Information on EC 2.4.2.2 - pyrimidine-nucleoside phosphorylase

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The expected taxonomic range for this enzyme is: Bacteria, Eukaryota

EC NUMBER
COMMENTARY
2.4.2.2
-
RECOMMENDED NAME
GeneOntology No.
pyrimidine-nucleoside phosphorylase
REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT
LITERATURE
2'-deoxyuridine + phosphate = uracil + 2-deoxy-alpha-D-ribose 1-phosphate
show the reaction diagram
molecular basis of catalytic chamber-assisted unfolding and cleavage of human insulin by human insulin-degrading enzyme, IDE utilizes the interaction of its exosite with the N-terminus of the insulin A chain, overview. The unique size, shape, charge distribution, and exosite of the IDE catalytic chamber contribute to its high affinity for insulin
-
-
-
thymidine + phosphate = thymine + 2-deoxy-alpha-D-ribose 1-phosphate
show the reaction diagram
(2)
-
-
-
uridine + phosphate = uracil + alpha-D-ribose 1-phosphate
show the reaction diagram
(1)
-
-
-
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
pentosyl group transfer
-
-
-
-
pentosyl group transfer
-
-
pentosyl group transfer
-
-
pentosyl group transfer
Q5SHF9
-
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
Metabolic pathways
-
-
pyrimidine deoxyribonucleosides degradation
-
-
Pyrimidine metabolism
-
-
pyrimidine ribonucleosides degradation
-
-
SYSTEMATIC NAME
IUBMB Comments
pyrimidine-nucleoside:phosphate alpha-D-ribosyltransferase
Unlike EC 2.4.2.3, uridine phosphorylase, and EC 2.4.2.4, thymidine phosphorylase, this enzyme can accept both the ribonucleoside uridine and the 2'-deoxyribonucleosides 2'-deoxyuridine and thymidine [3]. The reaction is reversible, and the enzyme does not distinguish between alpha-D-ribose 1-phosphate and 2-deoxy-alpha-D-ribose 1-phosphate in the synthetic direction.
SYNONYMS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
phosphorylase, pyrimidine nucleoside
-
-
-
-
Py-NPase
-
-
-
-
PYNP
-
-
-
-
pynpase
-
-
-
-
pynpase
-
-
pyrimidine nucleoside phosphorylase
-
-
-
-
pyrimidine nucleoside phosphorylase
-
-
pyrimidine nucleoside phosphorylase
-
-
pyrimidine nucleoside phosphorylase
Q5SHF9
-
pyrimidine ribonucleoside phosphorylase
-
-
-
-
TTHA1771
Q5SHF9
-
CAS REGISTRY NUMBER
COMMENTARY
9055-35-0
-
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
Enterobacter aerogenes CMCC 45103
-
-
-
Manually annotated by BRENDA team
Geobacillus stearothermophilus JTS 859
JTS 859
-
-
Manually annotated by BRENDA team
Haemophilus influenzae TH 6-2
TH 6-2
-
-
Manually annotated by BRENDA team
strain HB8
-
-
Manually annotated by BRENDA team
SUBSTRATE
PRODUCT                      
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
LITERATURE
(Substrate)
COMMENTARY
(Product)
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
(2'-deoxy-2'-fluoro-beta-D-arabinofuranosyl)uracil + phosphate
uracil + 2'-deoxy-2'-fluoro-beta-D-arabinofuranose 1-phosphate
show the reaction diagram
Q72HS4
-
1.6% substrate conversion after 30 min
-
?
1,2,4-triazole-3-carboxamide + alpha-D-ribose 1-phosphate
ribavirin + phosphate
show the reaction diagram
-
-
-
-
-
2'-deoxy-2'-fluorouridine + phosphate
?
show the reaction diagram
-
-
poor activity, 0.4% substrate conversion after 30 min
-
?
2'-deoxy-2'-fluorouridine + phosphate
uracil + 2'-deoxy-2'-fluoro-ribose 1-phosphate
show the reaction diagram
Q72HS4
-
15.6% substrate conversion after 30 min
-
?
4-thiothymidine + phosphate
4-thiothymin + alpha-D-ribose 1-phosphate
show the reaction diagram
-
uridine phosphorylase activity, see also EC 2.4.2.3
-
-
r
4-thiouridine + phosphate
4-thiouracil + alpha-D-ribose 1-phosphate
show the reaction diagram
-
thymidine phosphorylase activity, see also EC 2.4.2.4
-
-
r
5'-deoxy-5-fluorouridine + phosphate
5-fluorouracil + alpha-D-deoxyribose 1-phosphate
show the reaction diagram
-
-
5-fluorouracil is a chemotherapeutic agent and inactivated by dihydropyrimidine dihydrogenase activity
-
?
5-bromodeoxyuridine + phosphate
5-bromouracil + deoxyribose 1-phosphate
show the reaction diagram
-
-
-
-
?
5-bromodeoxyuridine + phosphate
5-bromouracil + deoxyribose 1-phosphate
show the reaction diagram
-
95% of the activity with uridine
-
-
?
5-bromodeoxyuridine + phosphate
5-bromouracil + deoxyribose 1-phosphate
show the reaction diagram
-
74% of the activity with uridine
-
-
?
5-bromodeoxyuridine + phosphate
5-bromouracil + 2-deoxy-D-ribose 1-phosphate
show the reaction diagram
-
-
-
-
?
5-bromouridine + phosphate
5-bromouracil + alpha-D-ribose 1-phosphate
show the reaction diagram
-
-
-
-
-
5-bromouridine + phosphate
5-bromouracil + alpha-D-ribose 1-phosphate
show the reaction diagram
-
40% of the activity with uridine
-
-
?
5-bromouridine + phosphate
5-bromouracil + alpha-D-ribose 1-phosphate
show the reaction diagram
-
95% of the activity with uridine
-
-
?
5-chlorodeoxyuridine + phosphate
5-chlorouracil + 2-deoxy-D-ribose 1-phosphate
show the reaction diagram
-
-
-
-
?
5-fluorodeoxyuridine + phosphate
5-fluorouracil + 2-deoxy-D-ribose 1-phosphate
show the reaction diagram
-
-
-
-
?
5-fluorouridine + phosphate
5-fluorouracil + alpha-D-ribose 1-phosphate
show the reaction diagram
-
-
-
-
?
5-iododeoxyuridine + phosphate
5-iodouracil + 2'-deoxy-D-ribose 1-phosphate
show the reaction diagram
-
-
-
-
?
5-iododeoxyuridine + phosphate
5-iodouracil + 2-deoxy-D-ribose 1-phosphate
show the reaction diagram
-
-
-
-
?
5-iodouridine + phosphate
5-iodouracil + alpha-D-ribose 1-phosphate
show the reaction diagram
-
-
-
-
?
5-methyluridine + phosphate
5-methyluracil + alpha-D-ribose 1-phosphate
show the reaction diagram
-
-
-
-
?
5-methyluridine + phosphate
5-methyluracil + alpha-D-ribose 1-phosphate
show the reaction diagram
-
-
-
-
?
5-methyluridine + phosphate
5-methyluracil + alpha-D-ribose 1-phosphate
show the reaction diagram
-
27% of the activity with uridine
-
-
?
5-methyluridine + phosphate
5-methyluracil + alpha-D-ribose 1-phosphate
show the reaction diagram
Geobacillus stearothermophilus JTS 859
-
-
-
-
?
5-trifluoromethyl-2'-deoxyuridine + phosphate
5-trifluoromethyluracil + 2-deoxy-D-ribose 1-phosphate
show the reaction diagram
-
-
-
-
?
cytidine + phosphate
cytosine + alpha-D-ribose 1-phosphate
show the reaction diagram
-
no activity
-
-
-
cytidine + phosphate
cytosine + alpha-D-ribose 1-phosphate
show the reaction diagram
-
8% of the activity with uridine
-
-
?
deoxyuridine + phosphate
uracil + 2-deoxy-D-ribose 1-phosphate
show the reaction diagram
-
-
-
-
?
deoxyuridine + phosphate
uracil + 2-deoxy-D-ribose 1-phosphate
show the reaction diagram
-
12% of the activity with uridine
-
-
?
deoxyuridine + phosphate
uracil + 2-deoxy-D-ribose 1-phosphate
show the reaction diagram
-
2'-deoxyuridine
-
-
?
deoxyuridine + phosphate
uracil + 2-deoxy-D-ribose 1-phosphate
show the reaction diagram
-
179% of the activity with uridine, by measuring the conversion of uracil to uridine or deoxyuridine a slight preference for ribose 1-phosphate over deoxyribose 1-phosphate is observed
-
-
r
deoxyuridine + phosphate
uracil + 2-deoxy-D-ribose 1-phosphate
show the reaction diagram
-
147% of the activity with uridine
-
-
?
deoxyuridine + phosphate
uracil + 2-deoxy-D-ribose 1-phosphate
show the reaction diagram
Geobacillus stearothermophilus JTS 859
-
2'-deoxyuridine
-
-
?
deoxyuridine + phosphate
uracil + 2-deoxy-D-ribose 1-phosphate
show the reaction diagram
Haemophilus influenzae TH 6-2
-
147% of the activity with uridine
-
-
?
deoxyuridine + phosphate
uracil + 2'-deoxy-D-ribose 1-phosphate
show the reaction diagram
-
-
-
-
?
pyrimidine nucleoside + phosphate
pyrimidine base + alpha-D-ribose 1-phosphate
show the reaction diagram
-
most pyrimidine nucleosides except for 4-amino-substituted species like deoxycytidine
or alpha-D-deoxyribose 1-phosphate
-
r
pyrimidine nucleoside + phosphate
pyrimidine base + alpha-D-ribose 1-phosphate
show the reaction diagram
Q5SHF9
most pyrimidine nucleosides except for 4-amino-substituted species like deoxycytidine
or alpha-D-deoxyribose 1-phosphate
-
r
ribofuranosyl thymine + phosphate
thymine + alpha-D-ribose 1-phosphate
show the reaction diagram
Haemophilus influenzae, Haemophilus influenzae TH 6-2
-
-
-
-
?
thymidine + phosphate
thymine + 2-deoxy-D-ribose 1-phosphate
show the reaction diagram
-
-
-
-
?
thymidine + phosphate
thymine + 2-deoxy-D-ribose 1-phosphate
show the reaction diagram
-
-
-
-
?
thymidine + phosphate
thymine + 2-deoxy-D-ribose 1-phosphate
show the reaction diagram
-
21% of the activity with uridine
-
-
?
thymidine + phosphate
thymine + 2-deoxy-D-ribose 1-phosphate
show the reaction diagram
-
139% of the activity with uridine
-
-
?
thymidine + phosphate
thymine + 2-deoxy-D-ribose 1-phosphate
show the reaction diagram
-
99% of the activity with uridine
-
-
?
thymidine + phosphate
thymine + 2-deoxy-D-ribose 1-phosphate
show the reaction diagram
Geobacillus stearothermophilus JTS 859
-
-
-
-
?
thymidine + phosphate
thymine + 2-deoxy-D-ribose 1-phosphate
show the reaction diagram
Haemophilus influenzae TH 6-2
-
99% of the activity with uridine
-
-
?
thymidine + phosphate
thymine + alpha-D-deoxyribose 1-phosphate
show the reaction diagram
Q5SHF9
-
-
-
r
thymidine + phosphate
thymine + 2'-deoxy-D-ribose 1-phosphate
show the reaction diagram
-
-
-
-
?
thymidine + phosphate
thymine + 2'-deoxy-D-ribose 1-phosphate
show the reaction diagram
-
-
-
-
?
thymidine + phosphate
thymine + 2'-deoxy-D-ribose 1-phosphate
show the reaction diagram
Q72HS4
-
-
-
?
thymidine + phosphate
thymine + 2'-deoxy-D-ribose 1-phosphate
show the reaction diagram
Q5XA29
-
-
-
?
uracil arabinoside + phosphate
uracil + alpha-D-arabinose 1-phosphate
show the reaction diagram
-
-
-
-
?
uridine + phosphate
uracil + alpha-D-ribose 1-phosphate
show the reaction diagram
-
-
-
-
-
uridine + phosphate
uracil + alpha-D-ribose 1-phosphate
show the reaction diagram
-
-
-
-
?
uridine + phosphate
uracil + alpha-D-ribose 1-phosphate
show the reaction diagram
-
-
-
-
?
uridine + phosphate
uracil + alpha-D-ribose 1-phosphate
show the reaction diagram
-
preferred substrate
-
-
?
uridine + phosphate
uracil + alpha-D-ribose 1-phosphate
show the reaction diagram
Q5XA29
preferred substrate
-
-
?
uridine + phosphate
uracil + alpha-D-ribose 1-phosphate
show the reaction diagram
-
by measuring the conversion of uracil to uridine or deoxyuridine a slight preference for ribose 1-phosphate over deoxyribose 1-phosphate is observed
-
r
uridine + phosphate
uracil + alpha-D-ribose 1-phosphate
show the reaction diagram
-
prefered substrate
-
-
?
uridine + phosphate
uracil + alpha-D-ribose 1-phosphate
show the reaction diagram
-
prefered substrate
-
-
?
uridine + phosphate
uracil + alpha-D-ribose 1-phosphate
show the reaction diagram
Q72HS4
prefered substrate
-
-
?
uridine + phosphate
uracil + alpha-D-ribose 1-phosphate
show the reaction diagram
Geobacillus stearothermophilus JTS 859
-
-
-
-
?
uridine + phosphate
uracil + alpha-D-ribose 1-phosphate
show the reaction diagram
Haemophilus influenzae TH 6-2
-
-
-
-
?
deoxyuridine + phosphate
uracil + 2'-deoxy-D-ribose 1-phosphate
show the reaction diagram
Q5XA29
-
-
-
?
additional information
?
-
-
no activity with deoxycytidine
-
-
-
additional information
?
-
-
reversible phosphorolysis of pyrimidines in the nucleotide synthesis salvage pathway
-
-
-
additional information
?
-
-
predictive profiling of 5-fluorouracil resistance in human cancer cells, overview
-
-
-
additional information
?
-
P39142
the function of His82, Arg169, and Lys188 is to stabilize the uridine in a high-energy conformation by means of electrostatic interactions. These residues are involved in catalysis. The function of Asp162 is likely to activate Lys188 for phosphorolytic catalysis through polarization effects
-
-
-
additional information
?
-
-
no substrate: cytidine, deoxycytidine, 2',2'-difluoro-2'-deoxycytidine, 2',2'-difluoro-2'-deoxyuridine, uracil arabinoside, thymine arabinoside, (E)-5-(2-bromovinyl)-deoxyuridine, azidothymidine, stavudine, 6-azauridine
-
-
-
additional information
?
-
Q5XA29
no substrate: cytidine, deoxycytidine, adenosine, inosine, or guanosine
-
-
-
NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
LITERATURE
(Substrate)
COMMENTARY
(Product)
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
pyrimidine nucleoside + phosphate
pyrimidine base + alpha-D-ribose 1-phosphate
show the reaction diagram
-
most pyrimidine nucleosides except for 4-amino-substituted species like deoxycytidine
or alpha-D-deoxyribose 1-phosphate
-
r
pyrimidine nucleoside + phosphate
pyrimidine base + alpha-D-ribose 1-phosphate
show the reaction diagram
Q5SHF9
most pyrimidine nucleosides except for 4-amino-substituted species like deoxycytidine
or alpha-D-deoxyribose 1-phosphate
-
r
additional information
?
-
-
reversible phosphorolysis of pyrimidines in the nucleotide synthesis salvage pathway
-
-
-
additional information
?
-
-
predictive profiling of 5-fluorouracil resistance in human cancer cells, overview
-
-
-
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
3,5-bis-(4-chlorobenzoyl)-alpha-D-ribose 1-phosphate
-
competitive inhibition with regard to inosine
5-benzylacyclouridine
-
-
5-chloro-6-(1-[2-iminopyrrolidinyl]methyl)uracil
-
-
uridine
-
above 15 mM, competitive inhibition of cleavage of 5-bromodeoxyuridine
7-deazaxanthine
-
-
additional information
-
inhibitory effects between pyrimidines and analogues
-
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.58
2'-deoxyuridine
-
-
0.25
4-thiothymidine
-
pH 7.5, 25C, thymidine phosphorylase activity
0.13
4-thiouridine
-
pH 7.5, 25C, uridine phosphorylase activity
0.1
5-bromodeoxyuridine
-
-
0.03
5-Bromouridine
-
-
0.169
5-fluorodeoxyuridine
-
pH 7.6, 37C
0.047
5-fluorouridine
-
pH 7.6, 37C
0.144
5-iododeoxyuridine
-
pH 7.6, 37C
0.069
5-iodouridine
-
pH 7.6, 37C
0.07
5-methyluridine
-
-
0.32
5-methyluridine
-
-
0.13
deoxyuridine
-
-
0.38
phosphate
-
reaction with uridine
0.388
phosphate
-
cosubstrate uridine, pH 7.6, 37C
0.797
phosphate
-
cosubstrate deoxythymidine, pH 7.6, 37C
0.43
thymidine
Q72HS4
pH 7.0, 60C
0.473
thymidine
-
pH 7.6, 37C
1.093
thymidine
-
pH 7.6, 37C
1.3
thymidine
-
pH 7.0, 60C
1.9
thymidine
-
-
0.092
uridine
-
pH 7.6, 37C
0.15
uridine
Q72HS4
pH 7.0, 60C
0.158
uridine
Q5XA29
wild-type, pH 7.5, temperature not specified in the publication
0.171
uridine
Q5XA29
mutant H169A, pH 7.5, temperature not specified in the publication
0.31
uridine
-
pH 7.6, 37C
1.38
uridine
-
-
2.3
uridine
-
pH 7.0, 60C
0.578
deoxyuridine
-
pH 7.6, 37C
additional information
additional information
-
kinetics
-
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
63
4-thiothymidine
-
pH 7.5, 25C, thymidine phosphorylase activity
49
4-thiouridine
-
pH 7.5, 25C, uridine phosphorylase activity
11.9
5-fluorodeoxyuridine
-
pH 7.6, 37C
6.1
5-fluorouridine
-
pH 7.6, 37C
9.5
5-iododeoxyuridine
-
pH 7.6, 37C
6.4
5-iodouridine
-
pH 7.6, 37C
24.6
deoxyuridine
-
pH 7.6, 37C
8.6
phosphate
-
cosubstrate uridine, pH 7.6, 37C
17.5
phosphate
-
cosubstrate deoxythymidine, pH 7.6, 37C
21.6
thymidine
-
pH 7.6, 37C
83
thymidine
-
pH 7.0, 60C
679
thymidine
Q72HS4
pH 7.0, 60C
1.7
uridine
Q5XA29
mutant H169A, pH 7.5, temperature not specified in the publication
8.5
uridine
-
pH 7.6, 37C
15
uridine
Q5XA29
wild-type, pH 7.5, temperature not specified in the publication
277
uridine
-
pH 7.0, 60C
277
uridine
Q72HS4
pH 7.0, 60C
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
70
5-fluorodeoxyuridine
-
pH 7.6, 37C
3977
130
5-fluorouridine
-
pH 7.6, 37C
2938
66
5-iododeoxyuridine
-
pH 7.6, 37C
8023
93
5-iodouridine
-
pH 7.6, 37C
7447
43
deoxyuridine
-
pH 7.6, 37C
1060
22
phosphate
-
cosubstrate deoxythymidine, pH 7.6, 37C; cosubstrate uridine, pH 7.6, 37C
16
46
thymidine
-
pH 7.6, 37C
166
92
uridine
-
pH 7.6, 37C
261
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.25
uridine
-
-
additional information
additional information
-
-
-
IC50 VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.113
3,5-bis-(4-chlorobenzoyl)-alpha-D-ribose 1-phosphate
-
substrate deoxythymidine, pH 7.6, 37C
0.128
3,5-bis-(4-chlorobenzoyl)-alpha-D-ribose 1-phosphate
-
substrate 5-fluorodeoxyuridine, pH 7.6, 37C
0.159
3,5-bis-(4-chlorobenzoyl)-alpha-D-ribose 1-phosphate
-
substrate 5-iododeoxyuridine pH 7.6, 37C
0.188
3,5-bis-(4-chlorobenzoyl)-alpha-D-ribose 1-phosphate
-
substrate uridine, pH 7.6, 37C
0.000003
5-benzylacyclouridine
-
substrate uridine, pH 7.6, 37C
0.000005
5-benzylacyclouridine
-
substrate deoxythymidine, pH 7.6, 37C
0.5
5-chloro-6-(1-[2-iminopyrrolidinyl]methyl)uracil
-
substrate deoxythymidine, pH 7.6, 37C; substrate uridine, pH 7.6, 37C
0.03
7-deazaxanthine
-
substrate deoxythymidine, pH 7.6, 37C
0.06
7-deazaxanthine
-
substrate uridine, pH 7.6, 37C
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
5.9
Q5XA29
substrate thymidine, pH 7.5, temperature not specified in the publication
8.1
Q5XA29
substrate uridine, pH 7.5, temperature not specified in the publication
11.2
Q5XA29
substrate deoxyuridine, pH 7.5, temperature not specified in the publication
45
-
purified recombinant enzyme
additional information
-
-
additional information
-
-
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
6.9
-
deoxynucleoside substrates
7.2
-
uridine, thymidine
7.4
-
ribonucleosides and uracil arabinoside
7.5
-
assay at
pH RANGE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
6 - 12
-
pH 6.0: about 75% of maximal activity, pH 12.0: about 80% of maximal activity
6.9 - 7.4
-
pH 6.9: optimum, pH 7.4: 75% of maximal activity
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
SOURCE
-
PyNPase level quantified by sandwich ELISA using monoclonal anti-human thimidine phosphorylase antibodies 104B and 232-2, no association between PyNPase level and recurrence, disease-free survival at year 2.2 better with low PyNPase levels and high PyNPase-to-dihydropyrimidine dihydrogenase ratio
Manually annotated by BRENDA team
-
PyNPase level quantified by sandwich ELISA using monoclonal anti-human thimidine phosphorylase antibodies 104B and 232-2
Manually annotated by BRENDA team
PDB
SCOP
CATH
ORGANISM
Staphylococcus aureus (strain COL)
Thermus thermophilus (strain HB8 / ATCC 27634 / DSM 579)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
45400
Q5SHF9
estimated according to amino acid composition
677372
78000
-
sedimentation velocity analysis
639637
85000
-
gel filtration
639641
86300
Q5SHF9
in the dimeric state of the purified protein
677372
92000
-
gel filtration
489698
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
dimer
-
2 * 54000, SDS-PAGE
dimer
-
2 * 46000, SDS-PAGE
dimer
Q5SHF9
revealed by dynamic light-scattering (DynaPro MS/X) of the purified protein, 20 mg protein/ml, 18C, 20 mM Tris-HCl pH 7.6, 200 mM sodium chloride, particle size distribution with a polydispersity of 18.7%
dimer
Geobacillus stearothermophilus JTS 859
-
2 * 54000, SDS-PAGE
-
dimer
Haemophilus influenzae TH 6-2
-
2 * 46000, SDS-PAGE
-
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
crystal structure of the enzyme with the substrate analog, pseudouridine, in its active site is solved to 2.1 A
-
hanging-drop vapor diffusion method, crystals of the protein-inhibitor complex with the substrate analog pseudouridine
-
in complex with the products, ribose 1-phosphate and uracil, at 1.8 A resolution. The biological unit is a hexamer with an alpha/beta monomeric fold. Residue His169 structurally aligns with Arg168 of the Escherichia coli uridine phosphorylase structure. A second active site residue, Lys162, is not present in previously determined uridine phosphorylase structures and interacts with O2 of uracil
Q5XA29
crystallized at 18C using the oil-microbatch method with PEG 4000 as a precipitant. A native data set is collected to 1.8 A resolution using synchrotron radiation. The crystal belongs to the monoclinic space group P2(1), with unit-cell parameters a = 58.83, b = 76.23, c = 103.86 A, beta = 91.3; crystals: monoclinic with dimensions of 0.13 * 0.13 * 0.07 mm, X-ray diffraction to 1.8 using synchrotron radiation at -173C, belong to space group P2(1), asymmetric unit contains dimer with local pseudo-twofold symmetry, unit-cell parameters: a: 58.83, b: 76.23, c: 103.86, beta: 91.3, oil-microbatch method: 1 week at 18C, drop: 0.5 microlitre precipitant solution (27.5% (w/v) PEG 4000 in 100 mM HEPES-NaOH pH 7.5, 10 mM CaCl2) + 0.5 microlitre protein solution (21.7 mg/ml in 20 mM Tris-HCl pH8, 200 NaCl)
Q5SHF9
pH STABILITY
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
5 - 9
-
stable
489698
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
60
-
stable below
489698
60
-
60 min, stable
639637
70
-
half-life: 25 min in 20 mM potassium phosphate, 15.1 h in 20 mM potassium phosphate and 1 mM 5-methyluridine
639641
70
-
half-life 1.6 h
723005
70
Q72HS4
half-life 1.6 h
723005
80
-
1 h, 20 mM potassium phosphate, complete inactivation
639641
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
repeated cycles of freezing and thawing inactivate the enzyme if phosphate is absent
-
the enzyme is quite labile during the course of purification, dithiothreitol and glycerol are required for stability
-
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-20C, in presence of phosphate, glycerol and dithiothreitol, stable for 6 months
-
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
recombinant enzymes
-
harvested with 5 mM 2-mercaptoethanol and 500 mM NaCl in Tris-HCl pH 8, extract desalted with HiPrep 2610, recombinant protein eluted with 0-300 mM NaCl from Super Q Toyopearl 650M column, eluted from Bio-Scale CHT-20-1 column with 10-100 mM NaCl in phosphate-NaOH pH 7, concentrated by ultracentrifugation with 10 kDa cut-off, eluted from HiLoad 16/60 Superdex 200 prep-grade column with 200 mM NaCl in Tris-HCl pH 8, concentrated by ultracentrifugation to 21.7 mg/ml, stored at -70 C
Q5SHF9
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expression in Escherichia coli
-
expression of enzymes with uridine and thymidine phosphorylase activities in strain BL21(DE3)
-
expression in Escherichia coli
-
expression in Escherichia coli
-
expression in Escherichia coli
-
expression in Escherichia coli
Q72HS4
pET-11a for expression in Escherichia coli BL21 (DE3)
Q5SHF9
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
H169A
Q5XA29
about 10% of wild-type activity
H169D
Q5XA29
about 1% of wild-type activity
H169N
Q5XA29
about 4% of wild-type activity
K162A
Q5XA29
complete loss of activity
V220D
Q5XA29
almost complete loss of activity
V220E
Q5XA29
less than 0.5% of wild-type activity
APPLICATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
synthesis
-
overexpression of enzyme in Escherichia coli, medium optimization and presence of uridine and 5-fluorouracil results in conversion of more than 90% of uridine to 5-fluorouridine by the cells
synthesis
Enterobacter aerogenes CMCC 45103
-
overexpression of enzyme in Escherichia coli, medium optimization and presence of uridine and 5-fluorouracil results in conversion of more than 90% of uridine to 5-fluorouridine by the cells
-
medicine
-
the cytostatic and antiviral activity of pyrimidine nucleoside analogues is markedly decreased by a Mycoplasma hyorhinis infection and the phosphorolytic activity of the mycoplasmas is responsible for this
synthesis
-
production of ribavirin, which is an antiviral drug