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Literature summary extracted from

  • Meyer, P.; Evrin, C.; Briozzo, P.; Joly, N.; Barzu, O.; Gilles, A.M.
    Structural and functional characterization of Escherichia coli UMP kinase in complex with its allosteric regulator GTP (2008), J. Biol. Chem., 283, 36011-36018.
    View publication on PubMed

Activating Compound

EC Number Activating Compound Comment Organism Structure
2.7.4.22 GTP
-
 Escherichia coli
2.7.4.22 GTP allosteric activator, the GTP-binding site, situated at 15 A from the UMP-binding site and at 24 A from the ATP-binding site, is delineated by two contiguous dimers. movements induced by binding GTP, overview. UTP and GTP are tightly coupled in both wild-type enzyme and N140A variant Escherichia coli

Cloned(Commentary)

EC Number Cloned (Comment) Organism
2.7.4.22 expression of His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3) Escherichia coli
2.7.4.22 His-tagged version expressed in Escherichia coli BL21(DE3) Escherichia coli

Crystallization (Commentary)

EC Number Crystallization (Comment) Organism
2.7.4.22 D159N, hanging drop vapor diffusion method Escherichia coli
2.7.4.22 purified recombinant UMP kinase mutant D159N, bound to GTP, by hanging drop vapour diffusion method, 0.004 ml of protein solution containing 4 mg/ml UMPK mutant in 50 mM Tris-HCl, pH 8.0, and 100 mM NaCl, mixed with 0.004 ml precipitant solution containing 22.5 mM GTP and 21.5% w/v PEG 400 in 100 mM sodium acetate, pH 4.6, equilibration against reservoir solution containing 43% PEG 400, 20°C, X-ray diffraction structure determination and analysis at 2.8-3.0 A resolution, molecular replacement and structure modelling Escherichia coli

Protein Variants

EC Number Protein Variants Comment Organism
2.7.4.22 D159N site-directed mutagenesis Escherichia coli
2.7.4.22 D159N used as reference enzyme Escherichia coli
2.7.4.22 D93A site-directed mutagenesis, inhibition by UTP appears significantly altered in the case of the D93A mutant as compared to the wild-type enzyme, the D93A substitution completely suppresses the related subunit-subunit hydrogen bonds between its main chain and Asp93, no inhibition by UTP in presence or absence of GTP. The Tm of the D93A variant is 10°C lower than that of the wild-type enzyme Escherichia coli
2.7.4.22 D93A/D159N D93 involved in hydrogen bond between the subunits of a dimer, mutation decreases the cooperativity for UTP binding and suppresses the reversal by GTP of UTP inhibition Escherichia coli
2.7.4.22 N140A site-directed mutagenesis, UTP and GTP are tightly coupled in both wild-type enzyme and N140A variant. The Tm of the mutant variant is 10°C lower than that of the wild-type enzyme Escherichia coli
2.7.4.22 N140A/D159N in N140A mutant protein is the cooperativity of inhibition caused by UTP suppressed Escherichia coli
2.7.4.22 N72A site-directed mutagenesis, no inhibition by UTP in presence or absence of GTP Escherichia coli
2.7.4.22 N72A/D159N N72 involved in hydrogen bond between the subunits Escherichia coli
2.7.4.22 N72A/D93A site-directed mutagenesis, the N72A mutation has less severe effects on enzyme activity regulation than the D93A substitution, reduced inhibition by UTP in absence of GTP, no inhibition in presence of GTP. The Tm of the mutant variant is 15°C lower than that of the wild-type enzyme Escherichia coli
2.7.4.22 N72A/D93A/D159N N72, D93 involved in hydrogen bonds between the subunits Escherichia coli

Inhibitors

EC Number Inhibitors Comment Organism Structure
2.7.4.22 UMP substrate inhibition Escherichia coli
2.7.4.22 UTP Mg-free competitive cooperative inhibitor, the inhibition is reversible by GTP. UTP and GTP are tightly coupled in both wild-type enzyme and N140A variant Escherichia coli

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
2.7.4.22 additional information
-
 additional information reaction kinetics Escherichia coli
2.7.4.22 0.0238
-
 UMP pH 7.4, 30°C, mutant D93A Escherichia coli
2.7.4.22 0.0238
-
 UMP D93A/D159N mutant protein, 2 mM ATP Escherichia coli
2.7.4.22 0.024
-
 UMP pH 7.4, 30°C, mutant N72A/D93A Escherichia coli
2.7.4.22 0.024
-
 UMP N72A/D93A/D159N mutant protein, 2 mM ATP Escherichia coli
2.7.4.22 0.0244
-
 UMP D93A/D159N mutant protein, 2 mM ATP, 0.5 mM GTP Escherichia coli
2.7.4.22 0.0246
-
 UMP N72A/D93A/D159N mutant protein, 2 mM ATP, 0.5 mM GTP Escherichia coli
2.7.4.22 0.0275
-
 UMP D93A/D159N mutant protein, 0.2 mM ATP, 0.5 mM GTP Escherichia coli
2.7.4.22 0.0321
-
 UMP pH 7.4, 30°C, mutant N72A Escherichia coli
2.7.4.22 0.0321
-
 UMP N72A/D159N mutant protein, 2 mM ATP Escherichia coli
2.7.4.22 0.0357
-
 UMP D93A/D159N mutant protein, 0.2 mM ATP Escherichia coli
2.7.4.22 0.046
-
 UMP pH 7.4, 30°C, wild-type enzyme Escherichia coli
2.7.4.22 0.046
-
 UMP D159N mutant protein, 2 mM ATP Escherichia coli
2.7.4.22 0.0478
-
 UMP N72A/D159N mutant protein, 2 mM ATP, 0.5 mM GTP Escherichia coli
2.7.4.22 0.05
-
 UMP D159N mutant protein, 0.2 mM ATP Escherichia coli
2.7.4.22 0.05
-
 UMP N140A/D159N mutant protein, 0.2 mM ATP Escherichia coli
2.7.4.22 0.051
-
 UMP D159N mutant protein, 2 mM ATP, 0.5 mM GTP Escherichia coli
2.7.4.22 0.0593
-
 UMP D159N mutant protein, 0.2 mM ATP, 0.5 mM GTP Escherichia coli
2.7.4.22 0.0739
-
 UMP N140A/D159N mutant protein, 0.2 mM ATP, 0.5 mM GTP Escherichia coli
2.7.4.22 0.0755
-
 UMP D93A/D159N mutant protein, 0.2 mM ATP, 0.5 mM GTP, 0.5 mM UTP Escherichia coli
2.7.4.22 0.0788
-
 UMP D93A/D159N mutant protein, 0.2 mM ATP, 0.5 mM UTP Escherichia coli
2.7.4.22 0.1086
-
 UMP N140A/D159N mutant protein, 0.2 mM ATP, 0.5 mM GTP, 0.5 mM UTP Escherichia coli
2.7.4.22 0.1812
-
 UMP N140A/D159N mutant protein, 0.2 mM ATP, 0.5 mM UTP Escherichia coli
2.7.4.22 0.19
-
 ATP pH 7.4, 30°C, wild-type enzyme Escherichia coli
2.7.4.22 0.19
-
 MgATP2- D159N mutant protein, 0.3 mM UMP Escherichia coli
2.7.4.22 0.2
-
 MgATP2- D159N mutant protein, 1 mM UMP, 0.5 mM GTP Escherichia coli
2.7.4.22 0.207
-
 UMP D159N mutant protein, 0.2 mM ATP, 0.5 mM GTP, 0.5 mM UTP Escherichia coli
2.7.4.22 0.21
-
 MgATP2- D159N mutant protein, 0.3 mM UMP Escherichia coli
2.7.4.22 0.21
-
 MgATP2- N140A/D159N mutant protein, 0.3 mM UMP, 0.5 mM GTP Escherichia coli
2.7.4.22 0.23
-
 MgATP2- N140A/D159N mutant protein, 0.3 mM UMP, 0.5 mM GTP, 0.5 mM UTP Escherichia coli
2.7.4.22 0.24
-
 MgATP2- D159N mutant protein, 0.3 mM UMP, 0.5 mM GTP Escherichia coli
2.7.4.22 0.25
-
 MgATP2- N140A/D159N mutant protein, 0.3 mM UMP Escherichia coli
2.7.4.22 0.26
-
 MgATP2- D159N mutant protein, 0.3 mM UMP, 0.5 mM GTP, 0.5 mM UTP Escherichia coli
2.7.4.22 0.27
-
 MgATP2- D93A/D159N mutant protein, 0.3 mM UMP Escherichia coli
2.7.4.22 0.27
-
 MgATP2- D93A/D159N mutant protein, 0.3 mM UMP, 0.5 mM GTP Escherichia coli
2.7.4.22 0.36
-
 ATP pH 7.4, 30°C, mutant D93A Escherichia coli
2.7.4.22 0.36
-
 MgATP2- D93A/D159N mutant protein, 0.3 mM UMP Escherichia coli
2.7.4.22 0.4
-
 MgATP2- N72A/D159N mutant protein, 1 mM UMP, 0.5 mM GTP Escherichia coli
2.7.4.22 0.41
-
 MgATP2- D93A/D159N mutant protein, 1 mM UMP, 0.5 mM GTP Escherichia coli
2.7.4.22 0.53
-
 MgATP2- N140A/D159N mutant protein, 0.3 mM UMP, 0.5 mM UTP Escherichia coli
2.7.4.22 0.59
-
 ATP pH 7.4, 30°C, mutants N72A and N72A/D93A Escherichia coli
2.7.4.22 0.59
-
 MgATP2- D93A/D159N mutant protein, 0.3 mM UMP, 0.5 mM GTP, 0.5 mM UTP Escherichia coli
2.7.4.22 0.59
-
 MgATP2- N72A/D159N mutant protein, 0.3 mM UMP Escherichia coli
2.7.4.22 0.59
-
 MgATP2- N72A/D93A/D159N mutant protein, 0.3 mM UMP Escherichia coli
2.7.4.22 0.64
-
 MgATP2- D93A/D159N mutant protein, 0.3 mM UMP, 0.5 mM UTP Escherichia coli
2.7.4.22 0.66
-
 MgATP2- N72A/D93A/D159N mutant protein, 1 mM UMP, 0.5 mM GTP Escherichia coli
2.7.4.22 0.77
-
 MgATP2- D159N mutant protein, 0.3 mM UMP, 0.5 mM UTP Escherichia coli
2.7.4.22 1.69
-
 UMP D159N mutant protein, 0.2 mM ATP, 0.5 mM UTP Escherichia coli

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
2.7.4.22 Mg2+ required Escherichia coli

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
2.7.4.22 ATP + UMP Escherichia coli the enzyme regulatory mechanisms involve GTP and UTP, overview ADP + UDP
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
2.7.4.22 Escherichia coli P0A7E9
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
2.7.4.22
-
 Escherichia coli
2.7.4.22 recombinant His-tagged wild-type and mutant enzymes from Escherichia coli strain BL21(DE3) by nickel affinity chromatography and gel filtration Escherichia coli

Specific Activity [micromol/min/mg]

EC Number Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
2.7.4.22 23.8
-
 D159N mutant protein, 0.3 mM UMP, 0.5 mM UTP Escherichia coli
2.7.4.22 30.4
-
 N140A/D159N mutant protein, 0.2 mM ATP, 0.5 mM UTP Escherichia coli
2.7.4.22 36.3
-
 D93A/D159N mutant protein, 0.2 mM ATP, 0.5 mM GTP, 0.5 mM UTP Escherichia coli
2.7.4.22 36.8
-
 D159N mutant protein, 0.2 mM ATP, 0.5 mM UTP Escherichia coli
2.7.4.22 38.1
-
 D93A/D159N mutant protein, 0.2 mM ATP, 0.5 mM UTP Escherichia coli
2.7.4.22 38.4
-
 D93A/D159N mutant protein, 0.2 mM ATP, 0.5 mM GTP Escherichia coli
2.7.4.22 40.7
-
 D93A/D159N mutant protein, 0.2 mM ATP Escherichia coli
2.7.4.22 46.1
-
 D159N mutant protein, 0.2 mM ATP Escherichia coli
2.7.4.22 46.6
-
 D159N mutant protein, 0.2 mM ATP, 0.5 mM GTP, 0.5 mM UTP Escherichia coli
2.7.4.22 48.9
-
 N140A/D159N mutant protein, 0.2 mM ATP Escherichia coli
2.7.4.22 51.4
-
 N72A/D93A/D159N mutant protein, 2 mM ATP, 0.5 mM GTP Escherichia coli
2.7.4.22 51.9
-
 D159N mutant protein, 0.2 mM ATP, 0.5 mM GTP Escherichia coli
2.7.4.22 52.7
-
 N72A/D93A/D159N mutant protein, 2 mM ATP Escherichia coli
2.7.4.22 55.7
-
 N140A/D159N mutant protein, 0.2 mM ATP, 0.5 mM GTP, 0.5 mM UTP Escherichia coli
2.7.4.22 57.6
-
 D93A/D159N mutant protein, 0.3 mM UMP, 0.5 mM GTP, 0.5 mM UTP Escherichia coli
2.7.4.22 58.2
-
 N140A/D159N mutant protein, 0.2 mM ATP, 0.5 mM GTP Escherichia coli
2.7.4.22 60.8
-
 D93A/D159N mutant protein, 0.3 mM UMP, 0.5 mM UTP Escherichia coli
2.7.4.22 61.5
-
 D93A/D159N mutant protein, 0.3 mM UMP Escherichia coli
2.7.4.22 61.6
-
 D159N mutant protein, 0.3 mM UMP Escherichia coli
2.7.4.22 61.6
-
 N72A/D159N mutant protein, 0.3 mM UMP Escherichia coli
2.7.4.22 62.3
-
 D93A/D159N mutant protein, 0.3 mM UMP, 0.5 mM GTP Escherichia coli
2.7.4.22 62.4
-
 D159N mutant protein, 0.3 mM UMP Escherichia coli
2.7.4.22 76.1
-
 N72A/D93A/D159N mutant protein, 1 mM UMP, 0.5 mM GTP Escherichia coli
2.7.4.22 77.1
-
 D93A/D159N mutant protein, 2 mM ATP Escherichia coli
2.7.4.22 78.3
-
 N140A/D159N mutant protein, 0.3 mM UMP, 0.5 mM UTP Escherichia coli
2.7.4.22 78.7
-
 N72A/D93A/D159N mutant protein, 0.3 mM UMP Escherichia coli
2.7.4.22 81.9
-
 N140A/D159N mutant protein, 0.3 mM UMP Escherichia coli
2.7.4.22 83.4
-
 D93A/D159N mutant protein, 2 mM ATP, 0.5 mM GTP Escherichia coli
2.7.4.22 84.2
-
 D159N mutant protein, 1 mM UMP, 0.5 mM GTP Escherichia coli
2.7.4.22 84.2
-
 N72A/D159N mutant protein, 1 mM UMP, 0.5 mM GTP Escherichia coli
2.7.4.22 90.7
-
 D93A/D159N mutant protein, 0.3 mM UMP Escherichia coli
2.7.4.22 91.7
-
 D159N mutant protein, 0.3 mM UMP, 0.5 mM GTP, 0.5 mM UTP Escherichia coli
2.7.4.22 92.5
-
 D159N mutant protein, 2 mM ATP Escherichia coli
2.7.4.22 92.5
-
 N72A/D159N mutant protein, 2 mM ATP Escherichia coli
2.7.4.22 93.9
-
 D93A/D159N mutant protein, 1 mM UMP, 0.5 mM GTP Escherichia coli
2.7.4.22 98.2
-
 N140A/D159N mutant protein, 0.3 mM UMP, 0.5 mM GTP, 0.5 mM UTP Escherichia coli
2.7.4.22 99.6
-
 D159N mutant protein, 0.3 mM UMP, 0.5 mM GTP Escherichia coli
2.7.4.22 100.3
-
 D159N mutant protein, 2 mM ATP, 0.5 mM GTP Escherichia coli
2.7.4.22 100.3
-
 N72A/D159N mutant protein, 2 mM ATP, 0.5 mM GTP Escherichia coli
2.7.4.22 110.5
-
 N140A/D159N mutant protein, 0.3 mM UMP, 0.5 mM GTP Escherichia coli

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
2.7.4.22 ATP + UMP the enzyme regulatory mechanisms involve GTP and UTP, overview Escherichia coli ADP + UDP
-
 ?
2.7.4.22 ATP + UMP movements induced by binding uridine nucleotides, GTP and UTP interaction mechanism, analysis, overview Escherichia coli ADP + UDP
-
 ?
2.7.4.22 MgATP2- + UMP
-
 Escherichia coli MgADP- + UDP
-
 ?
2.7.4.22 additional information Asp93 appears as one of the key amino acids in the transfer of information from the regulatory site to the active site Escherichia coli ?
-
 ?

Subunits

EC Number Subunits Comment Organism
2.7.4.22 hexamer quaternary structure, overview Escherichia coli
2.7.4.22 homohexamer crystal structure Escherichia coli

Synonyms

EC Number Synonyms Comment Organism
2.7.4.22 UMP kinase
-
 Escherichia coli

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
2.7.4.22 30
-
 assay at Escherichia coli

Temperature Stability [°C]

EC Number Temperature Stability Minimum [°C] Temperature Stability Maximum [°C] Comment Organism
2.7.4.22 additional information
-
 thermal stability of the N72A/D159N variant is close to that of the D159N variant, temperature of half-inactivation of the D93A/D159N variant is 10°C lower than that of the D159N variant, temperature of half-inactivation of the D93A/D159N variant is 15°C lower than that of the D159N variant, Escherichia coli

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
2.7.4.22 7.4
-
 assay at Escherichia coli

pH Range

EC Number pH Minimum pH Maximum Comment Organism
2.7.4.22 7.4 8.5
-
 Escherichia coli

Cofactor

EC Number Cofactor Comment Organism Structure
2.7.4.22 ATP
-
 Escherichia coli

Ki Value [mM]

EC Number Ki Value [mM] Ki Value maximum [mM] Inhibitor Comment Organism Structure
2.7.4.22 0.44
-
 UMP pH 7.4, 30°C, wild-type enzyme, in absence of GTP Escherichia coli
2.7.4.22 0.44
-
 UMP D159N mutant protein, 2 mM ATP Escherichia coli
2.7.4.22 10.9
-
 UMP pH 7.4, 30°C, mutant N72A/D93A, in absence of GTP Escherichia coli
2.7.4.22 10.9
-
 UMP N72A/D93A/D159N mutant protein, 2 mM ATP Escherichia coli