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Literature summary for 3.1.3.48 extracted from

  • Zhang, X.Y.; Bishop, A.C.
    Engineered inhibitor sensitivity in the WPD loop of a protein tyrosine phosphatase (2008), Biochemistry, 47, 4491-4500.
    View publication on PubMed

Cloned(Commentary)

Cloned (Comment) Organism
DNA and amino acid sequence determination and analysis, sequence comparisons, expression of the His6-tagged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3), subcloning in strain DH5alpha Homo sapiens

Protein Variants

Protein Variants Comment Organism
E187C site-directed mutagenesis, altered kinetics compared to the wild-type enzyme Homo sapiens
E187C/S188C site-directed mutagenesis, altered kinetics compared to the wild-type enzyme Homo sapiens
F183C site-directed mutagenesis, altered kinetics compared to the wild-type enzyme Homo sapiens
G184C site-directed mutagenesis, altered kinetics compared to the wild-type enzyme Homo sapiens
additional information development of an engineering method for insertion of a chemical off switch into the catalytically essential loop region, the socalled WPD loop, of a protein tyrosine phosphatase using a combination of point mutations and amino acid insertions, the mutants displays FlAsH-binding cysteine residues, overview. Inhibition of the FlAsH-sensitized TCPTP mutants is rapid and specific, and strong FlAsH sensitivity is observed in mutants that contain as few as two cysteine point mutations in their engineered WPD loops Homo sapiens
P181C site-directed mutagenesis, altered kinetics compared to the wild-type enzyme Homo sapiens
P181C/E187C site-directed mutagenesis, altered kinetics compared to the wild-type enzyme Homo sapiens
P181C/P186C site-directed mutagenesis, altered kinetics compared to the wild-type enzyme Homo sapiens
P186C site-directed mutagenesis, altered kinetics compared to the wild-type enzyme Homo sapiens
P186C/E187C site-directed mutagenesis, altered kinetics compared to the wild-type enzyme Homo sapiens
V185C site-directed mutagenesis, altered kinetics compared to the wild-type enzyme Homo sapiens

Inhibitors

Inhibitors Comment Organism Structure
FlAsH the small molecule biarsenical fluorescein derivative, FlAsH, is no inhibitor of the wild-type enzyme but of the engineered WPD insertion mutant enzyme that displays FlAsH-binding cysteine residues. Inhibition of the FlAsH-sensitized TCPTP mutants is rapid and specific, and strong FlAsH sensitivity is observed in mutants that contain as few as two cysteine point mutations in their engineered WPD loops, minimization of FlAsH-binding determinants, overview Homo sapiens
additional information no inhibition by EDTA at 1 mM Homo sapiens

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
additional information kinetics of WPD loop insertion cysteine mutants, overview Homo sapiens
0.54
-
4-nitrophenyl phosphate pH 7.0, 22°C, mutant P181C Homo sapiens
0.85
-
4-nitrophenyl phosphate pH 7.0, 22°C, mutant P181C/E187C Homo sapiens
1.3
-
4-nitrophenyl phosphate pH 7.0, 22°C, mutant G184C Homo sapiens
1.9
-
4-nitrophenyl phosphate pH 7.0, 22°C, mutant P181C/P186C Homo sapiens
2.3
-
4-nitrophenyl phosphate pH 7.0, 22°C, mutant E187C/S188C Homo sapiens
2.5
-
4-nitrophenyl phosphate pH 7.0, 22°C, wild-type enzyme Homo sapiens
2.5
-
4-nitrophenyl phosphate pH 7.0, 22°C, mutant P186C Homo sapiens
2.7
-
4-nitrophenyl phosphate pH 7.0, 22°C, mutant E187C Homo sapiens
2.7
-
4-nitrophenyl phosphate pH 7.0, 22°C, mutant V185C Homo sapiens
3.1
-
4-nitrophenyl phosphate pH 7.0, 22°C, mutant F183C Homo sapiens
3.1
-
4-nitrophenyl phosphate pH 7.0, 22°C, mutant P186C/E187C Homo sapiens

Organism

Organism UniProt Comment Textmining
Homo sapiens P17706
-
-

Purification (Commentary)

Purification (Comment) Organism
recombinant His6-tagged enzyme from Escherichia coli Homo sapiens

Source Tissue

Source Tissue Comment Organism Textmining
T-lymphocyte
-
Homo sapiens
-

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
4-nitrophenyl phosphate + H2O
-
Homo sapiens 4-nitrophenol + phosphate
-
?
DADEpYLIPQQG + H2O a phosphopeptide substrate Homo sapiens DADEYLIPQQG + phosphate
-
?

Synonyms

Synonyms Comment Organism
protein tyrosine phosphatase
-
Homo sapiens
PTP
-
Homo sapiens
T-cell PTP
-
Homo sapiens
TCPTP
-
Homo sapiens

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
22
-
assay at Homo sapiens

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.83
-
4-nitrophenyl phosphate pH 7.0, 22°C, mutant F183C Homo sapiens
1.7
-
4-nitrophenyl phosphate pH 7.0, 22°C, mutant P186C/E187C Homo sapiens
3.2
-
4-nitrophenyl phosphate pH 7.0, 22°C, mutant P181C/P186C Homo sapiens
3.5
-
4-nitrophenyl phosphate pH 7.0, 22°C, mutant E187C Homo sapiens
3.7
-
4-nitrophenyl phosphate pH 7.0, 22°C, mutant P186C Homo sapiens
3.9
-
4-nitrophenyl phosphate pH 7.0, 22°C, mutant V185C Homo sapiens
4.4
-
4-nitrophenyl phosphate pH 7.0, 22°C, mutant G184C Homo sapiens
5.1
-
4-nitrophenyl phosphate pH 7.0, 22°C, wild-type enzyme Homo sapiens
5.4
-
4-nitrophenyl phosphate pH 7.0, 22°C, mutant E187C/S188C Homo sapiens
6.4
-
4-nitrophenyl phosphate pH 7.0, 22°C, mutant P181C/E187C Homo sapiens
9.8
-
4-nitrophenyl phosphate pH 7.0, 22°C, mutant P181C Homo sapiens

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7
-
assay at Homo sapiens

Ki Value [mM]

Ki Value [mM] Ki Value maximum [mM] Inhibitor Comment Organism Structure
additional information
-
additional information inhibition kinetics of WPD loop insertion cysteine mutants with FlAsH, overview Homo sapiens

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
0.26
-
4-nitrophenyl phosphate pH 7.0, 22°C, mutant F183C Homo sapiens
0.53
-
4-nitrophenyl phosphate pH 7.0, 22°C, mutant P186C/E187C Homo sapiens
1.3
-
4-nitrophenyl phosphate pH 7.0, 22°C, mutant E187C Homo sapiens
1.5
-
4-nitrophenyl phosphate pH 7.0, 22°C, mutant P186C Homo sapiens
1.5
-
4-nitrophenyl phosphate pH 7.0, 22°C, mutant V185C Homo sapiens
1.7
-
4-nitrophenyl phosphate pH 7.0, 22°C, mutant P181C/P186C Homo sapiens
2.1
-
4-nitrophenyl phosphate pH 7.0, 22°C, wild-type enzyme Homo sapiens
2.4
-
4-nitrophenyl phosphate pH 7.0, 22°C, mutant E187C/S188C Homo sapiens
3.4
-
4-nitrophenyl phosphate pH 7.0, 22°C, mutant G184C Homo sapiens
7.6
-
4-nitrophenyl phosphate pH 7.0, 22°C, mutant P181C/E187C Homo sapiens
18
-
4-nitrophenyl phosphate pH 7.0, 22°C, mutant P181C Homo sapiens