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Literature summary for 2.7.7.27 extracted from

  • Meyer, C.R.; Yirsa, J.; Gott, B.; Preiss, J.
    A kinetic study of site-directed mutants of Escherichia coli ADP-glucose pyrophosphorylase: the role of residue 295 in allosteric regulation (1998), Arch. Biochem. Biophys., 352, 247-254.
    View publication on PubMed

Cloned(Commentary)

Cloned (Comment) Organism
expression is performed using bacterial strain G6MD3 Escherichia coli

Protein Variants

Protein Variants Comment Organism
G336D a higher activity enzyme form, 10fold decreased affinity for AMP than wild-type enzyme, higher apparent affinity for ATP than wild-type enzyme Escherichia coli
P295D extremely high activity in the absence of fructose 1,6-bisphosphate, 20fold decreased affinity for AMP than wild-type enzyme Escherichia coli
P295D/G336D the double mutant enzyme is more active in the absence of fructose 1,6-bisphosphate, with a higher affinity for fructose 1,6-bisphosphate and a lower apparent affinity for AMP than either single mutated enzyme Escherichia coli
P295E extremely high activity in the absence of fructose 1,6-bisphosphate, 10fold decreased affinity for AMP than wild-type enzyme, higher apparent affinity for ATP than wild-type enzyme Escherichia coli
P295G activity in the absence of fructose 1,6-bisphosphate is similar to wild-type enzyme Escherichia coli
P295G 3fold decreased affinity for AMP than wild-type enzyme, higher apparent affinity for ATP than wild-type enzyme Escherichia coli
P295N 3.4fold decreased affinity for AMP than wild-type enzyme, higher apparent affinity for ATP than wild-type enzyme Escherichia coli
P295Q 3.8fold decreased affinity for AMP than wild-type enzyme, higher apparent affinity for ATP than wild-type enzyme Escherichia coli

Inhibitors

Inhibitors Comment Organism Structure
AMP
-
Escherichia coli

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
additional information kinetic parameters Escherichia coli
2.2
-
ATP pH 7.0, 37°C, wild-type enzyme in the absence of fructose 1,6-bisphosphate, comparison of Km of wild-type and mutant enzymes in the presence and absence of fructose 1,6-bisphosphate Escherichia coli

Metals/Ions

Metals/Ions Comment Organism Structure
Mg2+
-
Escherichia coli

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
ATP + alpha-D-glucose 1-phosphate Escherichia coli
-
diphosphate + ADP-glucose
-
?

Organism

Organism UniProt Comment Textmining
Escherichia coli
-
-
-

Purification (Commentary)

Purification (Comment) Organism
wild-type and mutant enzymes, purification of wild-type and single mutant enzyme usually involves a 5 min/60°C heat treatment step, but the heat treatment step of the double mutant enzyme results in 20% loss of activity Escherichia coli

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
80 100 mutant enzymes Escherichia coli
100
-
wild-type enzyme Escherichia coli
110
-
double mutant enzymes Escherichia coli

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
ATP + alpha-D-glucose 1-phosphate
-
Escherichia coli diphosphate + ADP-glucose
-
?

Temperature Stability [°C]

Temperature Stability Minimum [°C] Temperature Stability Maximum [°C] Comment Organism
60
-
purification of wild-type and single mutant enzyme usually involves a 5 min/60°C heat treatment step, the heat treatment step of the double mutant enzyme results in 20% loss of activity Escherichia coli

Ki Value [mM]

Ki Value [mM] Ki Value maximum [mM] Inhibitor Comment Organism Structure
0.04
-
AMP pH 7.6, 37°C, wild-type enzyme Escherichia coli
0.094
-
AMP pH 7.0, 37°C, wild-type enzyme, in the presence of a saturating concentration of fructose 1,6-bisphosphate Escherichia coli
0.3
-
AMP pH 7.0, 37°C, mutant enzyme P295G, in the presence of a saturating concentration of fructose 1,6-bisphosphate, Escherichia coli
0.34
-
AMP pH 7.0, 37°C, mutant enzyme P295N, in the presence of a saturating concentration of fructose 1,6-bisphosphate Escherichia coli
0.38
-
AMP pH 7.0, 37°C, mutant enzyme P295Q, in the presence of a saturating concentration of fructose 1,6-bisphosphate Escherichia coli
0.95
-
AMP pH 7.0, 37°C, mutant enzyme G336D, in the presence of a saturating concentration of fructose 1,6-bisphosphate Escherichia coli
0.96
-
AMP pH 7.0, 37°C, mutant enzyme P295E, in the presence of a saturating concentration of fructose 1,6-bisphosphate Escherichia coli
2
-
AMP pH 7.0, 37°C, mutant enzyme P295D, in the presence of a saturating concentration of fructose 1,6-bisphosphate Escherichia coli
4.95
-
AMP pH 7.0, 37°C, double mutant enzyme P295D/G336D Escherichia coli