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Literature summary for 2.7.2.3 extracted from

  • Encalada, R.; Rojo-Dominguez, A.; Rodriguez-Zavala, J.S.; Pardo, J.P.; Quezada, H.; Moreno-Sanchez, R.; Saavedra, E.
    Molecular basis of the unusual catalytic preference for GDP/GTP in Entamoeba histolytica 3-phosphoglycerate kinase (2009), FEBS J., 276, 2037-2047.
    View publication on PubMed
Search for more literature for 2.7.2.3

Cloned(Commentary)

Cloned (Comment) Organism
mutant proteins cloned into the pGEM-T-easy vector, further cloned into the pET28 expression vector, wild-type and mutants overexpressed as N-terminal histidine-tailed recombinant proteins in Escherichia coli BL21DE3pLysS cells Entamoeba histolytica

Protein Variants

Protein Variants Comment Organism
E309M increases catalytic efficiency (Vm/Km) with ADP/ATP as a result of both, increased Vm and decreased Km values Entamoeba histolytica
E309Q increases catalytic efficiency (Vm/Km) with ADP/ATP as a result of both, increased Vm and decreased Km values Entamoeba histolytica
V311L increases catalytic efficiency (Vm/Km) with ADP/ATP as a result of both, increased Vm and decreased Km values. Is monomeric as the wild-type Entamoeba histolytica
Y239F increases catalytic efficiency (Vm/Km) with ADP/ATP as a result of both, increased Vm and decreased Km values Entamoeba histolytica
Y239F/E309M higher catalytic efficiency in the double mutant is achieved mainly due to an increased affinity for ADP/ATP with a concomitant diminished affinity for GDP/GTP. Is monomeric as the wild-type Entamoeba histolytica

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.28
-
 ADP recombinant purified mutant Y239F/E309M, at 37°C, pH 7.0 Entamoeba histolytica
0.348
-
 GTP recombinant purified mutant Y239F/E309M, at 37°C, pH 7.0 Entamoeba histolytica
0.42
-
 ATP wild-type, in cytosolic-enriched cellular extracts, at 37°C, pH 7.0 Saccharomyces cerevisiae
0.5
-
 ADP wild-type, in cytosolic-enriched cellular extracts, at 37°C, pH 7.0 Saccharomyces cerevisiae
0.587
-
 ATP recombinant purified mutant Y239F/E309M, at 37°C, pH 7.0 Entamoeba histolytica
0.637
-
 ATP recombinant purified mutant E309M, at 37°C, pH 7.0 Entamoeba histolytica
1.104
-
 ATP recombinant purified mutant V311L, at 37°C, pH 7.0 Entamoeba histolytica
1.313
-
 ATP recombinant purified mutant E309Q, at 37°C, pH 7.0 Entamoeba histolytica
1.439
-
 ATP recombinant purified wild-type, at 37°C, pH 7.0 Entamoeba histolytica
1.606
-
 ADP recombinant purified mutant E309M, at 37°C, pH 7.0 Entamoeba histolytica
1.696
-
 ADP recombinant purified mutant V311L, at 37°C, pH 7.0 Entamoeba histolytica
1.723
-
 ADP recombinant purified mutant E309Q, at 37°C, pH 7.0 Entamoeba histolytica
2.016
-
 ADP recombinant purified wild-type, at 37°C, pH 7.0 Entamoeba histolytica
2.244
-
 ATP recombinant purified mutant Y239F, at 37°C, pH 7.0 Entamoeba histolytica
2.693
-
 ADP recombinant purified mutant Y239F, at 37°C, pH 7.0 Entamoeba histolytica
5
-
 ATP wild-type, in cytosolic-enriched cellular extracts, at 37°C, pH 7.0 Entamoeba histolytica
7.4
-
 ADP wild-type, in cytosolic-enriched cellular extracts, at 37°C, pH 7.0 Entamoeba histolytica

Localization

Localization Comment Organism GeneOntology No. Textmining
cytosol
-
 Saccharomyces cerevisiae 5829
-
cytosol
-
 Entamoeba histolytica 5829
-

Metals/Ions

Metals/Ions Comment Organism Structure
MgCl2
-
 Entamoeba histolytica

Organism

Organism UniProt Comment Textmining
Entamoeba histolytica
-
 HM1:IMSS
-
Entamoeba histolytica HM1:IMSS
-
 HM1:IMSS
-
Saccharomyces cerevisiae
-
-
-
Saccharomyces cerevisiae BY4741
-
-
-

Purification (Commentary)

Purification (Comment) Organism
wild-type and mutants purified by metal-affinity chromatography, mutants purified more than 98% Entamoeba histolytica

Storage Stability

Storage Stability Organism
-22°C, 50% glycerol (v/v), 4-7 months Entamoeba histolytica

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
3-phospho-D-glycerate
-
 Saccharomyces cerevisiae ADP + 3-phospho-D-glyceroyl 1-phosphate
-
 r
3-phospho-D-glycerate
-
 Saccharomyces cerevisiae BY4741 ADP + 3-phospho-D-glyceroyl 1-phosphate
-
 r
ADP + 3-phospho-D-glyceroyl 1-phosphate
-
 Saccharomyces cerevisiae ATP + 3-phospho-D-glycerate
-
 r
ADP + 3-phospho-D-glyceroyl 1-phosphate
-
 Entamoeba histolytica ATP + 3-phospho-D-glycerate
-
 r
ADP + 3-phospho-D-glyceroyl 1-phosphate
-
 Saccharomyces cerevisiae BY4741 ATP + 3-phospho-D-glycerate
-
 r
ADP + 3-phospho-D-glyceroyl 1-phosphate
-
 Entamoeba histolytica HM1:IMSS ATP + 3-phospho-D-glycerate
-
 r
ATP + 3-phospho-D-glycerate
-
 Entamoeba histolytica ADP + 3-phospho-D-glyceroyl 1-phosphate
-
 r
ATP + 3-phospho-D-glycerate
-
 Entamoeba histolytica HM1:IMSS ADP + 3-phospho-D-glyceroyl 1-phosphate
-
 r
GTP + 3-phospho-D-glycerate
-
 Entamoeba histolytica GDP + 1,3-diphosphoglycerate
-
 ?
GTP + 3-phospho-D-glycerate
-
 Entamoeba histolytica HM1:IMSS GDP + 1,3-diphosphoglycerate
-
 ?

Subunits

Subunits Comment Organism
monomer
-
 Entamoeba histolytica

Synonyms

Synonyms Comment Organism
3-phosphoglycerate kinase
-
 Saccharomyces cerevisiae
3-phosphoglycerate kinase
-
 Entamoeba histolytica
PGK
-
 Saccharomyces cerevisiae
PGK
-
 Entamoeba histolytica

Temperature Stability [°C]

Temperature Stability Minimum [°C] Temperature Stability Maximum [°C] Comment Organism
50
-
 wild-type, mutants V311L and Y239F/E309Q incubated for 1-12 min do not show drastic reductions in activity at 50°C. 30% of their initial activities decay after 1 min incubation at 60°C Entamoeba histolytica

Cofactor

Cofactor Comment Organism Structure
ADP
-
 Saccharomyces cerevisiae
ATP exhibits 2.5times higher activity with ATP compared with GTP Saccharomyces cerevisiae
GTP
-
 Saccharomyces cerevisiae