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Literature summary for 2.7.1.48 extracted from

  • Tomoike, F.; Nakagawa, N.; Fukui, K.; Yano, T.; Kuramitsu, S.; Masui, R.
    Indispensable residue for uridine binding in the uridine-cytidine kinase family (2017), Biochem. Biophys. Rep., 11, 93-98 .
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

Cloned (Comment) Organism
gene UCK2, recombinant expression of GST-tagged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3) Homo sapiens
recombinant expression of wild-type and mutant enzymes in Escherichia coli strain Rosetta(DE3) Thermus thermophilus

Crystallization (Commentary)

Crystallization (Comment) Organism
analysis of cytidine-complex structure of hsUCK2 (PDB ID 1UEJ) Homo sapiens
purified recombinant Y93H ttCK in complex with cytidine and AMPPCP, X-ray diffraction structure determination and analysis at 2.49 A resolution Thermus thermophilus

Protein Variants

Protein Variants Comment Organism
H117Y site-directed mutagenesis, the mutation results in a loss of uridine phosphorylation activity of the enzyme Homo sapiens
Y93A site-directed mutagenesis, the mutation restores the uridine kinase activity that has been lost in the wild-type enzyme Thermus thermophilus
Y93C site-directed mutagenesis, the mutation restores the uridine kinase activity that has been lost in the wild-type enzyme Thermus thermophilus
Y93D site-directed mutagenesis, the mutation restores the uridine kinase activity that has been lost in the wild-type enzyme Thermus thermophilus
Y93E site-directed mutagenesis, the mutation restores the uridine kinase activity that has been lost in the wild-type enzyme Thermus thermophilus
Y93F site-directed mutagenesis, the mutation restores the uridine kinase activity that has been lost in the wild-type enzyme Thermus thermophilus
Y93G site-directed mutagenesis, the mutation restores the uridine kinase activity that has been lost in the wild-type enzyme Thermus thermophilus
Y93H site-directed mutagenesis, the mutation restores the uridine kinase activity that has been lost in the wild-type enzyme Thermus thermophilus
Y93I site-directed mutagenesis, the mutation restores the uridine kinase activity that has been lost in the wild-type enzyme Thermus thermophilus
Y93K site-directed mutagenesis, inactive mutant Thermus thermophilus
Y93L site-directed mutagenesis, the mutation restores the uridine kinase activity that has been lost in the wild-type enzyme Thermus thermophilus
Y93M site-directed mutagenesis, the mutation restores the uridine kinase activity that has been lost in the wild-type enzyme Thermus thermophilus
Y93N site-directed mutagenesis, the mutation restores the uridine kinase activity that has been lost in the wild-type enzyme Thermus thermophilus
Y93P site-directed mutagenesis, the mutation restores the uridine kinase activity that has been lost in the wild-type enzyme Thermus thermophilus
Y93Q site-directed mutagenesis, the mutation restores the uridine kinase activity that has been lost in the wild-type enzyme Thermus thermophilus
Y93R site-directed mutagenesis, inactive mutant Thermus thermophilus
Y93S site-directed mutagenesis, the mutation restores the uridine kinase activity that has been lost in the wild-type enzyme Thermus thermophilus
Y93T site-directed mutagenesis, the mutation restores the uridine kinase activity that has been lost in the wild-type enzyme Thermus thermophilus
Y93V site-directed mutagenesis, the mutation restores the uridine kinase activity that has been lost in the wild-type enzyme Thermus thermophilus
Y93W site-directed mutagenesis, the mutation does not restore the uridine kinase activity that has been lost in the wild-type enzyme Thermus thermophilus

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
additional information Michaelis-Menten kinetics in a coupled assay with lactate dehydrogenase and pyruvate kinase Homo sapiens
additional information
-
additional information Michaelis-Menten kinetics in a coupled assay with lactate dehydrogenase and pyruvate kinase Thermus thermophilus
0.042
-
cytidine pH 7.2, 25°C, recombinant wild-type enzyme UCK2 Homo sapiens
0.058
-
uridine pH 7.2, 25°C, recombinant wild-type enzyme UCK2 Homo sapiens
0.071
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93W Thermus thermophilus
0.072
-
cytidine pH 7.2, 25°C, recombinant wild-type enzyme ttCK Thermus thermophilus
0.1
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93F Thermus thermophilus
0.138
-
cytidine pH 7.2, 25°C, recombinant UCK2 mutant Homo sapiens
0.186
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93E Thermus thermophilus
0.215
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93C Thermus thermophilus
0.22
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93H Thermus thermophilus
0.225
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93S Thermus thermophilus
0.23
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93Q Thermus thermophilus
0.236
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93D Thermus thermophilus
0.36
-
uridine pH 7.2, 25°C, recombinant ttCK mutant Y93H Thermus thermophilus
0.383
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93A Thermus thermophilus
0.42
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93L Thermus thermophilus
0.47
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93N Thermus thermophilus
0.555
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93P Thermus thermophilus
0.574
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93V Thermus thermophilus
0.587
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93T Thermus thermophilus
0.62
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93G Thermus thermophilus
0.95
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93M Thermus thermophilus
0.965
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93I Thermus thermophilus
1.6
-
uridine pH 7.2, 25°C, recombinant ttCK mutant Y93Q Thermus thermophilus

Metals/Ions

Metals/Ions Comment Organism Structure
Mg2+ required Homo sapiens
Mg2+ required Thermus thermophilus

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
ATP + cytidine Homo sapiens
-
ADP + CMP
-
?
ATP + cytidine Thermus thermophilus
-
ADP + CMP
-
?
ATP + cytidine Thermus thermophilus DSM 579
-
ADP + CMP
-
?
ATP + cytidine Thermus thermophilus ATCC 27634
-
ADP + CMP
-
?
ATP + uridine Homo sapiens
-
ADP + UMP
-
?

Organism

Organism UniProt Comment Textmining
Homo sapiens Q9BZX2
-
-
Thermus thermophilus Q5SKR5
-
-
Thermus thermophilus ATCC 27634 Q5SKR5
-
-
Thermus thermophilus DSM 579 Q5SKR5
-
-

Purification (Commentary)

Purification (Comment) Organism
recombinant GST-tagged wild-type and mutant enzymes from Escherichia coli strain BL21(DE3) by glutathione affinity chromatography Homo sapiens
recombinant wild-type and mutant enzymes from Escherichia coli strain Rosetta(DE3) by heat treatment at 70°C for 20 min, followed by hydrophobic interaction chromatography, and two different steps of gel filtration Thermus thermophilus

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
ATP + cytidine
-
Homo sapiens ADP + CMP
-
?
ATP + cytidine
-
Thermus thermophilus ADP + CMP
-
?
ATP + cytidine
-
Thermus thermophilus DSM 579 ADP + CMP
-
?
ATP + cytidine
-
Thermus thermophilus ATCC 27634 ADP + CMP
-
?
ATP + uridine
-
Homo sapiens ADP + UMP
-
?
ATP + uridine reaction of ttCK enzyme mutants Y93H and Y93Q only, no activity with the wild-type enzyme Thermus thermophilus ADP + UMP
-
?
ATP + uridine reaction of ttCK enzyme mutants Y93H and Y93Q only, no activity with the wild-type enzyme Thermus thermophilus DSM 579 ADP + UMP
-
?
ATP + uridine reaction of ttCK enzyme mutants Y93H and Y93Q only, no activity with the wild-type enzyme Thermus thermophilus ATCC 27634 ADP + UMP
-
?

Subunits

Subunits Comment Organism
More secondary structure analysis of enzyme ttCK, overview Thermus thermophilus

Synonyms

Synonyms Comment Organism
hsUCK2
-
Homo sapiens
ttCK
-
Thermus thermophilus
UCK2
-
Homo sapiens
uridine-cytidine kinase 2
-
Homo sapiens

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
25
-
assay at Homo sapiens
25
-
assay at Thermus thermophilus

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.4
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93W Thermus thermophilus
1.1
-
uridine pH 7.2, 25°C, recombinant ttCK mutant Y93Q Thermus thermophilus
1.4
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93D Thermus thermophilus
1.5
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93E Thermus thermophilus
1.9
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93C Thermus thermophilus
2.8
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93Q Thermus thermophilus
2.9
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93G Thermus thermophilus
2.9
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93I Thermus thermophilus
3.1
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93T Thermus thermophilus
3.9
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93V Thermus thermophilus
4.1
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93N Thermus thermophilus
4.1
-
cytidine pH 7.2, 25°C, recombinant wild-type enzyme ttCK Thermus thermophilus
4.3
-
cytidine pH 7.2, 25°C, recombinant wild-type enzyme UCK2 Homo sapiens
4.5
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93A Thermus thermophilus
4.6
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93P Thermus thermophilus
5.2
-
uridine pH 7.2, 25°C, recombinant wild-type enzyme UCK2 Homo sapiens
5.9
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93F Thermus thermophilus
6.4
-
cytidine pH 7.2, 25°C, recombinant UCK2 mutant Homo sapiens
7.1
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93S Thermus thermophilus
7.2
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93M Thermus thermophilus
8.4
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93H Thermus thermophilus
9.2
-
uridine pH 7.2, 25°C, recombinant ttCK mutant Y93H Thermus thermophilus
10
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93L Thermus thermophilus

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7.2
-
assay at Homo sapiens
7.2
-
assay at Thermus thermophilus

General Information

General Information Comment Organism
malfunction mutagenesis of Tyr93 in ttCK reveals that the uridine phosphorylation activity is restored only by replacement of Tyr93 with His or Gln Thermus thermophilus
malfunction mutation H117Y of UCK2 results in a loss of uridine phosphorylation activity of the enzyme Homo sapiens
additional information uridine phosphorylation activity commonly depends on a single residue in the UCK family. Molecular docking analysis. Structure comparison of Thermus thermophilus enzyme ttCK and human enzyme hsUCK2 Thermus thermophilus
additional information uridine phosphorylation activity commonly depends on a single residue in the UCK family. Structure comparison of Thermus thermophilus enzyme ttCK and human enzyme hsUCK2 Homo sapiens
physiological function uridine-cytidine kinase (UCK) is one of the enzymes in the nucleoside salvage pathway. UCK generally converts both cytidine and uridine to nucleoside monophosphate using ATP as the phosphate donor Homo sapiens
physiological function uridine-cytidine kinase (UCK) is one of the enzymes in the nucleoside salvage pathway. UCK generally converts both cytidine and uridine to nucleoside monophosphate using ATP as the phosphate donor, but the UCK of Thermus thermophilus HB8 (ttCK) phosphorylates only cytidine. This cytidine-restricted activity is thought to depend on residue Tyr93 Thermus thermophilus

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
0.689
-
uridine pH 7.2, 25°C, recombinant ttCK mutant Y93Q Thermus thermophilus
4.04
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93I Thermus thermophilus
4.68
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93G Thermus thermophilus
5.05
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93V Thermus thermophilus
5.28
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93T Thermus thermophilus
5.63
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93W Thermus thermophilus
5.93
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93D Thermus thermophilus
7.58
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93M Thermus thermophilus
8.07
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93E Thermus thermophilus
8.29
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93P Thermus thermophilus
8.72
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93N Thermus thermophilus
8.84
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93C Thermus thermophilus
11.75
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93A Thermus thermophilus
12.17
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93Q Thermus thermophilus
23.81
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93L Thermus thermophilus
25.56
-
uridine pH 7.2, 25°C, recombinant ttCK mutant Y93H Thermus thermophilus
31.56
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93S Thermus thermophilus
38.18
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93H Thermus thermophilus
46.38
-
cytidine pH 7.2, 25°C, recombinant UCK2 mutant Homo sapiens
56.94
-
cytidine pH 7.2, 25°C, recombinant wild-type enzyme ttCK Thermus thermophilus
59
-
cytidine pH 7.2, 25°C, recombinant ttCK mutant Y93F Thermus thermophilus
89.66
-
uridine pH 7.2, 25°C, recombinant wild-type enzyme UCK2 Homo sapiens
102.38
-
cytidine pH 7.2, 25°C, recombinant wild-type enzyme UCK2 Homo sapiens