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Information on EC 2.7.7.19 - polynucleotide adenylyltransferase and Organism(s) Bos taurus and UniProt Accession P25500

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IUBMB Comments
Also acts slowly with CTP. Catalyses template-independent extension of the 3'- end of a DNA strand by one nucleotide at a time. Cannot initiate a chain de novo. The primer, depending on the source of the enzyme, may be an RNA or DNA fragment, or oligo(A) bearing a 3'-OH terminal group. See also EC 2.7.7.6 DNA-directed RNA polymerase.
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Bos taurus
UNIPROT: P25500
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Word Map
The taxonomic range for the selected organisms is: Bos taurus
The enzyme appears in selected viruses and cellular organisms
Synonyms
poly(a) polymerase, pap i, fam46c, gld-2, star-pap, fam46a, poly(a) polymerase i, pap ii, papd5, mtpap, more
SYNONYM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
poly(A) polymerase II
-
adenosine triphosphate:ribonucleic acid adenylyltransferase
-
-
-
-
AMP polynucleotidylexotransferase
-
-
-
-
ATP-polynucleotide adenylyltransferase
-
-
-
-
ATP:polynucleotidylexotransferase
-
-
-
-
neo-PAP
-
-
-
-
NTP polymerase
-
-
-
-
nucleotidyltransferase, polyadenylate
-
-
-
-
PAP I
-
-
-
-
poly(A) hydrolase
-
-
-
-
poly(A) polymerase
poly(A) synthetase
-
-
-
-
polyadenylate nucleotidyltransferase
-
-
-
-
polyadenylate polymerase
-
-
-
-
polyadenylate synthetase
-
-
-
-
polyadenylic acid polymerase
-
-
-
-
polyadenylic polymerase
-
-
-
-
RNA adenylating enzyme
-
-
-
-
RNA formation factors, PF1
-
-
-
-
terminal riboadenylate transferase
-
-
-
-
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
ATP + RNAn = diphosphate + RNAn+1
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
nucleotidyl group transfer
-
-
-
-
SYSTEMATIC NAME
IUBMB Comments
ATP:polynucleotide adenylyltransferase
Also acts slowly with CTP. Catalyses template-independent extension of the 3'- end of a DNA strand by one nucleotide at a time. Cannot initiate a chain de novo. The primer, depending on the source of the enzyme, may be an RNA or DNA fragment, or oligo(A) bearing a 3'-OH terminal group. See also EC 2.7.7.6 DNA-directed RNA polymerase.
CAS REGISTRY NUMBER
COMMENTARY hide
9026-30-6
-
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
ATP + RNA primer
diphosphate + RNA primer-A
show the reaction diagram
in absence of ATP: no activity with 8-Cl-ATP, and 8-amino-ATP results in chain termination, in presence of ATP: polyadenylation of the primer is blocked (inhibited) by 8-amino-ATP and 8-Cl-ATP
-
-
?
adenosine 5'-O-(1-thiotriphosphate) + RNA
diphosphate + ?
show the reaction diagram
-
SP-diastereomer
-
-
?
ATP + myosin short poly(A)
?
show the reaction diagram
-
-
-
-
?
ATP + oligo(A)n
diphosphate + oligo(A)n+1
show the reaction diagram
-
-
-
-
?
ATP + oligoadenylate
?
show the reaction diagram
-
-
-
-
?
ATP + poly(A)n
diphosphate + poly(A)n+1
show the reaction diagram
-
-
-
-
?
ATP + RNA
?
show the reaction diagram
ATP + RNA
diphosphate + RNA(A)n
show the reaction diagram
ATP + RNA (A)15
diphosphate + RNA (A)16
show the reaction diagram
-
-
-
-
?
ATP + RNA(A)15
diphosphate + RNA(A)16
show the reaction diagram
-
-
-
-
?
ATP + RNAn
diphosphate + RNAn+1
show the reaction diagram
-
adenine can bind in two different configurations in the PAP active site
-
-
?
additional information
?
-
-
overview on substrates and primers
-
-
?
NATURAL SUBSTRATE
NATURAL PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
ATP + RNA
?
show the reaction diagram
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
KCl
-
requirement is dependent on the primer and the divalent cation used
additional information
-
overview: ion requirements, poly(A) polymerases purified from different sources, and in some cases even from the same source, respond differently to the presence of Mg2+ and Mn2+
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
2'-dATP
-
-
3'-dATP
-
-
Bentonite
-
-
-
CaCl2
-
remaining activity 3%
CoCl2
-
remaining activity 75%
cordycepin
-
not
CTP
-
-
CuCl2
-
remaining activity 0.1%
diphosphate
GTP
-
-
heparin
-
-
Ionic strength
-
-
-
Mn2+
-
high concentrations
N-ethylmaleimide
-
inhibits Mn2+-activated enzyme
Na2HPO4
-
-
phosphate
-
-
polyamines
-
-
-
Polyphosphate
-
with 20 nM to 0.002 mM of polyphosphate the length of the products is reduced but inhibition is not complete
Polyvinyl sulfate
-
-
Proflavine
-
only at very high levels
Ribonucleoside triphosphates other than ATP
-
-
rifamycin AF/013
-
O-n-octyloxime of 3-formylrifamycin SV
UTP
-
-
Zn2+
-
inhibits PAP activity at concentrations above 10 µM in the presence of 5 mM MgCl2
additional information
-
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.05 - 2.191
ATP
0.01 - 0.3
oligo(A)n
-
0.2
oligoadenylate
-
in presence of Mg2+, pH 9.0, 35°C
0.0036 - 0.36
poly(A)n
-
0.51 - 24.99
RNA (A)15
-
0.007
short poly(A)
-
-
additional information
additional information
-
-
-
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.002 - 30
ATP
0.0025 - 34.5
RNA(A)15
-
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
28.33
-
35°C, pH 9.0
additional information
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
8
-
Mg2+-activated enzyme
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7 - 8.8
-
about 50% of maximum activity at pH 7.0 and pH 8.8
7 - 9
-
active in this range
7.2 - 9.2
-
pH 7.2: 55% of maximum activity, pH 9.2: 62% of maximum activity
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
35
-
assay at
37
-
assay at
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
-
Uniprot
Manually annotated by BRENDA team
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
UNIPROT
ENTRY NAME
ORGANISM
NO. OF AA
NO. OF TRANSM. HELICES
MOLECULAR WEIGHT[Da]
SOURCE
SEQUENCE
LOCALIZATION PREDICTION?
PAPOA_BOVIN
739
0
82441
Swiss-Prot
other Location (Reliability: 4)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
120000 - 140000
-
Mg2+-activated enzyme, gel filtration
140000 - 160000
-
gel filtration
57000
60000
62000
76990
-
predicted from nucleotide sequence
82400
-
predicted from nucleotide sequence
85000
-
x * 85000, SDS-PAGE, recombinant enzyme
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
monomer
additional information
-
binding of enzyme to nuclear poly(A) binding protein results in 80-fold increase in apparent affinity for RNA, mechanism
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
-
not glycosylated
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
in complex with an ATP analog
mutant C36S/C118V/A152C/C160S/C197S/C257S/C293S/C204V in complex with a chemically modified RNA, to 2.25 A resolution
Crystals of PAP complexed to 3'-dATP and Mn2+ are grown from a solution made of 22% (w/v) polyethylene glycol 8000, 100 mM Mes (pH 6), 120 mM ammonium sulfate, 5 mM CaCl2, 2 mM MnCl2, and 2 mM beta-mercaptoethanol. Cocrystallization of PAP with 3'-dATP and MgCl2 is performed under conditions similar to those reported, with the exception that 5 mM MgCl2 is used instead of 2 mM MnCl2. Crystals are of the space group p212121, with one monomer per asymmetric unit and a solvent content of ~55%.
-
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
C36S/C118V/A152C/C160S/C197S/C257S/C293S/C204V
introduction of a Cys residue in a mutant lacking all endogenous Cys residues. Mutant achieves maximum specific disulfide cross-linking efficiency. The resulting construct is active and, when mixed with a chemically modified RNA, yields crystals of an enzyme-RNA complex
C36S/C118V/C160S/C197S/C257S/C293S/C204V
mutation of all seven endogenous cysteine residues. Mutant is able to bind RNA at a level similar to that of wild-type, but no longer forms nonspecific disulfide cross-links with the modified RNA
D167N
-
strong reduction of catalytic efficiency
D167N/N202A
-
sharp decrease in catalytic efficiency
D167N/T317G
-
sharp decrease in catalytic efficiency
D167N/V247R
-
sharp decrease in catalytic efficiency
F100D
-
strong reduction of catalytic efficiency
G203H
-
strong reduction of catalytic efficiency
K228A
-
increase in Km for ATP
K232A
-
increase in Km for ATP
N202A
-
high Km value for RNA, strong reduction of catalytic efficiency, incorporates etheno-ATP and N1-methyl-ATP better than wild-type, but incorporates N6-methyl-ATP poorly
N239A
-
kcat for ATP is reduced fourfold
R199A
-
reduces kcat for RNA about 30-fold
T317G
-
increase in Km for ATP, tolerates etheno-ATP and GTP, rejects N6-methyl-ATP
V156A
-
strong reduction of catalytic efficiency
V247R
-
kcat for ATP is reduced 2000-fold, strong reduction of catalytic efficiency
Y237A
-
increase in Km for ATP
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
freezing and thawing accelerates inactivation
-
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-20°C, 50% glycerol, 80% inactivation in the first few weeks, the 20% remaining activity is stable for more than 2 years
-
-70°C or under liquid nitrogen, extensive inactivation
-
-70°C, stable
-
-80°C, 10-20 mg protein/ml, flash frozen
-
-80°C, storage for 5 days including 2 cycles of freezing and thawing results in 35% inactivation
-
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
homogenous preparation of bovine enzyme
-
overview: purification methods
-
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
expression in Escherichia coli Rosetta (DE3) pLysS cells
expression in Escherichia coli
-
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Bollum, F.J.; Chang, L.M.S.; Tsiapalis, C.M.; Dorson, J.W.
Nucleotide polymerizing enzymes from calf thymus gland
Methods Enzymol.
29E
70-81
1974
Bos taurus
-
Manually annotated by BRENDA team
Edmonds, M.
Poly(A) adding enzymes
The Enzymes,3rd Ed. (Boyer,P. D. ,ed. )
15
217-244
1982
Bos taurus, Cricetulus griseus, Coturnix sp., Escherichia coli, Homo sapiens, Mus musculus, Rattus norvegicus, Vaccinia virus
-
Manually annotated by BRENDA team
Winters, M.A.; Edmonds, M.
A poly(A) polymerase from calf thymus. Purification and properities of the enzyme
J. Biol. Chem.
248
4756-4762
1973
Bos taurus
Manually annotated by BRENDA team
Winters, M.A.; Edmonds, M.
A poly(A) polymerase from calf thymus. Characterization of the reaction product and the primer requirement
J. Biol. Chem.
248
4763-4768
1973
Bos taurus
Manually annotated by BRENDA team
Raabe, T.; Bollum, F.J.; Manley, J.L.
Primary structure and expression of bovine poly(A) polymerase
Nature
353
229-234
1991
Bos taurus
Manually annotated by BRENDA team
Tsiapalis, C.M.; Dorson, J.W.; Bollum, F.J.
Purification of terminal riboadenylate transferase from calf thymus gland
J. Biol. Chem.
250
4486-4496
1975
Bos taurus
Manually annotated by BRENDA team
Wahle, E.; Martin, G.; Schiltz, E.; Keller, W.
Isolation and expression of cDNA clones encoding mammalian poly(A) polymerase
EMBO J.
10
4251-4257
1991
Bos taurus
Manually annotated by BRENDA team
Wahle, E.
Purification and characterization of a mammalian polyadenylate polymerase involved in the 3 end processing of messenger RNA precursors
J. Biol. Chem.
266
3131-3139
1991
Bos taurus
Manually annotated by BRENDA team
Ohyama, Y.; Fukami, H.; Ohta, T.
Purification and characterization of a poly(A) polymerase from beef liver nuclei
J. Biochem.
88
337-348
1980
Bos taurus
Manually annotated by BRENDA team
Martin, G.; Keller, W.; Doublie, S.
Crystal structure of mammalian poly(A) polymerase in complex with an analog of ATP
EMBO J.
19
4193-4203
2000
Bos taurus (P25500), Bos taurus
Manually annotated by BRENDA team
Wittmann, T.; Wahle, E.
Purification and characterization of full-length mammalian poly(A) polymerase
Biochim. Biophys. Acta
1350
293-305
1997
Bos taurus
Manually annotated by BRENDA team
Kerwitz, Y.; Kuehn, U.; Lilie, H.; Knoth, A.; Scheuermann, T.; Friedrich, H.; Schwarz, E.; Wahle, E.
Stimulation of poly(A) polymerase through a direct interaction with the nuclear poly(A) binding protein allosterically regulated by RNA
EMBO J.
22
3705-3714
2003
Bos taurus
Manually annotated by BRENDA team
Martin, G.; Moglich, A.; Keller, W.; Doublie, S.
Biochemical and structural insights into substrate binding and catalytic mechanism of mammalian poly(A) polymerase
J. Mol. Biol.
341
911-925
2004
Bos taurus
Manually annotated by BRENDA team
Holbein, S.; Freimoser, F.M.; Werner, T.P.; Wengi, A.; Dichtl, B.
Cordycepin-hypersensitive growth links elevated polyphosphate levels to inhibition of poly(A) polymerase in Saccharomyces cerevisiae
Nucleic Acids Res.
36
353-363
2008
Bos taurus, Saccharomyces cerevisiae, Saccharomyces cerevisiae BY4741
Manually annotated by BRENDA team
Chen, L.S.; Du-Cuny, L.; Vethantham, V.; Hawke, D.H.; Manley, J.L.; Zhang, S.; Gandhi, V.
Chain termination and inhibition of mammalian poly(A) polymerase by modified ATP analogues
Biochem. Pharmacol.
79
669-677
2010
Bos taurus (P25500), Bos taurus
Manually annotated by BRENDA team
Yang, Q.; Faucher, F.; Coseno, M.; Heckman, J.; Doublie, S.
Purification, crystallization and preliminary X-ray diffraction of a disulfide cross-linked complex between bovine poly(A) polymerase and a chemically modified 15-mer oligo(A) RNA
Acta Crystallogr. Sect. F
67
241-244
2011
Bos taurus (P25500), Bos taurus
Manually annotated by BRENDA team