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PDBsum entry 1orf
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Hydrolase/hydrolase inhibitor
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PDB id
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1orf
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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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The oligomeric structure of human granzyme a is a determinant of its extended substrate specificity.
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Authors
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J.K.Bell,
D.H.Goetz,
S.Mahrus,
J.L.Harris,
R.J.Fletterick,
C.S.Craik.
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Ref.
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Nat Struct Biol, 2003,
10,
527-534.
[DOI no: ]
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PubMed id
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Abstract
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The cell death-inducing serine protease granzyme A (GzmA) has a unique
disulfide-linked quaternary structure. The structure of human GzmA bound to a
tripeptide CMK inhibitor, determined at a resolution of 2.4 A, reveals that the
oligomeric state contributes to substrate selection by limiting access to the
active site for potential macromolecular substrates and inhibitors. Unlike other
serine proteases, tetrapeptide substrate preferences do not correlate well with
natural substrate cleavage sequences. This suggests that the context of the
cleavage sequence within a macromolecular substrate imposes another level of
selection not observed with the peptide substrates. Modeling of inhibitors bound
to the GzmA active site shows that the dimer also contributes to substrate
specificity in a unique manner by extending the active-site cleft. The crystal
structure, along with substrate library profiling and mutagenesis, has allowed
us to identify and rationally manipulate key components involved in GzmA
substrate specificity.
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Figure 1.
Figure 1. Overall structure of dimeric human granzyme A. (a)
Ribbon diagram of dimeric human GzmA. The secondary structural
elements are colored in a gradient from N to C terminus. The
Ser195-His57-Asp102 catalytic triad and P1 coordinating Asp189
are shown in ball and stick (carbons, light green) as is the
D-Phe-Pro-Arg-chloromethylketone (carbons, cyan). The disulfide
linkage is depicted as space-filling model for residue 93 and
its symmetry mate. A sulfate ion contributed from the
crystallization solution is bound at the base of loop 184-B -197
by Arg186 and Arg188. (b) Surface representation mapped with
potentials shows the overall positive charge of the molecule
reflected by its pI > 9 and the distinct negative charge of
Asp189 emanating from the S1 pocket.
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Figure 3.
Figure 3. The active site of human granzyme A. (a)
Ball-and-stick representation of the bound inhibitor,
D-Phe-Pro-Arg-CMK (carbons, cyan) and residues that frame the
substrate binding pocket depicted in the context of the
molecular surface. The molecular surfaces of the proposed S1'
and S2' subsites are colored in magenta; S1 subsite, orange; S2,
blue; S3, red; S4, green. (b) Ligplot representation showing
direct interactions between GzmA and the bound inhibitor.
D-Phe-Pro-Arg-CMK bonds and carbons are cyan. Bonds between the
irreversible inhibitor and GzmA are magenta. (c) Stereo view of
the refined (2F[o] - F[c]) electron density for the CMK
inhibitor (carbons, yellow) bound to the GzmA active site
(carbons, gray).
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The above figures are
reprinted
by permission from Macmillan Publishers Ltd:
Nat Struct Biol
(2003,
10,
527-534)
copyright 2003.
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