BRENDA - Enzyme Database show
show all sequences of 2.1.1.56

Encephalitozoon cuniculi mRNA cap (guanine N-7) methyltransferase: methyl acceptor specificity, inhibition BY S-adenosylmethionine analogs, and structure-guided mutational analysis

Hausmann, S.; Zheng, S.; Fabrega, C.; Schneller, S.W.; Lima, C.D.; Shuman, S.; J. Biol. Chem. 280, 20404-20412 (2005)

Data extracted from this reference:

Cloned(Commentary)
Commentary
Organism
inserted into vector pET16b and transformed into Escherichia coli BL21(DE3), yeast strain YBS40 transformed with CEN TRP1 plasmids containing the wild-type and mutant alleles of ECM1
Encephalitozoon cuniculi
Crystallization (Commentary)
Crystallization
Organism
Ecm1-aza-S-adenosyl-L-methionine binary complex, by vapor diffusion, reveals that the inhibitor occupies the same site as S-adenosyl-L-methionine
Encephalitozoon cuniculi
Engineering
Amino acid exchange
Commentary
Organism
D70A
lethal, suppresses methyltransferase activity to less than 3% of the wild-type level, in vivo lethality correlates with loss of catalytic activity
Encephalitozoon cuniculi
D70E
growth at all temperatures
Encephalitozoon cuniculi
D70N
lethal
Encephalitozoon cuniculi
D78A
lethal, suppresses methyltransferase activity to less than 3% of the wild-type level, in vivo lethality correlates with loss of catalytic activity
Encephalitozoon cuniculi
D78E
growth at all temperatures except at 37°C
Encephalitozoon cuniculi
D78N
growth at all temperatures
Encephalitozoon cuniculi
D94A
suppresses methyltransferase activity to less than 3% of the wild-type level, in vivo lethality correlates with loss of catalytic activity
Encephalitozoon cuniculi
D94E
growth at all temperatures
Encephalitozoon cuniculi
D94N
lethal
Encephalitozoon cuniculi
E225A
growth at all temperatures except at 37°C
Encephalitozoon cuniculi
E225D
growth at all temperatures
Encephalitozoon cuniculi
E225Q
growth at all temperatures
Encephalitozoon cuniculi
F141A
suppresses methyltransferase activity to less than 3% of the wild-type level, in vivo lethality correlates with loss of catalytic activity
Encephalitozoon cuniculi
F214A
growth at all temperatures except at 37°C
Encephalitozoon cuniculi
F214L
growth at all temperatures, slow growth at 37°C
Encephalitozoon cuniculi
H144A
growth at all temperatures
Encephalitozoon cuniculi
K267A
growth at all temperatures
Encephalitozoon cuniculi
K54A
lethal, suppresses methyltransferase activity to less than 3% of the wild-type level, in vivo lethality correlates with loss of catalytic activity
Encephalitozoon cuniculi
K54Q
lethal
Encephalitozoon cuniculi
K54R
lethal
Encephalitozoon cuniculi
K75Q
slow growth at all temperatures
Encephalitozoon cuniculi
K75R
growth at all temperatures
Encephalitozoon cuniculi
K81Q
lethal
Encephalitozoon cuniculi
K81R
lethal
Encephalitozoon cuniculi
additional information
residues Lys-54, Asp-70, Asp-78, and Asp-94 that comprise the S-adenosyl-L-methionine-binding pocket are all essential for Ecm1 function in yeast, i.e. alanine substitutions at each of these positions are lethal, loss of function caused by the elimination of Phe-141 attests to the importance of its van der Waals interactions with the guanosine methyl acceptor seen in the Ecm1-GTP cocrystal
Encephalitozoon cuniculi
N51D
lethal
Encephalitozoon cuniculi
R106A
lethal
Encephalitozoon cuniculi
R106K
lethal
Encephalitozoon cuniculi
R106Q
lethal
Encephalitozoon cuniculi
R59A
growth at all temperatures
Encephalitozoon cuniculi
R84A
growth at all temperatures
Encephalitozoon cuniculi
Y124F
growth at all temperatures, slow growth at 37°C
Encephalitozoon cuniculi
Y145A
lethal
Encephalitozoon cuniculi
Y145F
growth at all temperatures
Encephalitozoon cuniculi
Y145L
growth at all temperatures
Encephalitozoon cuniculi
Y145S
lethal
Encephalitozoon cuniculi
Y212A
growth at all temperatures except at 37°C
Encephalitozoon cuniculi
Y212F
growth at all temperatures
Encephalitozoon cuniculi
Inhibitors
Inhibitors
Commentary
Organism
Structure
aza-S-adenosyl-L-methionine
-
Encephalitozoon cuniculi
carbocyclic aza-S-adenosyl-L-methionine
-
Encephalitozoon cuniculi
S-adenosyl-L-homocysteine
inhibits methylation of GTP in the presence of 0.005 mM S-adenosyl-L-methionine
Encephalitozoon cuniculi
sinefungin
-
Encephalitozoon cuniculi
KM Value [mM]
KM Value [mM]
KM Value Maximum [mM]
Substrate
Commentary
Organism
Structure
0.1
-
GpppA
-
Encephalitozoon cuniculi
1
-
GTP
-
Encephalitozoon cuniculi
2.4
-
GDP
-
Encephalitozoon cuniculi
Molecular Weight [Da]
Molecular Weight [Da]
Molecular Weight Maximum [Da]
Commentary
Organism
40000
-
1 * 40000, SDS-PAGE
Encephalitozoon cuniculi
Organism
Organism
Primary Accession No. (UniProt)
Commentary
Textmining
Encephalitozoon cuniculi
Q8SR66
-
-
Purification (Commentary)
Commentary
Organism
by nickel-affinity chromatography
Encephalitozoon cuniculi
Storage Stability
Storage Stability
Organism
-80 °C, Tris-HCl buffer, pH 7.5, 10% glycerol
Encephalitozoon cuniculi
Substrates and Products (Substrate)
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
additional information
does not methylate ATP, CTP, UTP, ITP, or m7GTP
674439
Encephalitozoon cuniculi
?
-
-
-
-
S-adenosyl-L-methionine + dGTP
-
674439
Encephalitozoon cuniculi
S-adenosyl-L-homocysteine + m7dGTP
-
-
-
-
S-adenosyl-L-methionine + GDP
-
674439
Encephalitozoon cuniculi
S-adenosyl-L-homocysteine + m7GDP
-
-
-
-
S-adenosyl-L-methionine + GpppA
-
674439
Encephalitozoon cuniculi
S-adenosyl-L-homocysteine + m7GpppA
-
-
-
-
S-adenosyl-L-methionine + GTP
-
674439
Encephalitozoon cuniculi
S-adenosyl-L-homocysteine + m7GTP
-
-
-
?
Subunits
Subunits
Commentary
Organism
monomer
1 * 40000, SDS-PAGE
Encephalitozoon cuniculi
pH Optimum
pH Optimum Minimum
pH Optimum Maximum
Commentary
Organism
7.5
-
-
Encephalitozoon cuniculi
pH Range
pH Minimum
pH Maximum
Commentary
Organism
6
7.5
methyl transfer by Ecm1 declines sharply below pH 6.0
Encephalitozoon cuniculi
Cofactor
Cofactor
Commentary
Organism
Structure
additional information
reaction is cofactor-independent
Encephalitozoon cuniculi
IC50 Value
IC50 Value
IC50 Value Maximum
Commentary
Organism
Inhibitor
Structure
0.0015
-
-
Encephalitozoon cuniculi
sinefungin
0.004
-
-
Encephalitozoon cuniculi
S-adenosyl-L-homocysteine
0.035
-
-
Encephalitozoon cuniculi
carbocyclic aza-S-adenosyl-L-methionine
0.1
-
-
Encephalitozoon cuniculi
aza-S-adenosyl-L-methionine
Cloned(Commentary) (protein specific)
Commentary
Organism
inserted into vector pET16b and transformed into Escherichia coli BL21(DE3), yeast strain YBS40 transformed with CEN TRP1 plasmids containing the wild-type and mutant alleles of ECM1
Encephalitozoon cuniculi
Cofactor (protein specific)
Cofactor
Commentary
Organism
Structure
additional information
reaction is cofactor-independent
Encephalitozoon cuniculi
Crystallization (Commentary) (protein specific)
Crystallization
Organism
Ecm1-aza-S-adenosyl-L-methionine binary complex, by vapor diffusion, reveals that the inhibitor occupies the same site as S-adenosyl-L-methionine
Encephalitozoon cuniculi
Engineering (protein specific)
Amino acid exchange
Commentary
Organism
D70A
lethal, suppresses methyltransferase activity to less than 3% of the wild-type level, in vivo lethality correlates with loss of catalytic activity
Encephalitozoon cuniculi
D70E
growth at all temperatures
Encephalitozoon cuniculi
D70N
lethal
Encephalitozoon cuniculi
D78A
lethal, suppresses methyltransferase activity to less than 3% of the wild-type level, in vivo lethality correlates with loss of catalytic activity
Encephalitozoon cuniculi
D78E
growth at all temperatures except at 37°C
Encephalitozoon cuniculi
D78N
growth at all temperatures
Encephalitozoon cuniculi
D94A
suppresses methyltransferase activity to less than 3% of the wild-type level, in vivo lethality correlates with loss of catalytic activity
Encephalitozoon cuniculi
D94E
growth at all temperatures
Encephalitozoon cuniculi
D94N
lethal
Encephalitozoon cuniculi
E225A
growth at all temperatures except at 37°C
Encephalitozoon cuniculi
E225D
growth at all temperatures
Encephalitozoon cuniculi
E225Q
growth at all temperatures
Encephalitozoon cuniculi
F141A
suppresses methyltransferase activity to less than 3% of the wild-type level, in vivo lethality correlates with loss of catalytic activity
Encephalitozoon cuniculi
F214A
growth at all temperatures except at 37°C
Encephalitozoon cuniculi
F214L
growth at all temperatures, slow growth at 37°C
Encephalitozoon cuniculi
H144A
growth at all temperatures
Encephalitozoon cuniculi
K267A
growth at all temperatures
Encephalitozoon cuniculi
K54A
lethal, suppresses methyltransferase activity to less than 3% of the wild-type level, in vivo lethality correlates with loss of catalytic activity
Encephalitozoon cuniculi
K54Q
lethal
Encephalitozoon cuniculi
K54R
lethal
Encephalitozoon cuniculi
K75Q
slow growth at all temperatures
Encephalitozoon cuniculi
K75R
growth at all temperatures
Encephalitozoon cuniculi
K81Q
lethal
Encephalitozoon cuniculi
K81R
lethal
Encephalitozoon cuniculi
additional information
residues Lys-54, Asp-70, Asp-78, and Asp-94 that comprise the S-adenosyl-L-methionine-binding pocket are all essential for Ecm1 function in yeast, i.e. alanine substitutions at each of these positions are lethal, loss of function caused by the elimination of Phe-141 attests to the importance of its van der Waals interactions with the guanosine methyl acceptor seen in the Ecm1-GTP cocrystal
Encephalitozoon cuniculi
N51D
lethal
Encephalitozoon cuniculi
R106A
lethal
Encephalitozoon cuniculi
R106K
lethal
Encephalitozoon cuniculi
R106Q
lethal
Encephalitozoon cuniculi
R59A
growth at all temperatures
Encephalitozoon cuniculi
R84A
growth at all temperatures
Encephalitozoon cuniculi
Y124F
growth at all temperatures, slow growth at 37°C
Encephalitozoon cuniculi
Y145A
lethal
Encephalitozoon cuniculi
Y145F
growth at all temperatures
Encephalitozoon cuniculi
Y145L
growth at all temperatures
Encephalitozoon cuniculi
Y145S
lethal
Encephalitozoon cuniculi
Y212A
growth at all temperatures except at 37°C
Encephalitozoon cuniculi
Y212F
growth at all temperatures
Encephalitozoon cuniculi
IC50 Value (protein specific)
IC50 Value
IC50 Value Maximum
Commentary
Organism
Inhibitor
Structure
0.0015
-
-
Encephalitozoon cuniculi
sinefungin
0.004
-
-
Encephalitozoon cuniculi
S-adenosyl-L-homocysteine
0.035
-
-
Encephalitozoon cuniculi
carbocyclic aza-S-adenosyl-L-methionine
0.1
-
-
Encephalitozoon cuniculi
aza-S-adenosyl-L-methionine
Inhibitors (protein specific)
Inhibitors
Commentary
Organism
Structure
aza-S-adenosyl-L-methionine
-
Encephalitozoon cuniculi
carbocyclic aza-S-adenosyl-L-methionine
-
Encephalitozoon cuniculi
S-adenosyl-L-homocysteine
inhibits methylation of GTP in the presence of 0.005 mM S-adenosyl-L-methionine
Encephalitozoon cuniculi
sinefungin
-
Encephalitozoon cuniculi
KM Value [mM] (protein specific)
KM Value [mM]
KM Value Maximum [mM]
Substrate
Commentary
Organism
Structure
0.1
-
GpppA
-
Encephalitozoon cuniculi
1
-
GTP
-
Encephalitozoon cuniculi
2.4
-
GDP
-
Encephalitozoon cuniculi
Molecular Weight [Da] (protein specific)
Molecular Weight [Da]
Molecular Weight Maximum [Da]
Commentary
Organism
40000
-
1 * 40000, SDS-PAGE
Encephalitozoon cuniculi
Purification (Commentary) (protein specific)
Commentary
Organism
by nickel-affinity chromatography
Encephalitozoon cuniculi
Storage Stability (protein specific)
Storage Stability
Organism
-80 °C, Tris-HCl buffer, pH 7.5, 10% glycerol
Encephalitozoon cuniculi
Substrates and Products (Substrate) (protein specific)
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
additional information
does not methylate ATP, CTP, UTP, ITP, or m7GTP
674439
Encephalitozoon cuniculi
?
-
-
-
-
S-adenosyl-L-methionine + dGTP
-
674439
Encephalitozoon cuniculi
S-adenosyl-L-homocysteine + m7dGTP
-
-
-
-
S-adenosyl-L-methionine + GDP
-
674439
Encephalitozoon cuniculi
S-adenosyl-L-homocysteine + m7GDP
-
-
-
-
S-adenosyl-L-methionine + GpppA
-
674439
Encephalitozoon cuniculi
S-adenosyl-L-homocysteine + m7GpppA
-
-
-
-
S-adenosyl-L-methionine + GTP
-
674439
Encephalitozoon cuniculi
S-adenosyl-L-homocysteine + m7GTP
-
-
-
?
Subunits (protein specific)
Subunits
Commentary
Organism
monomer
1 * 40000, SDS-PAGE
Encephalitozoon cuniculi
pH Optimum (protein specific)
pH Optimum Minimum
pH Optimum Maximum
Commentary
Organism
7.5
-
-
Encephalitozoon cuniculi
pH Range (protein specific)
pH Minimum
pH Maximum
Commentary
Organism
6
7.5
methyl transfer by Ecm1 declines sharply below pH 6.0
Encephalitozoon cuniculi
Other publictions for EC 2.1.1.56
No.
1st author
Pub Med
title
organims
journal
volume
pages
year
Activating Compound
Application
Cloned(Commentary)
Crystallization (Commentary)
Engineering
General Stability
Inhibitors
KM Value [mM]
Localization
Metals/Ions
Molecular Weight [Da]
Natural Substrates/ Products (Substrates)
Organic Solvent Stability
Organism
Oxidation Stability
Posttranslational Modification
Purification (Commentary)
Reaction
Renatured (Commentary)
Source Tissue
Specific Activity [micromol/min/mg]
Storage Stability
Substrates and Products (Substrate)
Subunits
Temperature Optimum [°C]
Temperature Range [°C]
Temperature Stability [°C]
Turnover Number [1/s]
pH Optimum
pH Range
pH Stability
Cofactor
Ki Value [mM]
pI Value
IC50 Value
Activating Compound (protein specific)
Application (protein specific)
Cloned(Commentary) (protein specific)
Cofactor (protein specific)
Crystallization (Commentary) (protein specific)
Engineering (protein specific)
General Stability (protein specific)
IC50 Value (protein specific)
Inhibitors (protein specific)
Ki Value [mM] (protein specific)
KM Value [mM] (protein specific)
Localization (protein specific)
Metals/Ions (protein specific)
Molecular Weight [Da] (protein specific)
Natural Substrates/ Products (Substrates) (protein specific)
Organic Solvent Stability (protein specific)
Oxidation Stability (protein specific)
Posttranslational Modification (protein specific)
Purification (Commentary) (protein specific)
Renatured (Commentary) (protein specific)
Source Tissue (protein specific)
Specific Activity [micromol/min/mg] (protein specific)
Storage Stability (protein specific)
Substrates and Products (Substrate) (protein specific)
Subunits (protein specific)
Temperature Optimum [°C] (protein specific)
Temperature Range [°C] (protein specific)
Temperature Stability [°C] (protein specific)
Turnover Number [1/s] (protein specific)
pH Optimum (protein specific)
pH Range (protein specific)
pH Stability (protein specific)
pI Value (protein specific)
Expression
General Information
General Information (protein specific)
Expression (protein specific)
KCat/KM [mM/s]
KCat/KM [mM/s] (protein specific)
733122
Sun
Yeast-based assays for the hig ...
SARS coronavirus
Antiviral Res.
104
156-164
2014
-
1
-
-
-
-
1
-
-
-
-
-
-
7
-
-
-
-
-
-
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1
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1
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-
-
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-
-
-
-
734586
Chen
Structure-function analysis of ...
SARS coronavirus
J. Virol.
87
6296-6305
2013
-
-
1
1
27
-
-
-
-
-
-
-
-
4
-
-
-
-
-
-
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1
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1
27
-
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-
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-
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-
-
-
-
-
-
-
-
-
-
-
735335
Jin
Characterization of the guanin ...
SARS coronavirus
Virus Res.
176
45-52
2013
-
-
-
-
18
-
-
-
-
-
-
-
-
5
-
-
-
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-
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6
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18
-
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6
-
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-
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-
-
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-
-
-
-
735240
Thillier
Synthesis of 5' cap-0 and cap- ...
Homo sapiens
RNA
18
856-868
2012
-
1
1
-
-
-
-
-
-
-
-
-
-
1
-
-
-
-
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-
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1
1
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-
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-
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-
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-
-
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-
-
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-
-
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-
-
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-
-
-
-
-
-
-
-
-
-
-
704920
Selisko
Biochemical characterization o ...
Dengue virus
J. Gen. Virol.
91
112-121
2010
-
-
-
-
-
-
1
-
-
-
-
1
-
3
-
-
-
-
-
-
-
-
1
-
-
-
-
-
-
-
-
-
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-
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1
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1
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1
-
-
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-
-
-
-
-
-
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-
-
-
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-
721099
Chung
Higher catalytic efficiency of ...
Dengue virus
Virology
402
52-60
2010
-
-
-
-
-
-
2
2
-
-
-
-
-
1
-
-
-
-
-
-
-
-
1
-
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-
-
2
-
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-
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2
-
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-
-
-
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-
2
2
-
2
-
-
-
-
-
-
-
-
-
-
-
-
1
-
-
-
-
2
-
-
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-
-
-
-
-
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-
721100
Dong
Biochemical and genetic charac ...
Dengue virus
Virology
405
568-578
2010
-
-
1
-
4
-
-
-
-
-
1
-
-
1
-
-
-
-
-
-
-
-
2
-
-
-
-
-
1
-
-
-
-
-
-
-
-
1
-
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4
-
-
-
-
-
-
-
1
-
-
-
-
-
-
-
-
-
2
-
-
-
-
-
1
-
-
-
-
1
1
-
-
-
721101
Zhang
Identification of aromatic ami ...
vesicular stomatitis virus
Virology
408
241-252
2010
-
-
-
-
16
-
-
-
-
-
-
-
-
1
-
-
-
-
-
-
-
-
1
-
-
-
-
-
-
-
-
1
-
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-
-
-
-
1
-
16
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
1
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
701751
Milani
Flaviviral methyltransferase/R ...
Dengue virus, Wesselsbron virus
Antiviral Res.
83
28-34
2009
-
-
-
-
-
-
4
-
-
-
-
-
-
4
-
-
2
-
-
-
-
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2
-
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-
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2
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-
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2
4
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-
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2
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-
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2
-
-
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-
-
-
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-
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-
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-
706020
Cowling
Enhanced mRNA cap methylation ...
Homo sapiens
Oncogene
29
930-936
2009
-
-
-
-
-
-
-
-
-
-
-
-
-
1
-
-
-
-
-
2
-
-
1
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-
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-
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-
-
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-
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1
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485450
Pugh
Effects of S-adenosylhomocyste ...
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5 Cap methylation of homologou ...
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Purification of mRNA guanylylt ...
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Pugh
Inhibition of Newcastle diseas ...
Avian avulavirus 1
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Ensinger
Modification of the 5 terminus ...
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-
-
-
-
-
2
3
1
-
-
1
-
-
1
-
-
1
1
2
8
-
-
-
-
-
1
-
-
1
-
-
-
-
-
-
1
-
-
-
-
-
-
-
2
3
1
-
-
-
-
1
-
1
1
2
8
-
-
-
-
-
1
-
-
-
-
-
-
-
-
-
485458
Martin
mRNA guanylyltransferase and m ...
Vaccinia virus, Vaccinia virus WR
J. Biol. Chem.
251
7313-7321
1976
-
-
-
-
-
-
-
-
1
1
-
-
-
11
-
-
1
-
-
-
-
-
13
-
1
-
-
-
1
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
1
1
-
-
-
-
-
1
-
-
-
-
13
-
1
-
-
-
1
-
-
-
-
-
-
-
-
-
485452
Martin
Purification of mRNA guanylylt ...
Vaccinia virus, Vaccinia virus WR
J. Biol. Chem.
250
9322-9329
1975
-
-
-
-
-
-
2
-
1
-
2
2
-
11
-
-
1
-
-
-
-
1
4
1
1
-
-
-
1
-
-
-
3
-
-
-
-
-
-
-
-
-
-
2
3
-
1
-
2
2
-
-
-
1
-
-
-
1
4
1
1
-
-
-
1
-
-
-
-
-
-
-
-
-
485453
Martin
Modification of RNA by mRNA gu ...
Vaccinia virus
J. Biol. Chem.
250
9330-9335
1975
-
-
-
-
-
-
6
-
1
1
-
-
-
1
-
-
-
-
-
-
-
-
4
-
1
-
-
-
-
-
-
1
-
-
-
-
-
-
1
-
-
-
-
6
-
-
1
1
-
-
-
-
-
-
-
-
-
-
4
-
1
-
-
-
-
-
-
-
-
-
-
-
-
-
485454
Ensinger
Modification of the 5-terminus ...
Vaccinia virus
Proc. Natl. Acad. Sci. USA
72
2525-2529
1975
-
-
-
-
-
-
2
-
1
2
-
-
-
1
-
-
-
-
-
-
-
-
4
-
1
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
2
-
-
1
2
-
-
-
-
-
-
-
-
-
-
4
-
1
-
-
-
-
-
-
-
-
-
-
-
-
-