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Information on EC 3.6.1.74 - mRNA 5'-phosphatase
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3.6.1.74 mRNA 5'-phosphataseIUBMB Comments
The enzyme, found in eukaryotes and some plus strand RNA viruses (e.g. alphavirus), is involved in mRNA capping. Unlike the eukaryotic enzyme, the viral enzyme requires a purine in the first position of the mRNA. The human enzyme is a multi domain protein that also has the activity of EC 2.7.7.50, mRNA guanylyltransferase.
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Word Map
- 3.6.1.74
-
guanylyltransferase
- proteasome
- integrins
- triphosphate
- gtp
-
vaccinia
- ntpase
-
nucleotidyl
-
flavivirus
- f1-atpase
-
inhibin
-
kappa-b
-
gamma-phosphate
-
coatomer
- nsp2
-
reovirus
- gtases
-
replicase
-
inhba
-
nonsegmented
- spt5
The enzyme appears in viruses and cellular organisms
Reaction Schemes
a 5'-triphospho-[mRNA]
+
=
+
Synonyms
subunit beta, mrna capping, rna triphosphatase, rna 5'-triphosphatase, mrna capping apparatus, cacet1p, hce1b, hcap1a, hce1a, mrna 5'-capping enzyme, 
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SYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
5'-polynucleotidase
-
-
-
-
CaCET1
CaCet1p
capping enzyme RNA 5'-triphosphatase-like 1
CET1
Cet1 RNA triphosphatase
Ctl1
mRNA 5'-capping enzyme triphosphatase 1
mRNA 5'-triphosphatase
mRNA capping
mRNA capping apparatus
mRNA capping enzyme
mRNA-capping enzyme
nonstructural protein 2
nsP2
polynucleotide 5'-phosphohydrolase
-
-
-
-
RNA 5'-triphosphatase
RNA triphosphatase
RNGTT
subunit beta
Tpase
CET1
-
-
-
-
RNGTT
-
-
-
-
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REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT
LITERATURE
a 5'-triphospho-[mRNA] + H2O = a 5'-diphospho-[mRNA] + phosphate
-
-
-
-
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PATHWAY SOURCE
PATHWAYS
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-
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SYSTEMATIC NAME
IUBMB Comments
5'-triphospho-mRNA 5'-phosphohydrolase
The enzyme, found in eukaryotes and some plus strand RNA viruses (e.g. alphavirus), is involved in mRNA capping. Unlike the eukaryotic enzyme, the viral enzyme requires a purine in the first position of the mRNA. The human enzyme is a multi domain protein that also has the activity of EC 2.7.7.50, mRNA guanylyltransferase.
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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
a 5'-triphospho-[mRNA] + H2O
a 5'-diphospho-[mRNA] + phosphate
-
-
-
?
a 5'-triphospho-[mRNA] + H2O
a 5'-diphospho-[mRNA] + phosphate
[gamma-32P]ATP-terminated RNA
-
-
?
a 5'-triphospho-[mRNA] + H2O
a 5'-diphospho-[mRNA] + phosphate
-
-
-
?
a 5'-triphospho-[mRNA] + H2O
a 5'-diphospho-[mRNA] + phosphate
[gamma-32P]ATP-terminated RNA
-
-
?
a 5'-triphospho-[mRNA] + H2O
a 5'-diphospho-[mRNA] + phosphate
-
-
-
?
a 5'-triphospho-[mRNA] + H2O
a 5'-diphospho-[mRNA] + phosphate
-
-
-
?
a 5'-triphospho-[mRNA] + H2O
a 5'-diphospho-[mRNA] + phosphate
[gamma-32P]ATP-terminated RNA
-
-
?
a 5'-triphospho-[mRNA] + H2O
a 5'-diphospho-[mRNA] + phosphate
-
[gamma-32P]ATP-terminated RNA
-
-
?
a 5'-triphospho-[mRNA] + H2O
a 5'-diphospho-[mRNA] + phosphate
-
-
-
?
a 5'-triphospho-[mRNA] + H2O
a 5'-diphospho-[mRNA] + phosphate
-
-
-
-
?
a 5'-triphospho-[mRNA] + H2O
a 5'-diphospho-[mRNA] + phosphate
-
-
-
?
a 5'-triphospho-[mRNA] + H2O
a 5'-diphospho-[mRNA] + phosphate
-
-
-
?
a 5'-triphospho-[mRNA] + H2O
a 5'-diphospho-[mRNA] + phosphate
-
-
-
?
a 5'-triphospho-[mRNA] + H2O
a 5'-diphospho-[mRNA] + phosphate
-
-
-
?
a 5'-triphospho-[mRNA] + H2O
a 5'-diphospho-[mRNA] + phosphate
-
-
-
?
pppA-terminated poly(A) + H2O
ppA-terminated poly(A) + phosphate
-
-
-
?
pppA-terminated poly(A) + H2O
ppA-terminated poly(A) + phosphate
-
-
-
-
?
pppA-terminated poly(A) + H2O
ppA-terminated poly(A) + phosphate
-
-
-
?
pppApCpC + H2O
ppApCpC + phosphate
[gamma32P]ATP-terminated trimeric RNA or [alpha32P]ATP-terminated trimeric RNA
-
-
?
pppApCpC + H2O
ppApCpC + phosphate
[gamma32P]ATP-terminated trimeric RNA or [alpha32P]ATP-terminated trimeric RNA
-
-
?
pppG-terminated mRNA + H2O
ppG-terminated mRNA + phosphate
lower activity
-
-
?
pppG-terminated mRNA + H2O
ppG-terminated mRNA + phosphate
lower activity
-
-
?
additional information
?
-
cap formation by the eukaryotic GTase (EC 2.7.7.50) requires an RNA with a 5'-diphosphate end as the substrate
-
-
-
additional information
?
-
-
cap formation by the eukaryotic GTase (EC 2.7.7.50) requires an RNA with a 5'-diphosphate end as the substrate
-
-
-
additional information
?
-
cap formation by the eukaryotic GTase (EC 2.7.7.50) requires an RNA with a 5'-diphosphate end as the substrate
-
-
-
additional information
?
-
cap formation by the eukaryotic GTase (EC 2.7.7.50) requires an RNA with a 5'-diphosphate end as the substrate. Inability of the human capping enzyme to bind to the yeast GTase
-
-
-
additional information
?
-
-
cap formation by the eukaryotic GTase (EC 2.7.7.50) requires an RNA with a 5'-diphosphate end as the substrate. Inability of the human capping enzyme to bind to the yeast GTase
-
-
-
additional information
?
-
-
the purified recombinant hCAP1a gene product, hCAP1a, exhibits both mRNA 5'-triphosphatase and mRNA guanylyltransferase activities. The N-terminal 213 amino acid fragment containing a tyrosine-specific protein phosphatase motif catalyzes the RNA 5'-triphosphatase activity, and the C-terminal 369 amino acid fragment exhibits the mRNA guanylyltransferase activity. The recombinant protein hCAP1a releases [32P]Pi from [gamma-32P]triphosphate-terminated to RNA, but it is less active with [gamma-32P]GTP, indicating that it recognize the RNA chain
-
-
-
additional information
?
-
the purified recombinant hCAP1a gene product, hCAP1a, exhibits both mRNA 5'-triphosphatase and mRNA guanylyltransferase activities. The N-terminal 213 amino acid fragment containing a tyrosine-specific protein phosphatase motif catalyzes the RNA 5'-triphosphatase activity, and the C-terminal 369 amino acid fragment exhibits the mRNA guanylyltransferase activity. The recombinant protein hCAP1a releases [32P]Pi from [gamma-32P]triphosphate-terminated to RNA, but it is less active with [gamma-32P]GTP, indicating that it recognize the RNA chain
-
-
-
additional information
?
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-
the purified recombinant hCAP1b gene product, hCAP1b, shows RNA 5'-triphosphatase activity, but neither enzyme-GMP covalent complex formation nor cap structure formation is detected. The recombinant protein hCAP1b releases [32P]Pi from [gamma-32P]triphosphate-terminated to RNA, but it is less active with [gamma-32P]GTP, indicating that it recognize the RNA chain
-
-
-
additional information
?
-
the purified recombinant hCAP1b gene product, hCAP1b, shows RNA 5'-triphosphatase activity, but neither enzyme-GMP covalent complex formation nor cap structure formation is detected. The recombinant protein hCAP1b releases [32P]Pi from [gamma-32P]triphosphate-terminated to RNA, but it is less active with [gamma-32P]GTP, indicating that it recognize the RNA chain
-
-
-
additional information
?
-
beta-subunit enzyme Cet1 exhibits an RNA 5'-triphosphatase activity which specifically removes the gamma-phosphate from the triphosphate-terminated RNA substrate, but not from nucleoside triphosphates. Interaction between the Cet1 and the Ceg1 subunits is also studied by the West-Western procedure using recombinant Ceg1-[32P]GMP as probe. The recombinant Ceg1 is incubated with [alpha-32P]GTP to form a Ceg1-[32P]pG intermediate, method development and evaluation
-
-
-
additional information
?
-
-
beta-subunit enzyme Cet1 exhibits an RNA 5'-triphosphatase activity which specifically removes the gamma-phosphate from the triphosphate-terminated RNA substrate, but not from nucleoside triphosphates. Interaction between the Cet1 and the Ceg1 subunits is also studied by the West-Western procedure using recombinant Ceg1-[32P]GMP as probe. The recombinant Ceg1 is incubated with [alpha-32P]GTP to form a Ceg1-[32P]pG intermediate, method development and evaluation
-
-
-
additional information
?
-
enzyme Cet1 releases the gamma-phosphate from the terminal ATP of RNA
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-
-
additional information
?
-
enzyme Cet1 releases the gamma-phosphate from the terminal ATP of RNA
-
-
-
additional information
?
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enzyme Cet1 releases the gamma-phosphate from the terminal ATP of RNA
-
-
-
additional information
?
-
recombinant CTL1 releases the gamma-phosphate from the 5'-end of RNA to produce a diphosphate terminus. The enzyme is specific for polynucleotide RNA in the presence of magnesium, but becomes specific for nucleotide triphosphates in the presence of manganese. No activity with [gamma32P]ATP-terminated pppA
-
-
-
additional information
?
-
recombinant CTL1 releases the gamma-phosphate from the 5'-end of RNA to produce a diphosphate terminus. The enzyme is specific for polynucleotide RNA in the presence of magnesium, but becomes specific for nucleotide triphosphates in the presence of manganese. No activity with [gamma32P]ATP-terminated pppA
-
-
-
additional information
?
-
-
recombinant CTL1 releases the gamma-phosphate from the 5'-end of RNA to produce a diphosphate terminus. The enzyme is specific for polynucleotide RNA in the presence of magnesium, but becomes specific for nucleotide triphosphates in the presence of manganese. No activity with [gamma32P]ATP-terminated pppA
-
-
-
additional information
?
-
RNA triphosphatase activity is assayed by liberation of 32Pi from gamma-32P-labeled triphosphate-terminated poly(A) and quantitatively analyzed on polyethyleneimine-cellulose plates via thin-layer chromatography
-
-
-
additional information
?
-
-
RNA triphosphatase activity is assayed by liberation of 32Pi from gamma-32P-labeled triphosphate-terminated poly(A) and quantitatively analyzed on polyethyleneimine-cellulose plates via thin-layer chromatography
-
-
-
additional information
?
-
the enzyme is specific for the gamma-phosphoryl group at the 5'-terminus of RNA, it does not hydrolyze ATP. It can hydrolyze the gamma-phosphoryl group of pppGp, but the RNA substrates with longer chain length are preferred
-
-
-
additional information
?
-
-
the enzyme is specific for the gamma-phosphoryl group at the 5'-terminus of RNA, it does not hydrolyze ATP. It can hydrolyze the gamma-phosphoryl group of pppGp, but the RNA substrates with longer chain length are preferred
-
-
-
additional information
?
-
beta-subunit enzyme Cet1 exhibits an RNA 5'-triphosphatase activity which specifically removes the gamma-phosphate from the triphosphate-terminated RNA substrate, but not from nucleoside triphosphates. Interaction between the Cet1 and the Ceg1 subunits is also studied by the West-Western procedure using recombinant Ceg1-[32P]GMP as probe. The recombinant Ceg1 is incubated with [alpha-32P]GTP to form a Ceg1-[32P]pG intermediate, method development and evaluation
-
-
-
additional information
?
-
the enzyme is specific for the gamma-phosphoryl group at the 5'-terminus of RNA, it does not hydrolyze ATP. It can hydrolyze the gamma-phosphoryl group of pppGp, but the RNA substrates with longer chain length are preferred
-
-
-
additional information
?
-
RNA triphosphatase activity is assayed by liberation of 32Pi from gamma-32P-labeled triphosphate-terminated poly(A) and quantitatively analyzed on polyethyleneimine-cellulose plates via thin-layer chromatography
-
-
-
additional information
?
-
enzyme Cet1 releases the gamma-phosphate from the terminal ATP of RNA
-
-
-
additional information
?
-
enzyme Cet1 releases the gamma-phosphate from the terminal ATP of RNA
-
-
-
additional information
?
-
recombinant CTL1 releases the gamma-phosphate from the 5'-end of RNA to produce a diphosphate terminus. The enzyme is specific for polynucleotide RNA in the presence of magnesium, but becomes specific for nucleotide triphosphates in the presence of manganese. No activity with [gamma32P]ATP-terminated pppA
-
-
-
additional information
?
-
recombinant CTL1 releases the gamma-phosphate from the 5'-end of RNA to produce a diphosphate terminus. The enzyme is specific for polynucleotide RNA in the presence of magnesium, but becomes specific for nucleotide triphosphates in the presence of manganese. No activity with [gamma32P]ATP-terminated pppA
-
-
-
additional information
?
-
nonstructural protein Nsp2 of Semliki Forest virus specifically cleaves the gamma,beta-triphosphate bond at the 5'-end of RNA. The same activity is demonstrated for the N-terminal fragment of Semliki Forest virus Nsp2-N (residues 1-470). The C-terminal part of Semliki Forest virus Nsp2-C (residues 471-799) has no RNA triphosphatase activity
-
-
-
additional information
?
-
-
nonstructural protein Nsp2 of Semliki Forest virus specifically cleaves the gamma,beta-triphosphate bond at the 5'-end of RNA. The same activity is demonstrated for the N-terminal fragment of Semliki Forest virus Nsp2-N (residues 1-470). The C-terminal part of Semliki Forest virus Nsp2-C (residues 471-799) has no RNA triphosphatase activity
-
-
-
additional information
?
-
nonstructural protein Nsp2 of Sindbis virus specifically cleaves the gamma,beta-triphosphate bond at the 5'-end of RNA
-
-
-
additional information
?
-
-
nonstructural protein Nsp2 of Sindbis virus specifically cleaves the gamma,beta-triphosphate bond at the 5'-end of RNA
-
-
-
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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
a 5'-triphospho-[mRNA] + H2O
a 5'-diphospho-[mRNA] + phosphate
-
-
-
?
a 5'-triphospho-[mRNA] + H2O
a 5'-diphospho-[mRNA] + phosphate
-
-
-
?
a 5'-triphospho-[mRNA] + H2O
a 5'-diphospho-[mRNA] + phosphate
-
-
-
?
a 5'-triphospho-[mRNA] + H2O
a 5'-diphospho-[mRNA] + phosphate
-
-
-
?
a 5'-triphospho-[mRNA] + H2O
a 5'-diphospho-[mRNA] + phosphate
-
-
-
?
a 5'-triphospho-[mRNA] + H2O
a 5'-diphospho-[mRNA] + phosphate
-
-
-
-
?
a 5'-triphospho-[mRNA] + H2O
a 5'-diphospho-[mRNA] + phosphate
-
-
-
?
a 5'-triphospho-[mRNA] + H2O
a 5'-diphospho-[mRNA] + phosphate
-
-
-
?
a 5'-triphospho-[mRNA] + H2O
a 5'-diphospho-[mRNA] + phosphate
-
-
-
?
a 5'-triphospho-[mRNA] + H2O
a 5'-diphospho-[mRNA] + phosphate
-
-
-
?
a 5'-triphospho-[mRNA] + H2O
a 5'-diphospho-[mRNA] + phosphate
-
-
-
?
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METALS and IONS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
Mg2+
Mn2+
additional information
Mg2+
activates optimally at 5 mM Mg2+, inhibition above
Mg2+
required, optimum concentrations between 0.03 and 0.5 mM
Mn2+
enzyme CTL1 is completely inactive on trimer RNA in presence of Mn2+, and enzyme CTL1 exhibits ATPase activity only in presence of Mn2+
additional information
Mg2+ and Mn2+ can alter the enzyme substrate specificity, overview
additional information
Mg2+ and Mn2+ can alter the enzyme substrate specificity, overview
additional information
-
Mg2+ and Mn2+ can alter the enzyme substrate specificity, overview
additional information
the substrate specificity of Cet1 is unaltered by Mg2+ or Mn2+
additional information
the substrate specificity of Cet1 is unaltered by Mg2+ or Mn2+
additional information
-
the substrate specificity of Cet1 is unaltered by Mg2+ or Mn2+
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INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
Mn2+
enzyme CTL1 is completely inactive on trimer RNA in presence of Mn2+, and enzyme CTL1 exhibits ATPase activity only in presence of Mn2+
additional information
no inhibition pf Cet1 by 10 mM sodium vanadate; no inhibition pf CTL1 by 10 mM sodium vanadate
-
additional information
no inhibition pf Cet1 by 10 mM sodium vanadate; no inhibition pf CTL1 by 10 mM sodium vanadate
-
additional information
-
no inhibition pf Cet1 by 10 mM sodium vanadate; no inhibition pf CTL1 by 10 mM sodium vanadate
-
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.00299
a 5'-triphospho-[mRNA]
Nsp2-N (N-terminal fragment), pH 7.0, 25°C
-
additional information
additional information
kinetics, overview
-
additional information
additional information
-
kinetics, overview
-
additional information
additional information
capping apparatus kinetics, overview
-
additional information
additional information
-
capping apparatus kinetics, overview
-
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TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.092
a 5'-triphospho-[mRNA]
Nsp2-N (N-terminal fragment), pH 7.0, 25°C
-
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kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
30.77
a 5'-triphospho-[mRNA]
Nsp2-N (N-terminal fragment), pH 7.0, 25°C
-
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pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
7.9
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pH RANGE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
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TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
30
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TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
20 - 42
optimal activity at 25°C, inactivation at 42°C, recombinant enzyme
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ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
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SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
SOURCE
additional information
additional information
-
hCAP1a mRNAs is detected in all tissues tested, RT-PCR expression analysis
brenda
additional information
hCAP1a mRNAs is detected in all tissues tested, RT-PCR expression analysis
brenda
additional information
-
hCAP1b mRNAs is detected in all tissues tested, but to a lesser extent than hCAP1a, RT-PCR expression analysis
brenda
additional information
hCAP1b mRNAs is detected in all tissues tested, but to a lesser extent than hCAP1a, RT-PCR expression analysis
brenda
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LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY
GeneOntology No.
LITERATURE
SOURCE
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GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
evolution
malfunction
metabolism
physiological function
additional information
evolution
CTL1 is the second member of the yeast RNA triphosphatase family. Enzyme CTL1 resembles the C-terminus of Cet1, which has RNA triphosphatase activity
evolution
gene CES5 is identical to gene CET1, which encodes the RNA triphosphatase component of the yeast capping apparatus
evolution
metazoans including humans contain a separate cap methyltransferase (RNA guanine-7-methyltransferase, RNMT), but the RNA 5'-triphosphatase and guanylyltransferase activities are contained as N- and C-terminal domains in a bifunctional capping enzyme (RNGTT) encoded by a single gene
evolution
RNA 5'-triphosphatase activity is widely distributed among prokaryotes, eukaryotes, and viruses
evolution
the nucleotide sequences of hCAP1a and hCAP1b are identical from the 5' flanking region to codon 503, but are entirely different thereafter. The N-terminal portion of the predicted amino acid sequence contains the motif of the tyrosine specific protein phosphatase active site, [LIVMF]-H-C-x(2)-G-x(3)-[STC]-[STAG]-x-[LIVMFY]. The C-terminal portion of the protein is highly homologous to the known mRNA guanylyltransferases and contains all the motifs, i.e. motifs I, III, IIIa, IV, V, VI, and P, which are commonly observed in viral and cellular mRNA guanylyltransferases except that the motif VI is missing in hCAP1b
evolution
-
gene CES5 is identical to gene CET1, which encodes the RNA triphosphatase component of the yeast capping apparatus
-
evolution
-
RNA 5'-triphosphatase activity is widely distributed among prokaryotes, eukaryotes, and viruses
-
evolution
-
CTL1 is the second member of the yeast RNA triphosphatase family. Enzyme CTL1 resembles the C-terminus of Cet1, which has RNA triphosphatase activity
-
malfunction
ceg1-25 mutation of the yeast mRNA guanylyltransferase (capping enzyme) causes a temperature-sensitive growth defect. Truncated mutant protein Cet1(201-549) has RNA triphosphatase activity, heterodimerizes with and stimulates Ceg1 in vitro, and suffices when expressed in single copy for cell growth in vivo. The more extensively truncated mutant Cet1(246-549) also has RNA triphosphatase activity but fails to stimulate Ceg1 in vitro and is lethal when expressed in single copy in vivo
malfunction
deletion mutant analysis of hCAP1a shows that the N-terminal 213 amino acid fragment containing a tyrosine-specific protein phosphatase motif catalyzes the RNA 5'-triphosphatase activity, and the C-terminal 369 amino acid fragment exhibits the mRNA guanylyltransferase activity. When RNA 5'-triphosphatase activity is assayed using [gamma-32 P]pppG-RNA, mutant His-hCAP1a(1-213) releases [32P]phosphate, while no [32P]phosphate is released by mutant hCAP1a(229-597). Furthermore, the N-terminal half molecule, hCAP1a(1-213) exhibits no NTPase activity, indicating that this half molecule still retains the specificity for RNA
malfunction
HCE1A and HCE1B have been identified as variants of HCE1. Hce1p possesses both TPase and GTase activities, but Hce1a and Hce1b proteins have deletions within the ORF and lack GTase activity. The failure of Hce1a and Hce1b proteins to complement an Saccharomyces cerevisiae cet1DELTA null mutation may be due to the failure of complex formation between the yeast GTase and the human capping enzyme
malfunction
-
ceg1-25 mutation of the yeast mRNA guanylyltransferase (capping enzyme) causes a temperature-sensitive growth defect. Truncated mutant protein Cet1(201-549) has RNA triphosphatase activity, heterodimerizes with and stimulates Ceg1 in vitro, and suffices when expressed in single copy for cell growth in vivo. The more extensively truncated mutant Cet1(246-549) also has RNA triphosphatase activity but fails to stimulate Ceg1 in vitro and is lethal when expressed in single copy in vivo
-
metabolism
during transcription by RNA polymerase II, a cap structure is formed on the 5'-termini of most nascent nuclear pre-mRNAs. Capping of mRNA has been shown to be important for the stabilization, processing, nuclear export, and efficient translation of mRNA. Capping involves at least three enzymes called mRNA 5'-triphosphatase (TPase), mRNA 5'-guanylyltransferase (GTase), and cap methyltransferase (MTase). TPase converts the 5'-triphosphate end of a nascent RNA chain into a diphosphate end, and GTase transfers the GMP moiety of GTP to the newly produced 5'-diphosphate end of RNA to form a blocking structure. Thereafter, MTase attaches a methyl group to the 7 position of the terminal guanosine of RNA. As GTase and TPase are essential for the synthesis of the core cap structure, they are called mRNA 5'-capping enzymes. The physical association of mRNA 5'-guanylyltransferase (GTase) and mRNA 5'-triphosphatase (TPase) is essential for the function of the capping enzyme in vivo
metabolism
subunit Cet1 is a monomer in solution, it binds with recombinant subunit Ceg1 (EC 2.7.7.50) in vitro to form a Cet1-Ceg1 heterodimer. The interaction of Cet1 with Ceg1 elicits over 10fold stimulation of the guanylyltransferase activity of Ceg1, the Cet1-Ceg1 interaction is essential. Potential for regulating mRNA cap formation through protein-protein interactions
metabolism
-
during transcription by RNA polymerase II, a cap structure is formed on the 5'-termini of most nascent nuclear pre-mRNAs. Capping of mRNA has been shown to be important for the stabilization, processing, nuclear export, and efficient translation of mRNA. Capping involves at least three enzymes called mRNA 5'-triphosphatase (TPase), mRNA 5'-guanylyltransferase (GTase), and cap methyltransferase (MTase). TPase converts the 5'-triphosphate end of a nascent RNA chain into a diphosphate end, and GTase transfers the GMP moiety of GTP to the newly produced 5'-diphosphate end of RNA to form a blocking structure. Thereafter, MTase attaches a methyl group to the 7 position of the terminal guanosine of RNA. As GTase and TPase are essential for the synthesis of the core cap structure, they are called mRNA 5'-capping enzymes. The physical association of mRNA 5'-guanylyltransferase (GTase) and mRNA 5'-triphosphatase (TPase) is essential for the function of the capping enzyme in vivo
-
metabolism
-
subunit Cet1 is a monomer in solution, it binds with recombinant subunit Ceg1 (EC 2.7.7.50) in vitro to form a Cet1-Ceg1 heterodimer. The interaction of Cet1 with Ceg1 elicits over 10fold stimulation of the guanylyltransferase activity of Ceg1, the Cet1-Ceg1 interaction is essential. Potential for regulating mRNA cap formation through protein-protein interactions
-
physiological function
capping enzyme RNA 5'-triphosphatase-like 1 (CTL1) is not essential for cell viability and no genetic or physical interactions with the capping enzyme genes are observed. Enzyme CTL1 is probably involved in an RNA processing event other than mRNA capping. CTL1 has no obvious role in 5' mRNA cap formation
physiological function
cellular capping enzymes are bifunctional. The mRNA triphosphatase Cet1 part removes the gamma-phosphate from the 5'-end of RNA, and the Ceg1 part shows RNA guanylyltransferase activity and adds GMP to the resulting diphosphate end in a 5'-5'-orientation. The cap structure is then modified by one or more methyltransferases
physiological function
during transcription by RNA polymerase II, a cap structure is formed on the 5'-termini of most nascent nuclear pre-mRNAs. Capping of mRNA has been shown to be important for the stabilization, processing, nuclear export, and efficient translation of mRNA. Capping involves at least three enzymes called mRNA 5'-triphosphatase (TPase), mRNA 5'-guanylyltransferase (GTase), and cap methyltransferase (MTase). TPase converts the 5'-triphosphate end of a nascent RNA chain into a diphosphate end, and GTase transfers the GMP moiety of GTP to the newly produced 5'-diphosphate end of RNA to form a blocking structure. Thereafter, MTase attaches a methyl group to the 7 position of the terminal guanosine of RNA. As GTase and TPase are essential for the synthesis of the core cap structure, they are called mRNA 5'-capping enzymes. The physical association of mRNA 5'-guanylyltransferase (GTase) and mRNA 5'-triphosphatase (TPase) is essential for the function of the capping enzyme in vivo
physiological function
during transcription by RNA polymerase II, a cap structure is formed on the 5'-termini of most nascent nuclear pre-mRNAs. Capping of mRNA has been shown to be important for the stabilization, processing, nuclear export, and efficient translation of mRNA. Capping involves at least three enzymes called mRNA 5'-triphosphatase (TPase), mRNA 5'-guanylyltransferase (GTase), and cap methyltransferase (MTase). TPase converts the 5'-triphosphate end of a nascent RNA chain into a diphosphate end, and GTase transfers the GMP moiety of GTP to the newly produced 5'-diphosphate end of RNA to form a blocking structure. Thereafter, MTase attaches a methyl group to the 7 position of the terminal guanosine of RNA. As GTase and TPase are essential for the synthesis of the core cap structure, they are called mRNA 5'-capping enzymes. The physical association of mRNA 5'-guanylyltransferase (GTase) and mRNA 5'-triphosphatase (TPase) is essential for the function of the capping enzyme in vivo. In humans, GTase and TPase activities always reside in one enzyme, HCE1
physiological function
enzyme Nsp2 has several distinct functions. It has nucleoside triphosphatase (NTPase) activity at its N-terminal half, which is vital for the virus replication. Nsp2 has RNA helicase activity, which utilizes NTP hydrolysis as the energy source. The C-terminal part of the protein is a papain-like protease responsible for the autocatalytic cleavages of the nonstructural polyprotein. The C-terminal part has a nuclear localization sequence, which is responsible for sequestering of about half of the molecules to the nucleus during infection. Furthermore, Nsp2 regulates transcription of the subgenomic 26 S RNA. Nsp2 has yet an additional activity required for capping of the virus mRNAs. Capping of cellular mRNAs occurs in the nucleus and comprises four different reactions. RNA 5'-triphosphatase removes the gamma-phosphate from the 5'-end of the nascent RNA molecule (pppRNA -> ppRNA). Guanylyltransferase reacts with a GTP molecule to form a covalent complex with GMP, which is then transferred from guanylyltransferase to the 5'-end of RNA to form G(5')ppp(5')NpRNA. Methylation by guanine-7N-methyltransferase yields an RNA molecule with the cap0 structure (m7GpppNpRNA). Further methylation by nucleoside-2'-O-methyltransferase of the riboses of the penultimate and the adjacent nucleotides yields cap1 and cap2 structures, respectively. Unlike host cellular mRNAs, the capping of alphavirus RNAs takes place in the cytoplasm and is carried out by reactions that differ from the nuclear reactions. Nsp2 hydrolyzes only the gamma,beta-triphosphate bond at the 5'-end of RNA
physiological function
enzyme Nsp2 has several distinct functions. It has nucleoside triphosphatase (NTPase) activity at its N-terminal half, which is vital for the virus replication. Nsp2 has RNA helicase activity, which utilizes NTP hydrolysis as the energy source. The C-terminal part of the protein is a papain-like protease responsible for the autocatalytic cleavages of the nonstructural polyprotein. The C-terminal part has a nuclear localization sequence, which is responsible for sequestering of about half of the molecules to the nucleus during infection. Furthermore, Nsp2 regulates transcription of the subgenomic 26 S RNA. Nsp2 has yet an additional activity required for capping of the virus mRNAs. Capping of cellular mRNAs occurs in the nucleus and comprises four different reactions. RNA 5'-triphosphatase removes the gamma-phosphate from the 5'-end of the nascent RNA molecule (pppRNA -> ppRNA). Guanylyltransferase reacts with a GTP molecule to form a covalent complex with GMP, which is then transferred from guanylyltransferase to the 5'-end of RNA to form G(5')ppp(5')NpRNA. Methylation by guanine-7N-methyltransferase yields an RNA molecule with the cap0 structure (m7GpppNpRNA). Further methylation by nucleoside-2'-O-methyltransferase of the riboses of the penultimate and the adjacent nucleotides yields cap1 and cap2 structures, respectively. Unlike host cellular mRNAs, the capping of alphavirus RNAs takes place in the cytoplasm and is carried out by reactions that differ from the nuclear reactions
physiological function
enzyme variant Hce1p displays both mRNA 5'-triphosphatase (TPase) and mRNA 5'-guanylyltransferase (GTase, EC 2.7.7.50) activities, and it forms a cap structure at the 5'-triphosphate end of RNA, demonstrating that it indeed specifies an active mRNA 5'-capping enzyme. The recombinant proteins derived from HCE1A and HCE1B possess only TPase activity. When expressed from ADH1 promoter, HCE1, but not HCE1A and HCE1B, complements Saccharomyces cerevisiae CEG1 and CET1, the genes for GTase and TPase in yeast, respectively. These results demonstrate that the N-terminal part of Hce1p is responsible for TPase activity and the C-terminal part is essential for GTase activity. In addition, the human TPase domain cannot functionally substitute for the yeast enzyme in vivo
physiological function
recombinant enzyme hCAP1b shows RNA 5'-triphosphatase activity, but neither enzyme-GMP covalent complex formation nor cap structure formation is detected
physiological function
the 5' cap structure of eukaryotic mRNA consists of 7-methylguanosine linked to the end of the transcript via a 5'-5'-triphosphate bridge. Capping occurs by a series of three enzymatic reactions in which the 5'-triphosphate end of nascent pre-mRNA is hydrolyzed to a 5'-diphosphate by RNA triphosphatase, capped with GMP by RNA guanylyltransferase, and then methylated at N7 of guanine by RNA (guanine-7) methyltransferase. RNA capping is essential for cell growth. RNA triphosphatase activity is essential for eukaryotic cell growth. The mammalian capping enzyme Mce1 (a bifunctional triphosphatase-guanylyltransferase) substitutes for Cet1 in vivo. Essential Saccharomyces cerevisiae gene, CES5, when present in high copy, suppresses the temperature-sensitive growth defect caused by the ceg1-25 mutation of the yeast mRNA guanylyltransferase (capping enzyme, EC 2.7.7.50). CES5 is identical to CET1 encoding the RNA triphosphatase component of the yeast capping apparatus. CES5 is a multicopy suppressor of ceg1-25
physiological function
the capping enzyme from cellular sources is a bifunctional enzyme having the activities of mRNA guanylyltransferase and RNA 5'-triphosphatase. The beta-subunit enzyme Cet1 exhibits an RNA 5'-triphosphatase activity which specifically removes the gamma-phosphate from the triphosphate-terminated RNA substrate, but not from nucleoside triphosphates, while the alpha subunit Ceg1 exhibits the mRNA guanylyltransferase activity, which is essential for growth of yeast cells. The gene for RNA 5'-triphosphatase (CET1) is essential for cell viability
physiological function
the capping enzyme selectively binds to the phosphorylated, elongating form of RNA polymerase II (polII) for specific capping of pol II transcripts. Caps are formed on nascent pre-mRNAs by the sequential action of RNA 5'-triphosphatase, which removes the gamma-phosphate of the initiating nucleotide, RNA guanylyltransferase, which transfers GMP from GTP to the resulting diphosphate end, and RNA (guanine-7) methyltransferase, which methylates the guanine N7 position of the newly formed GpppN termini
physiological function
the mammalian mRNA capping enzyme is a bifunctional enzyme containing RNA 5'-triphosphatase and mRNA guanylyl-transferase activities in a single polypeptide
physiological function
the mRNA-capping enzyme from Saccharomyces cerevisiae catalyzes (a) removal of the gamma-phosphoryl group from the 5'-end of the newly formed mRNA and (b) guanylylation of the resulting diphosphoryl end (EC 2.7.7.50)
physiological function
-
during transcription by RNA polymerase II, a cap structure is formed on the 5'-termini of most nascent nuclear pre-mRNAs. Capping of mRNA has been shown to be important for the stabilization, processing, nuclear export, and efficient translation of mRNA. Capping involves at least three enzymes called mRNA 5'-triphosphatase (TPase), mRNA 5'-guanylyltransferase (GTase), and cap methyltransferase (MTase). TPase converts the 5'-triphosphate end of a nascent RNA chain into a diphosphate end, and GTase transfers the GMP moiety of GTP to the newly produced 5'-diphosphate end of RNA to form a blocking structure. Thereafter, MTase attaches a methyl group to the 7 position of the terminal guanosine of RNA. As GTase and TPase are essential for the synthesis of the core cap structure, they are called mRNA 5'-capping enzymes. The physical association of mRNA 5'-guanylyltransferase (GTase) and mRNA 5'-triphosphatase (TPase) is essential for the function of the capping enzyme in vivo
-
physiological function
-
the capping enzyme from cellular sources is a bifunctional enzyme having the activities of mRNA guanylyltransferase and RNA 5'-triphosphatase. The beta-subunit enzyme Cet1 exhibits an RNA 5'-triphosphatase activity which specifically removes the gamma-phosphate from the triphosphate-terminated RNA substrate, but not from nucleoside triphosphates, while the alpha subunit Ceg1 exhibits the mRNA guanylyltransferase activity, which is essential for growth of yeast cells. The gene for RNA 5'-triphosphatase (CET1) is essential for cell viability
-
physiological function
-
the mRNA-capping enzyme from Saccharomyces cerevisiae catalyzes (a) removal of the gamma-phosphoryl group from the 5'-end of the newly formed mRNA and (b) guanylylation of the resulting diphosphoryl end (EC 2.7.7.50)
-
physiological function
-
the 5' cap structure of eukaryotic mRNA consists of 7-methylguanosine linked to the end of the transcript via a 5'-5'-triphosphate bridge. Capping occurs by a series of three enzymatic reactions in which the 5'-triphosphate end of nascent pre-mRNA is hydrolyzed to a 5'-diphosphate by RNA triphosphatase, capped with GMP by RNA guanylyltransferase, and then methylated at N7 of guanine by RNA (guanine-7) methyltransferase. RNA capping is essential for cell growth. RNA triphosphatase activity is essential for eukaryotic cell growth. The mammalian capping enzyme Mce1 (a bifunctional triphosphatase-guanylyltransferase) substitutes for Cet1 in vivo. Essential Saccharomyces cerevisiae gene, CES5, when present in high copy, suppresses the temperature-sensitive growth defect caused by the ceg1-25 mutation of the yeast mRNA guanylyltransferase (capping enzyme, EC 2.7.7.50). CES5 is identical to CET1 encoding the RNA triphosphatase component of the yeast capping apparatus. CES5 is a multicopy suppressor of ceg1-25
-
physiological function
-
cellular capping enzymes are bifunctional. The mRNA triphosphatase Cet1 part removes the gamma-phosphate from the 5'-end of RNA, and the Ceg1 part shows RNA guanylyltransferase activity and adds GMP to the resulting diphosphate end in a 5'-5'-orientation. The cap structure is then modified by one or more methyltransferases
-
physiological function
-
capping enzyme RNA 5'-triphosphatase-like 1 (CTL1) is not essential for cell viability and no genetic or physical interactions with the capping enzyme genes are observed. Enzyme CTL1 is probably involved in an RNA processing event other than mRNA capping. CTL1 has no obvious role in 5' mRNA cap formation
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additional information
Mg2+ and Mn2+ can alter the enzyme substrate specificity, overview
additional information
Mg2+ and Mn2+ can alter the enzyme substrate specificity, overview
additional information
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Mg2+ and Mn2+ can alter the enzyme substrate specificity, overview
additional information
neither the full-length human capping enzyme nor its TPase domain interact with the yeast GTase. The failure of the human TPase activity to complement an Saccharomyces cerevisiae cet1DELTA null mutation is attributable, at least in part, to the inability of the human capping enzyme to associate with the yeast GTase, and the physical association of GTase and TPase is essential for the function of the capping enzyme in vivo
additional information
-
neither the full-length human capping enzyme nor its TPase domain interact with the yeast GTase. The failure of the human TPase activity to complement an Saccharomyces cerevisiae cet1DELTA null mutation is attributable, at least in part, to the inability of the human capping enzyme to associate with the yeast GTase, and the physical association of GTase and TPase is essential for the function of the capping enzyme in vivo
additional information
-
the active site for RNA 5'-triphosphatase resides in the N-terminal 213 amino acids fragment, whereas mRNA guanylyltransferase activity resides in the C-terminal 289 amino acid fragment
additional information
the active site for RNA 5'-triphosphatase resides in the N-terminal 213 amino acids fragment, whereas mRNA guanylyltransferase activity resides in the C-terminal 289 amino acid fragment
additional information
when His-Cet1(205-549) is incubated with 5'-[gamma-32P]GTP-terminated RNA, [32P]phosphate is released, while with His-Cet1(1-265) it is not, indicating that the active site resides in the C-terminal 345-amino acids fragment (39 kDa)
additional information
-
when His-Cet1(205-549) is incubated with 5'-[gamma-32P]GTP-terminated RNA, [32P]phosphate is released, while with His-Cet1(1-265) it is not, indicating that the active site resides in the C-terminal 345-amino acids fragment (39 kDa)
additional information
-
when His-Cet1(205-549) is incubated with 5'-[gamma-32P]GTP-terminated RNA, [32P]phosphate is released, while with His-Cet1(1-265) it is not, indicating that the active site resides in the C-terminal 345-amino acids fragment (39 kDa)
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additional information
-
Mg2+ and Mn2+ can alter the enzyme substrate specificity, overview
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Show AA Sequence and Transmembrane Helices (2501 entries)
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SUBUNIT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
additional information
?
x * 80000, beta-subunit, SDS-PAGE, x * 61849, beta-subunit, sequence calculation
?
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x * 80000, beta-subunit, SDS-PAGE, x * 61849, beta-subunit, sequence calculation
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additional information
subunit Cet1 is a monomer in solution, it binds with recombinant subunit Ceg1 (EC 2.7.7.50) in vitro to form a Cet1-Ceg1 heterodimer
additional information
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subunit Cet1 is a monomer in solution, it binds with recombinant subunit Ceg1 (EC 2.7.7.50) in vitro to form a Cet1-Ceg1 heterodimer
additional information
the yeast Saccharomyces cerevisiae mRNA capping enzyme is composed of two subunits of alpha (mRNA guanylyltransferase, 52 kDa) and beta (RNA 5'-triphosphatase)
additional information
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the yeast Saccharomyces cerevisiae mRNA capping enzyme is composed of two subunits of alpha (mRNA guanylyltransferase, 52 kDa) and beta (RNA 5'-triphosphatase)
additional information
-
the yeast Saccharomyces cerevisiae mRNA capping enzyme is composed of two subunits of alpha (mRNA guanylyltransferase, 52 kDa) and beta (RNA 5'-triphosphatase)
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additional information
-
subunit Cet1 is a monomer in solution, it binds with recombinant subunit Ceg1 (EC 2.7.7.50) in vitro to form a Cet1-Ceg1 heterodimer
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PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
E345A
site-directed mutagenesis, the mutant cannot complement Saccharomyces cerevisiae CEG1 deficient mutants
K294A
site-directed mutagenesis, the mutant cannot complement Saccharomyces cerevisiae CEG1 deficient mutants
K458A
site-directed mutagenesis, the mutant cannot complement Saccharomyces cerevisiae CEG1 deficient mutants
K460A
site-directed mutagenesis, the mutant cannot complement Saccharomyces cerevisiae CEG1 deficient mutants
R299A
site-directed mutagenesis, the mutant cannot complement Saccharomyces cerevisiae CEG1 deficient mutants
K192N
site-directed mutagenesis, the mutation in the nucleotide-binding site completely abolishes RNA triphosphatase and nucleoside triphosphatase activities of Semliki Forest virus Nsp2 and Nsp2-N (N-terminal fragment)
additional information
additional information
CaCET1 rescues CET1-deficient Saccharomyces cerevisiae cells when expressed under the control of the ADH1 promoter, whereas the human capping enzyme derivatives that are active for TPase activity but defective in mRNA 5'-guanylyltransferase (GTase) activity do not. Yeast two-hybrid analysis reveals that Candida albicans Cet1p can bind to the Saccharomyces cerevisiae GTase in addition to its endogenous partner, the Candida albicans GTase. In contrast, neither the full-length human capping enzyme nor its TPase domain interact with the yeast GTase. The failure of the human TPase activity to complement an Saccharomyces cerevisiae cet1DELTA null mutation is attributable, at least in part, to the inability of the human capping enzyme to associate with the yeast GTase, and the physical association of GTase and TPase is essential for the function of the capping enzyme in vivo. Complementation of an Saccharomyces cerevisiae ceg1DELTA null mutation by Candida albicans CET1 and interaction of CaCet1p with ScCeg1p (alpha-subunit with mRNA 5'-guanylyltransferase (GTase) activity)
additional information
-
CaCET1 rescues CET1-deficient Saccharomyces cerevisiae cells when expressed under the control of the ADH1 promoter, whereas the human capping enzyme derivatives that are active for TPase activity but defective in mRNA 5'-guanylyltransferase (GTase) activity do not. Yeast two-hybrid analysis reveals that Candida albicans Cet1p can bind to the Saccharomyces cerevisiae GTase in addition to its endogenous partner, the Candida albicans GTase. In contrast, neither the full-length human capping enzyme nor its TPase domain interact with the yeast GTase. The failure of the human TPase activity to complement an Saccharomyces cerevisiae cet1DELTA null mutation is attributable, at least in part, to the inability of the human capping enzyme to associate with the yeast GTase, and the physical association of GTase and TPase is essential for the function of the capping enzyme in vivo. Complementation of an Saccharomyces cerevisiae ceg1DELTA null mutation by Candida albicans CET1 and interaction of CaCet1p with ScCeg1p (alpha-subunit with mRNA 5'-guanylyltransferase (GTase) activity)
additional information
-
CaCET1 rescues CET1-deficient Saccharomyces cerevisiae cells when expressed under the control of the ADH1 promoter, whereas the human capping enzyme derivatives that are active for TPase activity but defective in mRNA 5'-guanylyltransferase (GTase) activity do not. Yeast two-hybrid analysis reveals that Candida albicans Cet1p can bind to the Saccharomyces cerevisiae GTase in addition to its endogenous partner, the Candida albicans GTase. In contrast, neither the full-length human capping enzyme nor its TPase domain interact with the yeast GTase. The failure of the human TPase activity to complement an Saccharomyces cerevisiae cet1DELTA null mutation is attributable, at least in part, to the inability of the human capping enzyme to associate with the yeast GTase, and the physical association of GTase and TPase is essential for the function of the capping enzyme in vivo. Complementation of an Saccharomyces cerevisiae ceg1DELTA null mutation by Candida albicans CET1 and interaction of CaCet1p with ScCeg1p (alpha-subunit with mRNA 5'-guanylyltransferase (GTase) activity)
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additional information
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construction of C-terminal deletion and of N-terminal mutants (His-CAP1a (1-213) and His-CAP1a (229-597)) of hCAP1a
additional information
construction of C-terminal deletion and of N-terminal mutants (His-CAP1a (1-213) and His-CAP1a (229-597)) of hCAP1a
additional information
construction of His-tagged C- and N-terminal deletion mutants, His-Cet1(1-265) and His-Cet1(205-549), respectively, and expression in Escherichia coli
additional information
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construction of His-tagged C- and N-terminal deletion mutants, His-Cet1(1-265) and His-Cet1(205-549), respectively, and expression in Escherichia coli
additional information
generation of ceg1-25 mutation of the yeast mRNA guanylyltransferase (capping enzyme) causing a temperature-sensitive growth defect. Construction of truncated mutant Cet1 proteins, Cet1(201-549) and Cet1(246-549). Mutant Cet1(201-549) has RNA triphosphatase activity, heterodimerizes with and stimulates Ceg1 in vitro, and suffices when expressed in single copy for cell growth in vivo, but the more extensively truncated mutant Cet1(246-549) fails to stimulate Ceg1 in vitro and is lethal when expressed in single copy in vivo, while it also has RNA triphosphatase activity. Construction and complementation of a DELTAcet1 mutant by mouse capping enzyme, overview
additional information
-
generation of ceg1-25 mutation of the yeast mRNA guanylyltransferase (capping enzyme) causing a temperature-sensitive growth defect. Construction of truncated mutant Cet1 proteins, Cet1(201-549) and Cet1(246-549). Mutant Cet1(201-549) has RNA triphosphatase activity, heterodimerizes with and stimulates Ceg1 in vitro, and suffices when expressed in single copy for cell growth in vivo, but the more extensively truncated mutant Cet1(246-549) fails to stimulate Ceg1 in vitro and is lethal when expressed in single copy in vivo, while it also has RNA triphosphatase activity. Construction and complementation of a DELTAcet1 mutant by mouse capping enzyme, overview
additional information
-
construction of His-tagged C- and N-terminal deletion mutants, His-Cet1(1-265) and His-Cet1(205-549), respectively, and expression in Escherichia coli
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additional information
-
generation of ceg1-25 mutation of the yeast mRNA guanylyltransferase (capping enzyme) causing a temperature-sensitive growth defect. Construction of truncated mutant Cet1 proteins, Cet1(201-549) and Cet1(246-549). Mutant Cet1(201-549) has RNA triphosphatase activity, heterodimerizes with and stimulates Ceg1 in vitro, and suffices when expressed in single copy for cell growth in vivo, but the more extensively truncated mutant Cet1(246-549) fails to stimulate Ceg1 in vitro and is lethal when expressed in single copy in vivo, while it also has RNA triphosphatase activity. Construction and complementation of a DELTAcet1 mutant by mouse capping enzyme, overview
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PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
recombinant GST-tagged Cet1p from CET1-deficient Saccharomyces cerevisiae strain by glutathione affinity chromatography
recombinant GST-tagged enzyme variants from Escherichia coli by glutathione affinity separation
recombinant His-tagged enzyme from Escherichia coli strain BL21(DE3)pLysS by nickel affinity chromatography
recombinant His-tagged wild-type and mutant enzymes from Escherichia coli strain BL21(DE3) by nickel affinity chromatography
recombinant His-tagged wild-type and mutant enzymes from Escherichia coli strain BL21(DE3)pLysS by nickel affinity chromatography
recombinant His7-tagged Cet1 from Escherichia coli strain BL21(DE3) by nickel affinity chromatography, dialysis, cation exchange chromatography, dialysis, and anion exchange chromatography
recombinant His7-tagged wild-type and truncated mutant CTL1s from Escherichia coli strain BL21(DE3) by nickel affinity chromatography, anion exchange chromatography, and cation exchange chromatography
recombinant N-terminally His-tagged wild-type and mutant enzymes from Escherichia coli strain BL21(DE3) by nickel affinity chromatography
recombinant N-terminally His-tagged wild-type enzyme from Escherichia coli strain BL21(DE3) by nickel affinity chromatography
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CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
gene CET1, DNA and amino acid sequence determination and analysis, sequence comparison, functional expression as GST-tagged enzyme in a CET1-deficient Saccharomyces cerevisiae strain under control of the ADH1 promoter. Complementation of the Saccharomyces cerevisiae ceg1DELTA null mutation by Candida albicans CET1 and interaction of CaCet1p with ScCeg1p
gene CET1, encodes the beta-subunit with RNA 5'-triphosphatase activity, DNA and amino acid sequence determination and analysis, recombinant expression in Escherichia coli
gene Cet1, recombinant expression of His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3). Gene CES5, DNA and amino acid sequence determination and analysis, CES5 is a multicopy suppressor of ceg1-25
gene CET1, recombinant expression of wild-type His7-tagged Cet1 and of C-terminally truncated mutant Cet1 (265-549) in Escherichia coli strain BL21(DE3)
gene CTL1, DNA and amino acid sequence determination and analysis, sequence comparisons, recombinant expression of the GFP-tagged enzyme in Saccharomyces cerevisiae strain YSB613 cells in nucleus and cytoplasm, recombinant expression of His7-tagged Cet1 in Escherichia coli strain BL21(DE3)
gene hCAP1a, DNA and amino acid sequence determination and analysis, sequence comparisons, recombinant expression of His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3)pLysS, RT-PCR expression analysis
gene hCAP1b, DNA and amino acid sequence determination and analysis, sequence comparisons, recombinant expression of His-tagged enzyme in Escherichia coli strain BL21(DE3)pLysS, RT-PCR expression analysis
gene nsP2, recombinant expression of N-terminally His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3)
gene nsP2, recombinant expression of N-terminally His-tagged wild-type enzyme in Escherichia coli strain BL21(DE3)
gene RNGTT, DNA and amino acid sequence deterination and analysis, genetic organization, human mRNA capping enzyme maps to chromosome 6q16 between STS markers AFMB298ZG9 and AFM185XD10
three highly related cDNAs, HCE1 (human mRNA capping enzyme 1), HCE1A, and HCE1B, are cloned from a HeLa cDNA library. DNA and amino acid sequence determination and analysis, sequence comparisons. The HCE1 cDNA has the longest ORF, which can encode a 69 kDa protein. A short region of 69 bp in the 34-half of the HCE1 ORF is missing in HCE1A and HCE1B, and, additionally, HCE1B has an early translation termination signal, which suggests that the latter two cDNAs represent alternatively spliced product. Semiquantitative RT-PCR enzyme expression analysis. Recombinant expression of GST-tagged enzyme variants in Escherichia coli. When expressed from ADH1 promoter, HCE1, but not HCE1A and HCE1B, functionally complements Saccharomyces cerevisiae CEG1 and CET1, the genes for GTase and TPase in yeast, respectively
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REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Tsukamoto, T.; Shibagaki, Y.; Imajoh-Ohmi, S.; Murakoshi, T.; Suzuki, M.; Nakamura, A.; Gotoh, H.; Mizumoto, K.
Isolation and characterization of the yeast mRNA capping enzyme beta subunit gene encoding RNA 5'-triphosphatase, which is essential for cell viability
Biochem. Biophys. Res. Commun.
239
116-122
1997
Saccharomyces cerevisiae (O13297), Saccharomyces cerevisiae, Saccharomyces cerevisiae ATCC 204508 (O13297)
brenda
Tsukamoto, T.; Shibagaki, Y.; Murakoshi, T.; Suzuki, M.; Nakamura, A.; Gotoh, H.; Mizumoto, K.
Cloning and characterization of two human cDNAs encoding the mRNA capping enzyme
Biochem. Biophys. Res. Commun.
243
101-108
1998
Homo sapiens, Homo sapiens (O60942)
brenda
Yamada-Okabe, T.; Mio, T.; Matsui, M.; Kashima, Y.; Arisawa, M.; Yamada-Okabe, H.
Isolation and characterization of the Candida albicans gene for mRNA 5'-triphosphatase Association of mRNA 5-triphosphatase and mRNA 5-guanylyltransferase activities is essential for the function of mRNA 5-capping enzyme in vivo
FEBS Lett.
435
49-54
1998
Homo sapiens (O60942), Homo sapiens, Candida albicans (O93803), Candida albicans, Candida albicans ATCC MYA-2876 (O93803)
brenda
Pillutla, R.; Shimamoto, A.; Furuichi, Y.; Shatkin, A.
Human mRNA capping enzyme (RNGTT) and cap methyltransferase (RNMT) map to 6q16 and 18p11.22-p11.23, respectively
Genomics
54
351-353
1998
Homo sapiens (O60942)
-
brenda
Itoh, N.; Mizumoto, K.; Kaziro, Y.
Messenger RNA guanylyltransferase from Saccharomyces cerevisiae. II. Catalytic properties
J. Biol. Chem.
259
13930-13936
1984
Saccharomyces cerevisiae (O13297), Saccharomyces cerevisiae, Saccharomyces cerevisiae ATCC 204508 (O13297)
brenda
Vasiljeva, L.; Merits, A.; Auvinen, P.; Krinen, L.
Identification of a novel function of the alphavirus capping apparatus. RNA 5-triphosphatase activity of Nsp2
J. Biol. Chem.
275
17281-17287
2000
Sindbis virus (P03317), Sindbis virus, Semliki forest virus (P08411), Semliki forest virus
brenda
Kiong Ho, C.; Schwer, B.; Shuman, S.
Genetic, physical, and functional interactions between the triphosphatase and guanylyltransferase components of the mRNA capping apparatus
Mol. Cell. Biol.
18
5189-5198
1998
Saccharomyces cerevisiae (O13297), Saccharomyces cerevisiae, Saccharomyces cerevisiae ATCC 204508 (O13297)
brenda
Yamada-Okabe, T.; Doi, R.; Shimmi, O.; Arisawa, M.; Yamada-Okabe, H.
Isolation and characterization of a human cDNA for mRNA 5'-capping enzyme
Nucleic Acids Res.
26
1700-1706
1998
Homo sapiens (O60942), Homo sapiens
brenda
Rodriguez, C.; Takagi, T.; Cho, E.; Buratowski, S.
A Saccharomyces cerevisiae RNA 5'-triphosphatase related to mRNA capping enzyme
Nucleic Acids Res.
27
2181-2188
1999
Saccharomyces cerevisiae (O13297), Saccharomyces cerevisiae (Q03220), Saccharomyces cerevisiae, Saccharomyces cerevisiae ATCC 204508 (O13297), Saccharomyces cerevisiae ATCC 204508 (Q03220)
brenda
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