Information on EC 6.5.1.4 - RNA 3'-terminal-phosphate cyclase (ATP)

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

EC NUMBER
COMMENTARY hide
6.5.1.4
-
RECOMMENDED NAME
GeneOntology No.
RNA 3'-terminal-phosphate cyclase (ATP)
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
ATP + RNA 3'-terminal-phosphate = AMP + diphosphate + RNA terminal-2',3'-cyclic-phosphate
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
cyclization
-
-
esterification
-
-
cyclic phosphomonoester
-
Ligation
-
-
-
-
SYSTEMATIC NAME
IUBMB Comments
RNA-3'-phosphate:RNA ligase (cyclizing, AMP-forming)
Adenosine 5'-(gamma-thio)triphosphate can act instead of ATP. cf. EC 6.5.1.5, RNA-3'-phosphate cyclase (GTP).
CAS REGISTRY NUMBER
COMMENTARY hide
85638-41-1
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
-
-
-
Manually annotated by BRENDA team
zebra fish
-
-
Manually annotated by BRENDA team
fruit fly
-
-
Manually annotated by BRENDA team
strain K12
-
-
Manually annotated by BRENDA team
strain YMC10
Uniprot
Manually annotated by BRENDA team
mouse
-
-
Manually annotated by BRENDA team
no activity in Bacillus subtilis
-
-
-
Manually annotated by BRENDA team
no activity in Haemophilus influenzae
-
-
-
Manually annotated by BRENDA team
no activity in Helicobacter pylori
-
-
-
Manually annotated by BRENDA team
no activity in Mycoplasma genitalium
-
-
-
Manually annotated by BRENDA team
no activity in Synechocystis sp.
-
-
-
Manually annotated by BRENDA team
-
SwissProt
Manually annotated by BRENDA team
-
SwissProt
Manually annotated by BRENDA team
fission yeast
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
metabolism
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
ATP + (Ap)npAp
AMP + ?
show the reaction diagram
-
pure oligoribonucleotide
-
?
ATP + (Up)10pGp
AMP + ?
show the reaction diagram
-
pure oligoribonucleotide
-
?
ATP + 5S rRNA
AMP + ?
show the reaction diagram
ATP + 5S rRNA-pCp
AMP + ?
show the reaction diagram
-
-
-
?
ATP + AAAACAAAAGp
AMP + ?
show the reaction diagram
-
-
?
ATP + AAAAUAAAAGCp
AMP + ?
show the reaction diagram
ATP + AAAAUAAAAGp
AMP + ?
show the reaction diagram
ATP + AAAAUAAAAGpCp
AMP + diphosphate + AAAAUAAAAGpC-2',3'-cyclic phosphate
show the reaction diagram
ATP + AUGp
AMP + ?
show the reaction diagram
-
pure oligoribonucleotide
-
?
ATP + CCCCACCCCGp
AMP + ?
show the reaction diagram
ATP + CCCCCACCCCGCp
AMP + ?
show the reaction diagram
-
pure oligoribonucleotide
-
?
ATP + DNA 5'-terminal-phosphate
AMP + AppDNA
show the reaction diagram
-
RtcA also catalyzes adenylylation of 5'-phosphate ends of DNA strands to form AppDNA product
-
-
?
ATP + human U14 snoRNA
AMP + ?
show the reaction diagram
ATP + oligodeoxyribonucleotide 3'-terminal phosphate
AMP + diphosphate + oligodeoxyribonucleotide-2',3'-cyclic-phosphate
show the reaction diagram
500fold poorer substrate than oligoribonucleotide 3'-terminal phosphate
-
-
-
ATP + RNA 2'-terminal-phosphate
?
show the reaction diagram
ATP + RNA 3'-terminal-phosphate
?
show the reaction diagram
ATP + RNA 3'-terminal-phosphate
AMP + diphosphate + RNA terminal-2',3'-cyclic phosphate
show the reaction diagram
-
RtcA catalyzes the synthesis of RNA 2',3'-cyclic phosphate ends via an ATP-dependent pathway comprising three nucleotidyl transfer steps: reaction of RtcA with ATP to form a covalent RtcA-(histidinyl-N)-AMP intermediate and release diphosphate, transfer of AMP from RtcA to an RNA 3'-phosphate to form an RNA(3')pp(5')A intermediate, and attack by the terminal nucleoside O2' on the 3'-phosphate to form an RNA 2',3'-cyclic phosphate product and release AMP
-
-
?
ATP + RNA 3'-terminal-phosphate
AMP + diphosphate + RNA terminal-2',3'-cyclic-phosphate
show the reaction diagram
ATP + RNA 3'-terminal-phosphate
AMP + diphosphate + RNA terminal-2',3'-cyclic-phosphodiester
show the reaction diagram
ATP + RNA 5'-terminal-phosphate
AMP + AppRNA
show the reaction diagram
-
RtcA also catalyzes adenylylation of 5'-phosphate ends of RNA strands to form AppRNA product
-
-
?
ATP + tobacco mosaic virus RNA
AMP + ?
show the reaction diagram
-
fragments modified by ligation of pGp, pAp, or pCp
-
?
ATP + tRNA-pNp
AMP + ?
show the reaction diagram
-
-
-
?
ATPgammaS + RNA 3'-terminal-phosphate
AMP + thiodiphosphate + RNA terminal-2',3'-cyclic-phosphate
show the reaction diagram
CTP + RNA 3'-terminal-phosphate
CMP + diphosphate + RNA terminal-2',3'-cyclic-phosphate
show the reaction diagram
GTP + RNA 3'-terminal-phosphate
GMP + diphosphate + RNA terminal-2',3'-cyclic-phosphate
show the reaction diagram
RNA-N3'p + ATP
RNA-N-2',3'-cyclic phosphate + AMP + diphosphate
show the reaction diagram
-
-
-
-
ir
UTP + RNA 3'-terminal-phosphate
UMP + diphosphate + RNA terminal-2',3'-cyclic-phosphate
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
ATP + RNA 3'-terminal-phosphate
?
show the reaction diagram
additional information
?
-
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Ca2+
-
only 5% of the activation by Mg2+
K+
-
optimal concentration: 150-200 mM
Na+
-
optimal concentration: 150-200 mM
additional information
-
no activity with Ca2+, Zn2+, or Cu2+
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
iodoacetamide
-
-
SDS
-
-
Sodium phosphate
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Vanadyl-ribonucleoside complex
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-
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0017 - 0.02
ATP
0.006
ATPgammaS
0.1 - 0.2
GTP
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
additional information
?
Escherichia coli
-
the rate of 2'-phosphate cyclization by RtcA is five orders of magnitude slower than 3'-phosphate cyclization, notwithstanding that RtcA binds with similar affinity to RNA3'p and RNA2'p substrates
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SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.021
-
-
additional information
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5.5 - 6
-
pH-optimum for DNA 5'-adenylylation Tris acetate buffer
8 - 8.5
-
pH-optimum for RNA 3'-cyclization
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
8 - 10.5
-
8.0-9.0: maximal activity, 10.5: 70% of maximal activity
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
30
-
similar rates of reaction at 25C, 30C, and 37C
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0 - 37
-
0C: 25% of maximal activity, 12C: 60% of maximal activity, 25-37C: maximal activity
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
PDB
SCOP
CATH
ORGANISM
UNIPROT
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Pyrococcus horikoshii (strain ATCC 700860 / DSM 12428 / JCM 9974 / NBRC 100139 / OT-3)
Pyrococcus horikoshii (strain ATCC 700860 / DSM 12428 / JCM 9974 / NBRC 100139 / OT-3)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
38000 - 40000
-
glycerol density gradient sedimentation, gel filtration
39000
-
SDS-PAGE
39400
-
calculated from cDNA
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
homodimer
-
the apoenzyme of Escherichia coli RtcA crystallizes as a disulfide-linked homodimer and reveales a fold composed of four tandem modules, each comprising a four-stranded beta sheet overlying two alpha helices, X-ray diffraction
monomer
-
1 * 38000-40000, SDS-PAGE
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
hanging drop vapor diffusion technique, 2 orthorhombic crystal forms, space group P2(1)2(1)2(1), unit-cell dimensions a : 101.8 A, b : 126.6 A and c : 128.8 A, and P2(1)2(1)2 with unit cell dimensions a : 125.8 A, b : 133.5 A, c : 51.0 A
-
crystal structure of the enzyme in complex with a 3'-phosphate terminated RNA and adenosine in the AMP-binding pocket
crystallized in the states St-Rtc, St-Rtc+Mn, St-Rtc+ATP, St-Rtc+AMP and St-Rtc-AMP
-
native enzyme, with AMP covalently bound, in complex with AMP, with ATP, or Mn2+, at 2.25 A, 2.25A, 2.9 A, 2.4 A and 3.2 A resolutions, respectively. Upon binding of Mg2+ to residue E10, the triphosphate group of the trapped ATP changes its conformation, with help of ligands R17 and R39. The Nepsilon atom of H307 attacks the alpha-phosphate group to form a new P-N bond. When a truncated RNA is bound, its 3'-phosphate group may be forced to react with the phosphate group of AMP, and the activiated 3'-phosphate group may be attacked by the 2'-hydroxyl group to generate the 2',3'-cyclic phosphodiester
-
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4
-
3 d, 50% loss of activity
20
-
3-4 h, 50% loss of activity
42
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5 min, 55% loss of activity
50
-
5 min, 95% loss of activity
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
not affected by several cycles of freezing and thawing
-
resistant to protease digestion
-
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-70C, stable for at least 3 months
-
-70C, stable for at least 6 months
-
4C, 50% loss of activity after 3 days
-
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
recombinant enzyme
-
using Ni-NTA chromatography
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
bacterially overexpressed as a fusion protein
-
cDNA characterized
-
cDNA characterized, complements yeast strains depleted of Rcl1p
-
cloned and overexpressed
-
cloned and overexpressed in Escherichia coli as fusion protein with glutathione S-transferase
-
expressed in Escherichia coli
-
expressed in Escherichia coli as a His-tagged fusion protein
expressed in Escherichia coli BL21-Codon Plus (DE3)
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expressed sequence tags encoding cyclase like protein identified
expression in Escherichia coli
-
gene encoding cyclase like protein present
gene rtcA cloned and overexpressed
gene rtcA encoding cyclase cloned and overexpressed
-
overexpressed in Escherichia coli
-
overexpression in Escherichia coli
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
C308A
-
recombinant mutant protein
D287A
-
activity very low, 1% of wild-type enzyme
E270A
-
reduced activity compared to wild-type enzyme, about 80%
F135A
-
activity very low, 3% of wild-type enzyme
F251A
-
considerably reduced activity compared to wild-type enzyme, about 30%
H309G
-
enzyme inactive
P131G
-
considerably reduced activity compared to wild-type enzyme, about 35%
Q104A
-
almost same activiy like wild-type enzyme
Q288A
-
activity very low, 2% of wild-type enzyme
S129A
-
almost same activiy like wild-type enzyme
Y284A
-
considerably reduced activity compared to wild-type enzyme (12%)
D297A
-
enzyme inactive
E14A
-
activity very low, only 4% of RNA is cyclized compared to 95% with wild-type enzyme
H326A
-
almost same activiy like wild-type enzyme
H52A
-
half activity compared to wild-type enzyme
Q51A
-
almost same activiy like wild-type enzyme
R21A
-
enzyme inactive
R40A
-
enzyme inactive
R43A
-
enzyme inactive
Y294A
-
enzyme inactive
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