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Information on EC 2.7.1.48 - uridine/cytidine kinase and Organism(s) Thermus thermophilus and UniProt Accession Q5SKR5
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2.7.1.48 uridine/cytidine kinaseIUBMB Comments
The enzyme, found in prokaryotes and eukaryotes, phosphorylates both uridine and cytidine to their monophosphate forms. The enzyme from Escherichia coli prefers GTP to ATP. The human enzyme also catalyses the phosphorylation of several cytotoxic ribonucleoside analogs. cf. EC 2.7.1.213, cytidine kinase.
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Word Map
- 2.7.1.48
- thymidine
-
salvage
- uracil
-
phosphoribosyltransferase
- phosphorylase
- orotate
- ump
- 5-fluorouracil
- 5-fluorouridine
- deoxycytidine
- 5-azacytidine
- 6-azauridine
- 3-deazauridine
- orotidine
- pyrazofurin
- medicine
- 5-fluoro-2'-deoxyuridine
-
3huridine
- dihydrouracil
- drug development
- diagnostics
- biotechnology
The taxonomic range for the selected organisms is: Thermus thermophilus
The enzyme appears in selected viruses and cellular organisms
The enzyme appears in selected viruses and cellular organisms
Synonyms
uridine kinase, uridine-cytidine kinase, uridine/cytidine kinase, uridine-cytidine kinase 2, uckl1, uridine/cytidine kinase 2, uridine cytidine kinase 2, uridine phosphokinase, 
more
topSYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
ATP:uridine 5'-phosphotransferase
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kinase, uridine (phosphorylating)
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pyrimidine ribonucleoside kinase
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uridine kinase
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uridine phosphokinase
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REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
phospho group transfer
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PATHWAY SOURCE
PATHWAYS
BRENDA
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SYSTEMATIC NAME
IUBMB Comments
ATP:uridine/cytidine 5'-phosphotransferase
The enzyme, found in prokaryotes and eukaryotes, phosphorylates both uridine and cytidine to their monophosphate forms. The enzyme from Escherichia coli prefers GTP to ATP. The human enzyme also catalyses the phosphorylation of several cytotoxic ribonucleoside analogs. cf. EC 2.7.1.213, cytidine kinase.
CAS REGISTRY NUMBER
COMMENTARY
9026-39-5
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SUBSTRATE
PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
additional information
?
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molecular mechanisms of substrate specificities of uridine-cytidine kinase, overview
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ATP + uridine
ADP + UMP
substrate of mutant ttCKs Y93H and Y93Q, not of wild-type ttCK Y93
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?
ATP + uridine
ADP + UMP
reaction of ttCK enzyme mutants Y93H and Y93Q only, no activity with the wild-type enzyme
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-
?
NATURAL SUBSTRATE
NATURAL PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
ATP + uridine
ADP + UMP
substrate of mutant ttCKs Y93H and Y93Q, not of wild-type ttCK Y93
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-
?
COFACTOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
UTP
inhibition of wild-type ttCK Y93, no inhibition of mutant ttCK H93
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
additional information
additional information
-
Michaelis-Menten kineics
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additional information
additional information
Michaelis-Menten kineics
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additional information
additional information
Michaelis-Menten kinetics in a coupled assay with lactate dehydrogenase and pyruvate kinase
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additional information
additional information
reaction kinetics and thermodynamics of wild-type and Y93 mutant enzymes, overview
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TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
malfunction
physiological function
metabolism
uridine-cytidine kinase is the rate-limiting enzyme in the pyrimidine-nucleotide salvage pathway, overview
additional information
malfunction
mutagenesis of Tyr93 in ttCK reveals that the uridine phosphorylation activity is restored only by replacement of Tyr93 with His or Gln
malfunction
UCK from Thermus thermophilus HB8 loses catalytic activity on uridine due to lack of a substrate binding ability and possesses an unusual amino acid, i.e. tyrosine 93 (Tyr93) at the binding site, whereas histidine (His) is located in the other UCKs. Mutagenesis experiments reveal that a replacement of Tyr93 by His or glutamine (Gln) recovers catalytic activity on uridine
physiological function
in contrast to other UCK enzymes, UCK of Thermus thermophilus HB8 (ttCK) phosphorylates only cytidine
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
additional information
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mechanism underlying nucleoside specificity, overview. The histidine residue located near the functional group at position 4 of cytidine or uridine in most UCKs is substituted with tyrosine, Tyr93, in ttCK, due to the naturally occuring polymorphism Y93, the UCK homologue ttCK of Thermus thermophilus HB8 has, unlike other UCKs, a substrate specificity towards only cytidine and shows no inhibition by UTP, uridine does not bind to ttCK as substrate. Replacement of Tyr93 by histidine or glutamine endows ttCK with phosphorylation activity toward uridine
additional information
mechanism underlying nucleoside specificity, overview. The histidine residue located near the functional group at position 4 of cytidine or uridine in most UCKs is substituted with tyrosine, Tyr93, in ttCK, due to the naturally occuring polymorphism Y93, the UCK homologue ttCK of Thermus thermophilus HB8 has, unlike other UCKs, a substrate specificity towards only cytidine and shows no inhibition by UTP, uridine does not bind to ttCK as substrate. Replacement of Tyr93 by histidine or glutamine endows ttCK with phosphorylation activity toward uridine
additional information
molecular dynamics simulations on the wild-type Thermus thermophilus enzyme, two mutant ttCKs, and a human UCK bound to cytidine and three protonation forms of uridine to elucidate their substrate specificity, overview. Three residues, Tyr88, Tyr/His/Gln93 and Arg152 in ttCKs, are important for recognizing the substrates. Arg152 contributes to induce a closed form of the binding site to retain the substrate, and the N3 atom of uridine needs to be deprotonated. Although Tyr88 tightly binds cytidine, it does not sufficiently bind uridine because of lack of the hydrogen bonding. His/Gln93 complements the interaction of Tyr88 and raises the affinity of ttCK to uridine. The crucial distinction between Tyr and His or Gln is a role in the hydrogen-bonding network. Therefore, the ability to form both hydrogen-bonding donor and acceptor is required to bind both uridine and cytidine. Residue interactions and kinetics. Tyr59 and Phe90 form Pi-Pi stackings and CH-Pi interaction with cytidine, and Ile113 interacts with cytidine via a hydrophobic interaction. Tyr88, His93, and Arg152 form hydrogen bonds with the cytidine base moiety, and Asp60 and Arg142 anchor the ribose moiety, role of His/Gln93 in complementing the interaction between Tyr88 and the substrate
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
additional information
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mechanism underlying nucleoside specificity, overview
additional information
mechanism underlying nucleoside specificity, overview
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
additional information
secondary structure analysis of enzyme ttCK, overview
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
in complex with cytidine and beta,gamma-methyleneadenosine 5'-triphosphate, sitting drop vapor diffusion method, using 0.1 M NaCl, 0.1 M bicine, pH 9.0, and 20% (w/v) polyethylene glycol monomethyl ether 550
purified recombinant enzyme H93Y variant ttCK free or incomplex with CMP, sitting-drop vapor diffusion method, mixing of 500 nl of 25 mg/ml protein, with 1 mM CMP, 1 mM ADP, and 1 mM MgCl2 in case of the enzyme complex, with 500 nl well solution containing 67% 40% v/v isopropanol, 15% w/v PEG 8000, and 0.1 M imidazole, pH 6.5, and 33% distilled water for the free enzyme, and containing 50% w/v PEG 200, and 0.1 M Tris-HCl, pH 8.0, for the enzyme complex, 20°C, X-ray diffraction structure determination and analysis at 2.25 A resolution, molecular replacement
purified recombinant Y93H ttCK in complex with cytidine and AMPPCP, X-ray diffraction structure determination and analysis at 2.49 A resolution
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
Y93A
site-directed mutagenesis, the mutation restores the uridine kinase activity that has been lost in the wild-type enzyme
Y93C
site-directed mutagenesis, the mutation restores the uridine kinase activity that has been lost in the wild-type enzyme
Y93D
site-directed mutagenesis, the mutation restores the uridine kinase activity that has been lost in the wild-type enzyme
Y93E
site-directed mutagenesis, the mutation restores the uridine kinase activity that has been lost in the wild-type enzyme
Y93F
site-directed mutagenesis, the mutation restores the uridine kinase activity that has been lost in the wild-type enzyme
Y93G
site-directed mutagenesis, the mutation restores the uridine kinase activity that has been lost in the wild-type enzyme
Y93H
Y93I
site-directed mutagenesis, the mutation restores the uridine kinase activity that has been lost in the wild-type enzyme
Y93L
site-directed mutagenesis, the mutation restores the uridine kinase activity that has been lost in the wild-type enzyme
Y93M
site-directed mutagenesis, the mutation restores the uridine kinase activity that has been lost in the wild-type enzyme
Y93N
site-directed mutagenesis, the mutation restores the uridine kinase activity that has been lost in the wild-type enzyme
Y93P
site-directed mutagenesis, the mutation restores the uridine kinase activity that has been lost in the wild-type enzyme
Y93Q
Y93S
site-directed mutagenesis, the mutation restores the uridine kinase activity that has been lost in the wild-type enzyme
Y93T
site-directed mutagenesis, the mutation restores the uridine kinase activity that has been lost in the wild-type enzyme
Y93V
site-directed mutagenesis, the mutation restores the uridine kinase activity that has been lost in the wild-type enzyme
Y93W
site-directed mutagenesis, the mutation does not restore the uridine kinase activity that has been lost in the wild-type enzyme
additional information
Y93H
site-directed mutagenesis, the mutant enzyme shows restored activity on uridine in contrast to the wild-type enzyme. The mutant has higher binding affinities to cytidine than the wild-type
Y93H
site-directed mutagenesis, the mutation restores the uridine kinase activity that has been lost in the wild-type enzyme
Y93Q
site-directed mutagenesis, the mutant enzyme shows restored activity on uridine in contrast to the wild-type enzyme. The mutant has higher binding affinities to cytidine than the wild-type
Y93Q
site-directed mutagenesis, the mutation restores the uridine kinase activity that has been lost in the wild-type enzyme
additional information
-
the histidine residue located near the functional group at position 4 of cytidine or uridine in most UCKs is substituted with tyrosine, Tyr93, in ttCK, due to the naturally occuring polymorphism Y93, the UCK homologue ttCK of Thermus thermophilus HB8 has, unlike other UCKs, a substrate specificity towards only cytidine and shows no inhibition by UTP, uridine does not bind to ttCK as substrate. Replacement of Tyr93 by histidine or glutamine endows ttCK with phosphorylation activity toward uridine
additional information
the histidine residue located near the functional group at position 4 of cytidine or uridine in most UCKs is substituted with tyrosine, Tyr93, in ttCK, due to the naturally occuring polymorphism Y93, the UCK homologue ttCK of Thermus thermophilus HB8 has, unlike other UCKs, a substrate specificity towards only cytidine and shows no inhibition by UTP, uridine does not bind to ttCK as substrate. Replacement of Tyr93 by histidine or glutamine endows ttCK with phosphorylation activity toward uridine
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
ammonium sulfate precipitation, Toyopearl Phenyl-650 M column chromatography, Toyopearl SP-650 column chromatography and Superdex 200 gel filtration
recombinant ttCK from Escherichia coli strain Rosetta(DE3) by heat treatment at 60°C for 20 min, hydrophobic interaction chromatography, and gel filtration
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
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
enzyme H93Y variant ttCK, DNA and amino acid sequence determination and analysis, overexpression in Escherichia coli strain Rosetta(DE3)
recombinant expression of wild-type and mutant enzymes in Escherichia coli strain Rosetta(DE3)
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Tomoike, F.; Nakagawa, N.; Kuramitsu, S.; Masui, R.
A single amino acid limits the substrate specificity of Thermus thermophilus uridine-cytidine kinase to cytidine
Biochemistry
50
4597-4607
2011
Thermus thermophilus, Thermus thermophilus (Q5SKR5), Thermus thermophilus HB8 / ATCC 27634 / DSM 579, Thermus thermophilus HB8 / ATCC 27634 / DSM 579 (Q5SKR5)
Tomoike, F.; Nakagawa, N.; Kuramitsu, S.; Masui, R.
Structural and biochemical studies on the reaction mechanism of uridine-cytidine kinase
Protein J.
34
411-420
2015
Thermus thermophilus (Q5SKR5), Thermus thermophilus HB8 / ATCC 27634 / DSM 579 (Q5SKR5)
Tomoike, F.; Nakagawa, N.; Fukui, K.; Yano, T.; Kuramitsu, S.; Masui, R.
Indispensable residue for uridine binding in the uridine-cytidine kinase family
Biochem. Biophys. Rep.
11
93-98
2017
Thermus thermophilus (Q5SKR5), Homo sapiens (Q9BZX2), Homo sapiens, Thermus thermophilus DSM 579 (Q5SKR5), Thermus thermophilus ATCC 27634 (Q5SKR5)
Tanaka, W.; Shoji, M.; Tomoike, F.; Ujiie, Y.; Hanaoka, K.; Harada, R.; Kayanuma, M.; Kamiya, K.; Ishida, T.; Masui, R.; Kuramitsu, S.; Shigeta, Y.
Molecular mechanisms of substrate specificities of uridine-cytidine kinase
Biophys. Physicobiol.
13
77-84
2016
Thermus thermophilus (Q5SKR5), Thermus thermophilus DSM 579 (Q5SKR5), Thermus thermophilus ATCC 27634 (Q5SKR5)
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