf, 1997).
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The above figures are
reproduced from the cited reference
which is an Open Access publication published by Macmillan Publishers Ltd
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Secondary reference #2
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Title
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Prolyl oligopeptidase: an unusual beta-Propeller domain regulates proteolysis.
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Authors
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V.Fülöp,
Z.Böcskei,
L.Polgár.
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Ref.
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Cell, 1998,
94,
161-170.
[DOI no: ]
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PubMed id
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Figure 4.
Figure 4. Comparison of the Fold of the Noncatalytic Domain
of Prolyl Oligopeptidase with a Typical β-Propeller
Structure(A) The protein chain of the β-propeller domain of
prolyl oligopeptidase is colored as in Figure 2 and viewed
perpendicular to that, down the pseudo 7-fold axis. The β
sheets of the seven blades are joined in succession (β1/1 to
β7/4, cf. Figure 1) around the central axis. The “Velcro”
is not closed; there are only hydrophobic interactions between
the first (blue) and last (green) blades. Residues (Lys82,
Glu134, His180, Asp242, Lys389, and Lys390) narrowing the
entrance to the tunnel of the propeller are shown in a
ball-and-stick representation.(B) The structure of G-protein β
subunit (PDB entry 1tbg). The “Velcro” is closed between the
two termini of the polypeptide chain by the main chain hydrogen
bonds between the N terminus (blue) and the three antiparallel
β strands from the C terminus (green). (Drawn with MolScript
and rendered with Raster3D.)
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Figure 6.
Figure 6. Surface Representation of Prolyl
OligopeptidaseThe molecular surface is superimposed on the
polypeptide chain. The picture shows a slab of the molecule,
hence the cropping of the chain. The large cavity extends from
the central tunnel of the β propeller to the catalytic domain
and is accessible through the narrow hole at the bottom of the
propeller. The covalently bound inhibitor, Z-Pro-prolinal, is
shown in a ball-and-stick representation. The molecular surface
was calculated by the method published by [11], and the figure
was prepared using XOBJECTS (M. E. M. Noble, Oxford, unpublished
program).
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The above figures are
reproduced from the cited reference
with permission from Cell Press
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