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Literature summary extracted from

  • Keefe, R.G.; Maley, G.F.; Saxl, R.L.; Maley, F.
    A T4-phage deoxycytidylate deaminase mutant that no longer requires deoxycytidine 5'-triphosphate for activation (2000), J. Biol. Chem., 275, 12598-12602.
    View publication on PubMed

Activating Compound

EC Number Activating Compound Comment Organism Structure
3.5.4.12 5-hydroxymethyl-dCTP natural positive allosteric effector, enzyme is much more effectively regulated by its natural effector, 5-hydroxymethyl-dCTP, than by dCTP, binding of 5-hydroxymethyl-dCTP is much more pH dependent than dCTP Tequatrovirus T4
3.5.4.12 5-hydroxymethyl-dCTP natural positive allosteric effector, enzyme is much more effectively regulated by its natural effector, 5-hydroxymethyl-dCTP, than by dCTP, binding of 5-hydroxymethyl-dCTP is much more pH dependent than dCTP Escherichia phage T2
3.5.4.12 5-hydroxymethyl-dCTP natural positive allosteric effector, enzyme is much more effectively regulated by its natural effector, 5-hydroxymethyl-dCTP, than by dCTP, binding of 5-hydroxymethyl-dCTP is much more pH dependent than dCTP Enterobacteria phage T6
3.5.4.12 5-hydroxymethyl-dCTP natural activator, required for activity Tequatrovirus T4
3.5.4.12 dCTP required for activity Tequatrovirus T4
3.5.4.12 dCTP positive allosteric effector Tequatrovirus T4
3.5.4.12 dCTP positive allosteric effector Escherichia phage T2
3.5.4.12 dCTP positive allosteric effector Enterobacteria phage T6
3.5.4.12 dCTP hexameric form of enzyme is activated by dCTP, while the dimer is not Tequatrovirus T4
3.5.4.12 dCTP wild-type, dCTP required for activity, no activity in absence of dCTP and Mg2+ Tequatrovirus T4
3.5.4.12 dCTP severalfold activation, activation requires presence of Mg2+ Tequatrovirus T4
3.5.4.12 dCTP severalfold activation, activation requires presence of Mg2+ Escherichia phage T2
3.5.4.12 dCTP severalfold activation, activation requires presence of Mg2+ Enterobacteria phage T6
3.5.4.12 dCTP mutants R115E and R115Q: dCTP not required. R115Q: slight, 30-40% activation by 0.02 mM dCTP Tequatrovirus T4
3.5.4.12 additional information the phage dCMP deaminase expression is increased upon infection of Escherichia coli Tequatrovirus T4

Cloned(Commentary)

EC Number Cloned (Comment) Organism
3.5.4.12 expression of wild-type and mutant enzymes in Escherichia coli Tequatrovirus T4

Protein Variants

EC Number Protein Variants Comment Organism
3.5.4.12 F112A molecular weight analysis using HPLC gel filtration in the presence of SDS and dCTP: wild-type is a hexamer, F112A varies from hexamer to dimer Tequatrovirus T4
3.5.4.12 F112A site-directed mutagenesis, the mutant enzyme shows reduced turnover and activity compared to the wild-type enzyme Tequatrovirus T4
3.5.4.12 R115E mutant enzyme active in the absence of dCTP and Mg2+, little if any activation by dCTP, possessing turnover number or kcat that is about 15% that of wild-type enzyme, specific activity about 40-50% of wild-type enzyme. Molecular weight analysis using HPLC gel filtration in the presence of SDS and dCTP: wild-type is a hexamer, R115E a dimer Tequatrovirus T4
3.5.4.12 R115E site-directed mutagenesis, the mutant enzyme shows reduced turnover and activity compared to the wild-type enzyme, but no longer requires deoxycytidine 5-triphosphate for activation in contrast to the wild-type enzyme Tequatrovirus T4
3.5.4.12 R115Q mutant enzyme active in the absence of dCTP and Mg2+, possessing turnover number or kcat that is about 15% that of wild-type enzyme, specific activity about 40-50% of wild-type enzyme. Molecular weight analysis using HPLC size gel filtration in the presence of SDS and dCTP: wild-type is a hexamer, R115Q varies from hexamer to dimer Tequatrovirus T4
3.5.4.12 R115Q site-directed mutagenesis, the mutant enzyme shows reduced turnover and activity compared to the wild-type enzyme, but no longer requires deoxycytidine 5-triphosphate for activation in contrast to the wild-type enzyme Tequatrovirus T4

General Stability

EC Number General Stability Organism
3.5.4.12 2-mercaptoethanol stabilizes Tequatrovirus T4
3.5.4.12 2-mercaptoethanol stabilizes Escherichia phage T2
3.5.4.12 EDTA: denaturation, EDTA removes the two resident Zn2+ atoms /subunit Tequatrovirus T4
3.5.4.12 guanidine-HCl, 6 M, denaturation, complete restoration of activity on removal of denaturant by dilution or dialysis Tequatrovirus T4

Inhibitors

EC Number Inhibitors Comment Organism Structure
3.5.4.12 dTTP allosteric inhibitor Enterobacteria phage T6
3.5.4.12 dTTP allosteric inhibitor Escherichia phage T2
3.5.4.12 dTTP allosteric inhibitor; wild-type, 0.1 mM dTTP: 20% inhibition, mutants R115E and R115Q: 0.1 mM dTTP: no inhibition. 0.3 mM dTTP: wild-type 90% inhibition, mutants R115E and R115Q: 30% inhibition Tequatrovirus T4
3.5.4.12 EDTA
-
 Tequatrovirus T4

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
3.5.4.12 additional information
-
 additional information
-
 Escherichia phage T2
3.5.4.12 additional information
-
 additional information kinetic analysis, data and characterization of wild-type and mutants R115E, R115Q and F112A Tequatrovirus T4
3.5.4.12 0.021
-
 dCMP wild-type, presence of dCTP and Mg2+ Tequatrovirus T4
3.5.4.12 0.021
-
 dCMP recombinant wild-type enzyme, pH 8.0, 30°C Tequatrovirus T4
3.5.4.12 0.127
-
 dCMP R115Q, absence of dCTP Tequatrovirus T4
3.5.4.12 0.127
-
 dCMP recombinant mutant R115Q, pH 8.0, 30°C Tequatrovirus T4
3.5.4.12 0.137
-
 dCMP R115E, absence of dCTP Tequatrovirus T4
3.5.4.12 0.137
-
 dCMP recombinant mutant R115E, pH 8.0, 30°C Tequatrovirus T4

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
3.5.4.12 Mg2+
-
 Tequatrovirus T4
3.5.4.12 Mg2+ required for activation by dCTP and for inhibition by dTTP Tequatrovirus T4
3.5.4.12 Mg2+ required for activation by dCTP and for inhibition by dTTP Escherichia phage T2
3.5.4.12 Mg2+ required for activation by dCTP and for inhibition by dTTP Enterobacteria phage T6
3.5.4.12 Zn2+ two resident Zn2+ atoms /subunit Tequatrovirus T4

Molecular Weight [Da]

EC Number Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
3.5.4.12 21200
-
 6 * 21200, wild-type, particularly in the presence of dCTP: hexamer, mutants R115Q and F112A varies from hexamers to dimers or something in between depending on protein concentration, HPLC gel filtration in the presence of SDS Tequatrovirus T4
3.5.4.12 21200
-
 2 * 21200, mutant R115E: dimer, mutants R115Q and F112A varies from hexamers to dimers or something in between depending on protein concentration, HPLC gel filtration in the presence of SDS Tequatrovirus T4
3.5.4.12 38200 113000 recombinant mutant F112A, gel filtration, dependent on protein concentration, in presence and absence of fdCTP Tequatrovirus T4
3.5.4.12 39800 118000 recombinant mutant R115Q, gel filtration, dependent on protein concentration, in presence and absence of fdCTP Tequatrovirus T4
3.5.4.12 41600 45400 recombinant mutant R115E, gel filtration, dependent on protein concentration, in presence and absence of fdCTP Tequatrovirus T4
3.5.4.12 44300
-
 R115E, HPLC gel filtration Tequatrovirus T4
3.5.4.12 110000 122000 recombinant wild-type enzyme, gel filtration, dependent on protein concentration, in presence and absence of fdCTP Tequatrovirus T4
3.5.4.12 111900
-
 F112A, HPLC gel filtration Tequatrovirus T4
3.5.4.12 124500
-
 R115Q, HPLC gel filtration Tequatrovirus T4
3.5.4.12 127000
-
 wild-type, HPLC gel filtration Tequatrovirus T4

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
3.5.4.12 dCMP + H2O Tequatrovirus T4
-
 dUMP + NH3
-
 ?
3.5.4.12 dCMP + H2O Escherichia phage T2
-
 dUMP + NH3
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
3.5.4.12 Enterobacteria phage T6
-
 T2-, T4-, and T6-phage induced enzyme in Escherichia coli
-
3.5.4.12 Escherichia phage T2
-
 T2-, T4-, and T6-phage induced enzyme in Escherichia coli
-
3.5.4.12 Tequatrovirus T4
-
-
-
3.5.4.12 Tequatrovirus T4
-
 T2-, T4-, and T6-phage induced enzyme in Escherichia coli
-
3.5.4.12 Tequatrovirus T4
-
 wild-type and R115E and R115Q, mutant enzymes that no longer requires dCTP for activation, and mutant F112A
-
3.5.4.12 Tequatrovirus T4
-
 T4-phage infected Escherichia coli
-

Purification (Commentary)

EC Number Purification (Comment) Organism
3.5.4.12 recombinant wild-type and mutant enzymes from Escherichia coli Tequatrovirus T4
3.5.4.12 wild-type and mutants R115E, R115Q and F112A Tequatrovirus T4

Reaction

EC Number Reaction Comment Organism Reaction ID
3.5.4.12 dCMP + H2O = dUMP + NH3 regulation, allosteric end-product regulation Tequatrovirus T4
a
3.5.4.12 dCMP + H2O = dUMP + NH3 regulation, allosteric end-product regulation Escherichia phage T2
a
3.5.4.12 dCMP + H2O = dUMP + NH3 regulation, allosteric end-product regulation Enterobacteria phage T6
a
3.5.4.12 dCMP + H2O = dUMP + NH3 highly regulated allosteric enzyme Tequatrovirus T4
a
3.5.4.12 dCMP + H2O = dUMP + NH3 highly regulated allosteric enzyme Escherichia phage T2
a
3.5.4.12 dCMP + H2O = dUMP + NH3 highly regulated allosteric enzyme Enterobacteria phage T6
a

Renatured (Commentary)

EC Number Renatured (Comment) Organism
3.5.4.12 after denaturation with EDTA, mutants R115E and R115Q restored 54% and 60% of original activities, wild-type enzyme only marginally restored Tequatrovirus T4
3.5.4.12 after treatment with guanidine-HCl, 6 M, complete reactivation on removal of denaturant by dilution or dialysis Tequatrovirus T4

Specific Activity [micromol/min/mg]

EC Number Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
3.5.4.12 additional information
-
 mutants R115E and R115Q possessing specific activity about 40-50% of wild-type enzyme Tequatrovirus T4
3.5.4.12 90 100 mutant R115E Tequatrovirus T4
3.5.4.12 121
-
 mutant R115Q Tequatrovirus T4
3.5.4.12 220
-
 wild-type Tequatrovirus T4

Storage Stability

EC Number Storage Stability Organism
3.5.4.12 -40°C, solid ammonium sulfate to 80% saturation, stable for longer periods of storage Tequatrovirus T4
3.5.4.12 0-4°C, 0.1 M 2-mercaptoethanol, stable for extended periods Tequatrovirus T4

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
3.5.4.12 dCMP + H2O
-
 Tequatrovirus T4 dUMP + NH3
-
 ?
3.5.4.12 dCMP + H2O
-
 Escherichia phage T2 dUMP + NH3
-
 ?
3.5.4.12 dCMP + H2O
-
 Enterobacteria phage T6 dUMP + NH3
-
 ?

Subunits

EC Number Subunits Comment Organism
3.5.4.12 dimer 2 * 21200, mutant R115E: dimer, mutants R115Q and F112A varies from hexamers to dimers or something in between depending on protein concentration, HPLC gel filtration in the presence of SDS Tequatrovirus T4
3.5.4.12 dimer mutants R115Q and R115E, mutant F112A changes between dimeric and hexameric forms Tequatrovirus T4
3.5.4.12 hexamer 6 identical subunits Tequatrovirus T4
3.5.4.12 hexamer 6 identical subunits Escherichia phage T2
3.5.4.12 hexamer 6 identical subunits Enterobacteria phage T6
3.5.4.12 hexamer 6 * 21200, wild-type, particularly in the presence of dCTP: hexamer, mutants R115Q and F112A varies from hexamers to dimers or something in between depending on protein concentration, HPLC gel filtration in the presence of SDS Tequatrovirus T4
3.5.4.12 hexamer wild-type enzyme, mutant F112A changes between dimeric and hexameric forms Tequatrovirus T4
3.5.4.12 More hexameric form of enzyme is activated by dCTP, while the dimer is not Tequatrovirus T4

Synonyms

EC Number Synonyms Comment Organism
3.5.4.12 dCMP deaminase
-
 Tequatrovirus T4
3.5.4.12 deoxycytidylate deaminase
-
 Tequatrovirus T4

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
3.5.4.12 30
-
 assay at Tequatrovirus T4

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
3.5.4.12 additional information
-
 additional information mutants R115E and R115Q possessing turnover number or kcat that is about 15% that of wild-type enzyme Tequatrovirus T4
3.5.4.12 69
-
 dCMP R115E Tequatrovirus T4
3.5.4.12 69
-
 dCMP mutant R115E, pH 8.0, 30°C Tequatrovirus T4
3.5.4.12 85
-
 dCMP R115Q dimer Tequatrovirus T4
3.5.4.12 85
-
 dCMP mutant R115Q, pH 8.0, 30°C Tequatrovirus T4
3.5.4.12 255
-
 dCMP R115Q hexamer Tequatrovirus T4
3.5.4.12 466
-
 dCMP wild-type Tequatrovirus T4
3.5.4.12 466
-
 dCMP wild-type enzyme, pH 8.0, 30°C Tequatrovirus T4

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
3.5.4.12 8
-
 assay at Tequatrovirus T4
3.5.4.12 8
-
 assay at Escherichia phage T2