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Literature summary for 2.6.1.1 extracted from

  • Fernandez, F.J.; de Vries, D.; Pena-Soler, E.; Coll, M.; Christen, P.; Gehring, H.; Vega, M.C.
    Structure and mechanism of a cysteine sulfinate desulfinase engineered on the aspartate aminotransferase scaffold (2012), Biochim. Biophys. Acta, 1824, 339-349.
    View publication on PubMed

Cloned(Commentary)

Cloned (Comment) Organism
expressed in Escherichia coli Escherichia coli

Crystallization (Commentary)

Crystallization (Comment) Organism
the crystal triple mutant I33Q/Y214Q/R280Y is determined to relate the observed changes in reaction kinetics to the changes in active-site structure Escherichia coli

Protein Variants

Protein Variants Comment Organism
I33Q/Y214Q/R280Y substitution of three active-site residues in Escherichia coli L-aspartateartate aminotransferase highly reduces kcat for transamination reaction. Ratio of L-cysteine sulfinate desulfinase to transaminase activity is increased by 100000fold in mutant. kcat for desulfination of L-cysteine sulfinate increases to 0.5/sec (from 0.05/sec in wild-type enzyme). kcat for beta-decarboxylation of L-aspartateartate increases from below 0.0001/sec to 0.07/sec Escherichia coli

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
3.1
-
L-aspartate pH 7.5, 25°C, mutant I33Q/Y214Q/R280Y Escherichia coli
4
-
L-aspartate pH 7.5, 25°C, wild-type Escherichia coli
5.2
-
L-glutamate pH 7.5, 25°C, mutant I33Q/Y214Q/R280Y Escherichia coli
37
-
L-glutamate pH 7.5, 25°C, wild-type Escherichia coli

Organism

Organism UniProt Comment Textmining
Escherichia coli P00509
-
-

Purification (Commentary)

Purification (Comment) Organism
EcAspAT is purified by anion exchange chromatography in a Fractogel EMD DEAE 650-S column, pooled fractions are dialyzed against 20 mM sodium phosphate, pH 7.5, and loaded onto a ceramic hydroxyapatite column Escherichia coli

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
L-aspartate + 2-oxoglutarate
-
Escherichia coli oxaloacetate + L-glutamate
-
r

Synonyms

Synonyms Comment Organism
EcAspAT wild-type enzyme shows weak desulfination activity and decarboxylation activity Escherichia coli
L-aspartateartate aminotransferase
-
Escherichia coli

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
25
-
assay at Escherichia coli

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.03
-
L-glutamate pH 7.5, 25°C, mutant I33Q/Y214Q/R280Y Escherichia coli
0.13
-
L-aspartate pH 7.5, 25°C, mutant I33Q/Y214Q/R280Y Escherichia coli
530
-
L-aspartate pH 7.5, 25°C, wild-type Escherichia coli
670
-
L-glutamate pH 7.5, 25°C, wild-type Escherichia coli

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7.5
-
assay at Escherichia coli

Cofactor

Cofactor Comment Organism Structure
pyridoxal 5'-phosphate
-
Escherichia coli
pyridoxamine 5'-phosphate
-
Escherichia coli

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
0.006
-
L-glutamate pH 7.5, 25°C, mutant I33Q/Y214Q/R280Y Escherichia coli
0.042
-
L-aspartate pH 7.5, 25°C, mutant I33Q/Y214Q/R280Y Escherichia coli
18
-
L-glutamate pH 7.5, 25°C, wild-type Escherichia coli
133
-
L-aspartate pH 7.5, 25°C, wild-type Escherichia coli