Information on EC 2.1.2.8 - deoxycytidylate 5-hydroxymethyltransferase

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The enzyme appears in viruses and cellular organisms

EC NUMBER
COMMENTARY hide
2.1.2.8
-
RECOMMENDED NAME
GeneOntology No.
deoxycytidylate 5-hydroxymethyltransferase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
5,10-methylenetetrahydrofolate + H2O + deoxycytidylate = tetrahydrofolate + 5-hydroxymethyldeoxycytidylate
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
hydroxymethyl group transfer
-
-
-
-
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
Pyrimidine metabolism
-
-
One carbon pool by folate
-
-
SYSTEMATIC NAME
IUBMB Comments
5,10-methylenetetrahydrofolate:deoxycytidylate 5-hydroxymethyltransferase
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CAS REGISTRY NUMBER
COMMENTARY hide
9012-68-4
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
bacteriophage T6r+
Escherichia coli strain B infected with bacteriophage T6r+
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
T-even phage
-
-
-
Manually annotated by BRENDA team
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
5,10-methylenetetrahydrofolate + H2O + deoxycytidylate
tetrahydrofolate + 5-hydroxymethyldeoxycytidylate
show the reaction diagram
additional information
?
-
NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
5,10-methylenetetrahydrofolate + H2O + deoxycytidylate
tetrahydrofolate + 5-hydroxymethyldeoxycytidylate
show the reaction diagram
additional information
?
-
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
5,10-methylenetetrahydrofolate
tetrahydrofolate
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
10-methyl-5,8-dideazatetrahydrofolate
-
competitive inhibitor with respect to tetrahydrofolate
-
5,5'-dithiobis(2-nitrobenzoic acid)
-
complete loss of activity, second-order kinetics, inactivation is reversed by dithiothreitol, inactivation involves the modification of one thiol group per mol of dimeric enzyme
5-deazatetrahydrofolate
5-Fluoro-2'-deoxycytidine 5'-monophosphate
-
competitive inhibitor with respect to dCMP, not mechanism-based inhibitor
5-fluorodeoxyuridylate
5-hydroxymethyl-dCMP
-
competitor of dCMP, inhibits formation of 5-hydroxymethyl-dCMP
aminopterin
-
competitive inhibitor with respect to tetrahydrofolate
dihydrofolate
-
competitive inhibitor with respect to tetrahydrofolate
methotrexate
-
competitive inhibitor with respect to tetrahydrofolate
N-ethylmaleimide
-
complete loss of activity, second-order kinetics, pH-dependent inactivation, dCMP protects against inactivation, dCMP plus either methotrexate or aminopterin greatly enhances protection, inactivation involves the modification of one thiol group per mol of dimeric enzyme
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
dithiothreitol
-
low specific activity samples can be activated by a 30 min room temperature incubation in 50 mM DTT
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.36
deoxycytidylate
-
-
0.04
tetrahydrofolate
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-
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0008 - 12.7
deoxycytidylate
additional information
additional information
-
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.01
10-methyl-5,8-dideazatetrahydrofolate
-
-
-
0.11
5-deazatetrahydrofolate
-
-
0.025
5-Fluoro-2'-deoxycytidine 5'-monophosphate
-
-
0.1
5-hydroxymethyl-dCMP
-
-
0.001
aminopterin
-
-
0.021
dihydrofolate
-
-
0.016
methotrexate
-
-
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
3.1
-
dCMP formation
4.6
-
5-hydroxymethyl-dCMP formation
additional information
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
additional information
-
enzyme is no membrane component or very loosely attached to the membrane
-
Manually annotated by BRENDA team
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
28400
-
calculation from the amino acid sequence
51400
-
gel filtration
57000
-
gel filtration
63000
-
sedimentation equilibrium method
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
homodimer
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
crystal structure in the presence of substrate dCMP at 1.6 A resolution
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
20
-
room temperature, 24 h, stable
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
4°C, up to 10 days, only minor loss of activity
bacteriophage T6r+
-
frozen, 20 mM KH2PO4, pH 6.8, 6 months
-
room temperature, 24 h, stable
-
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
1026fold purification; Escherichia coli B infected with
-
Escherichia coli B infected with
-
Escherichia coli infected with
bacteriophage T6r+
-
purification of His6-tagged enzyme
purification of wild-type enzyme and mutants E60Q and E60D
-
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
cloning and overexpression of g42 in Escherichia coli K38; nucleotide sequence of g42, the dCMP hydroxymethylase gene, encodes a protein of 246 amino acids
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gene 42 encoding enzyme is sequenced and expressed in Escherichia coli
nucleotide sequence of dCMP hydroxymethylase gene encoding a protein of 246 amino acids
nucleotide sequence of g42, the dCMP hydroxymethylase gene, encodes a protein of 246 amino acids
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overexpression in Escherichia coli BL21; structural gene 42 encodes enzyme
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structural gene 42 encodes enzyme
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
C148G
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mutant is unable to catalyze a detectable 18O exchange in either the absence or presence of the folate cofactor
D179A
-
mutant not inactivated by 5-fluoro-dUMP
D179S
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mutant not inactivated by 5-fluoro-dUMP
E60D
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mutant with decreased enzyme activity, impaired enzyme function in vitro and in vivo, uncoupling of early steps from later steps in catalysis
E60Q
-
mutant with decreased enzyme activity, impaired enzyme function in vitro and in vivo, uncoupling of early steps from later steps in catalysis
additional information