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PDBsum entry 1a5y
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References listed in PDB file
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Key reference
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Title
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Visualization of the cysteinyl-Phosphate intermediate of a protein-Tyrosine phosphatase by X-Ray crystallography.
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Authors
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A.D.Pannifer,
A.J.Flint,
N.K.Tonks,
D.Barford.
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Ref.
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J Biol Chem, 1998,
273,
10454-10462.
[DOI no: ]
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PubMed id
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Abstract
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Protein-tyrosine phosphatases (PTPs) are signal transduction enzymes that
catalyze the dephosphorylation of phosphotyrosine residues via the formation of
a transient cysteinyl-phosphate intermediate. The mechanism of hydrolysis of
this intermediate has been examined by generating a Gln-262 --> Ala mutant of
PTP1B, which allows the accumulation and trapping of the intermediate within a
PTP1B crystal. The structure of the intermediate at 2.5-A resolution reveals
that a conformationally flexible loop (the WPD loop) is closed over the entrance
to the catalytic site, sequestering the phosphocysteine intermediate and
catalytic site water molecules and preventing nonspecific phosphoryltransfer
reactions to extraneous phosphoryl acceptors. One of the catalytic site water
molecules, the likely nucleophile, forms a hydrogen bond to the putative
catalytic base, Asp-181. In the wild-type enzyme, the nucleophilic water
molecule would be coordinated by the side chain of Gln-262. In combination with
our previous structural data, we can now visualize each of the reaction steps of
the PTP catalytic pathway. The hydrolysis of the cysteinyl-phosphate
intermediate of PTPs is reminiscent of GTP hydrolysis by the GTPases, in that
both families of enzymes utilize an invariant Gln residue to coordinate the
attacking nucleophilic water molecule.
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Figure 6.
Fig. 6. Stereo views depicting the sequences of the
reaction pathway catalyzed by PTP1B. A, PTP1B apoenzyme, with
WPD loop open. B, PTP1B C215S Tyr(P)-Michaelis complex, with WPD
loop closed; Gln-262 swings out of the catalytic site. C, PTP1B
Q262A cysteinyl-phosphate intermediate complex, with WPD loop
closed. D, PTP1B vanadate complex, representing the transition
state of cysteinyl-phosphate hydrolysis, with WPD loop closed;
Gln-262 swings back into the catalytic site. E, PTP1B
tungstate-product complex, with WPD loop open.
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Figure 7.
Fig. 7. Schematic of the reaction mechanism catalyzed by
PTP1B. A, formation of the cysteinyl-phosphate intermediate. B,
hydrolysis of the cysteinyl-phosphate intermediate.
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The above figures are
reprinted
by permission from the ASBMB:
J Biol Chem
(1998,
273,
10454-10462)
copyright 1998.
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