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Information on EC 2.7.7.19 - polynucleotide adenylyltransferase and Organism(s) Saccharomyces cerevisiae and UniProt Accession P29468

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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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Saccharomyces cerevisiae
UNIPROT: P29468
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Word Map
The taxonomic range for the selected organisms is: Saccharomyces cerevisiae
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
polyadenylate polymerase
-
adenosine triphosphate:ribonucleic acid adenylyltransferase
-
-
-
-
AMP polynucleotidylexotransferase
-
-
-
-
ATP-polynucleotide adenylyltransferase
-
-
-
-
ATP:polynucleotidylexotransferase
-
-
-
-
neo-PAP
-
-
-
-
NTP polymerase
-
-
-
-
nuclear speckle targeted PIPKIalpha regulated-poly(A) polymerase
-
-
nucleotidyltransferase, polyadenylate
-
-
-
-
PAP I
-
-
-
-
poly(A) hydrolase
-
-
-
-
poly(A) polymerase
poly(A) RNA polymerase protein 1
-
poly(A) synthetase
-
-
-
-
polyA polymerase
-
-
polyadenylate nucleotidyltransferase
-
-
-
-
polyadenylate polymerase
polyadenylate synthetase
-
-
-
-
polyadenylic acid polymerase
-
-
-
-
polyadenylic polymerase
-
-
-
-
RNA adenylating enzyme
-
-
-
-
RNA formation factors, PF1
-
-
-
-
terminal riboadenylate transferase
-
-
-
-
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
(A)n + ATP
(A)n+1 + diphosphate
show the reaction diagram
-
-
-
?
(A)n + CTP
(A)n-C + diphosphate
show the reaction diagram
-
-
-
?
(A)n + diphosphate
(A)n-1 + ATP
show the reaction diagram
-
-
-
?
(A)n + GTP
(A)n-G + diphosphate
show the reaction diagram
-
-
-
?
2-aminopurine riboside triphosphate + RNA
?
show the reaction diagram
-
-
-
?
ATP + RNA
diphosphate + RNA(A)n
show the reaction diagram
ATP + oligo(A)14
diphosphate + oligo(A)15
show the reaction diagram
-
-
-
-
?
ATP + oligo(A)17C
diphosphate + oligo(A)18C
show the reaction diagram
-
-
-
-
?
ATP + oligo(A)18
diphosphate + oligo(A)19
show the reaction diagram
-
-
-
-
?
ATP + RNA
diphosphate + RNA(A)n
show the reaction diagram
ATP + RNAn
diphosphate + RNAn+1
show the reaction diagram
-
-
-
-
?
CTP + RNAn
diphosphate + RNAn+1
show the reaction diagram
-
-
-
-
?
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
2-aminopurine riboside triphosphate + RNA
?
show the reaction diagram
-
-
-
?
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
KCl
-
optimal concentration: 60 mM
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
8-amino-ATP
-
C-8 substitution influences sugar pucker conformation, which may affect yPAP efficiently
8-aza-ATP
-
slight inhibition
8-azido-ATP
-
slight inhibition
8-bromo-ATP
-
halogen modification at C-8 may negatively affect the ability of the enzyme to activate the ATP substrate or to transfer the AMP group from the enzyme to the RNA substrate
8-chloro-ATP
-
-
alpha, beta-methylene-ATP
-
a nonreactive ATP analogue
Cordycepin triphosphate
-
5.0 mM
diphosphate
-
product inhibitor
N-ethylmaleimide
-
-
Polyphosphate
additional information
-
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
Dinucleotide
-
in presence of 0.5 mM ATP, activation in decreasing order: GP4G, GP3G, AP6A, GP2G, AP4A, AP2A, GP5G, AP5A, AP3A, activation at 0.01 mM is 4-10fold
phosphatidylinositol 4,5-bisphosphate
-
in the presence of 50 mM phosphatidylinositol 4,5-bisphosphate Star-PAP activity is markedly stimulated
additional information
-
other phosphoinositides besides phosphatidylinositol 4,5-bisphosphate do not affect Star-PAP activity, and no phosphoinositide has an effect on PAPalpha activity
-
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0468 - 0.711
(A)n
0.0197
2-aminopurine riboside triphosphate
-
0.0359 - 0.929
ATP
0.104 - 4.7
CTP
0.055 - 0.062
GTP
0.031 - 0.308
ATP
0.0005 - 0.037
oligo(A)14
-
0.0263
oligo(A)17C
-
incubation at 30 °C for 15 min in the presence of MgATP2-
-
0.0468 - 0.0642
oligo(A)18
-
additional information
additional information
-
dependence on divalent cation concentration
-
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0004 - 0.0063
3'-dATP
0.0019
8-amino-ATP
-
incubation at 37 °C for 20 min
0.0002
polyphosphate P15
-
competitive, 30°C, pH 7.0
-
0.0005
polyphosphate P4
-
competitive, 30°C, pH 7.0
-
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7
-
Mn2+-activated enzyme
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
physiological function
nuclear RNA-dependent ATPase Mtr4 and either the nuclear non-canonical poly(A) polymerases, Trf4 or Trf5 assemble into a Trf4/5Air1/2/Mtr4 polyadenylation complex TRAMP. Disrupting the Mtr4/Trf interaction disrupts specific TRAMP and exosome functions, including small nucleolar RNA processing. A 20 amino acid peptide, residues 98-117 in the N-terminus of Trf5 is important for TRAMP complex formation
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
47000
-
gel filtration
63000
-
1 * 63000, SDS-PAGE
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
monomer
-
1 * 63000, SDS-PAGE
additional information
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
hanging drop vapour diffusion method, using 20% (w/v) PEG 8000, 100 mM magnesium acetate, 100 mM imidazole (pH 6.2), 3% ethylene glycol
mutant D154A in complex with MgATP-RNA, hanging drop vapour diffusion method, in 0.1 M bis-Tris propane, pH 6.4, 0.2 M Li-acetate, and 16% PEG 3350
mutant D154A, trapped in complex with ATP and a five residue poly(A). Enzyme has undergone significant domain movement and shows a closed conformation with extensive interactions between substrates and all three polymerase domains
tethered to the 3'-end processing complex via Fip1 peptide, hanging drop vapour diffusion method, with 100 mM MES, pH 6.5, 8-10% PEG 20000
alone and in complex with 3’-dATP
-
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
D154A
K215A
N189A
N226A
V498Y/C485R
the mutant is unable to bind Fip1 but retains full polymerase activity
Y224F
K215A
-
74- and 56-fold increase in the Km for oligo(A)14 in comparison with wild-type enzyme, 2.8-3.6-fold decrease in Vmax
N226A
-
74- and 56-fold increase in the Km for oligo(A)14 in comparison with wild-type enzyme
Y224F
-
30-fold increase in the Km for oligo(A)14 in comparison with wild-type enzyme, 2.8-3.6-fold decrease in Vmax
Y224S
-
12-fold increase in the Km for oligo(A)14 in comparison with wild-type enzyme, 2.8-3.6-fold decrease in Vmax
additional information
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
30 - 48
remains stable at 30°C, the melting temperature for the wild type enzyme is at 48°C
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
substrates poly(A) and Mg-ATP induce the conformational change, resulting in stabilization of the closed enzyme state and enabling catalysis
a decay of PAP activity with a half-life of approximately 2 h (on ice) is observed when Nonidet-P 40 is omitted from the enzyme storage solution
-
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
Ni-affinity column chromatography
nickel affinity column chromatography and ion exchange chromatography
polynucleotide adenylyltransferase Trf5p copurifies with Mtr4p and Air1p of the polyadenylation complex
-
recombinant yeast PAP is purified by nickel affinity and anion exchange chromatography using NTA agarose and Source S resins
-
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
expressed in HEK-293 cells
-
expression in Escherichia coli
-
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
biotechnology
-
assay in microtiter format
drug development
-
polyadenylation inhibition represents one mode of action for 8-chloroadenosine and 8-aminoadenosine in hematological malignancies, among several possible mechanisms. RNA-directed drugs may offer a valuable strategy in targeting indolent cancers, such as multiple myeloma
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Lingner, J.; Radtke, I.; Wahle, E.; Keller, W.
Purification and characterization of poly(A) polymerase from Saccharomyces cerevisiae
J. Biol. Chem.
266
8741-8746
1991
Saccharomyces cerevisiae
Manually annotated by BRENDA team
Bard, J.; Zhelkovsky, A.M.; Helmling, S.; Earnest, T.N.; Moore, C.L.; Bohm, A.
Structure of yeast poly(A) polymerase alone and in complex with 3'-dATP
Science
289
1346-1349
2000
Saccharomyces cerevisiae
Manually annotated by BRENDA team
Sillero, M.A.; de Diego, A.; Silles, E.; Osorio, H.; Sillero, A.
Polyphosphates strongly inhibit the tRNA dependent synthesis of poly(A) catalyzed by poly(A) polymerase from Saccharomyces cerevisiae
FEBS Lett.
550
41-45
2003
Saccharomyces cerevisiae
Manually annotated by BRENDA team
Hooker, L.; Strong, R.; Adams, R.; Handa, B.; Merrett, J.H.; Martin, J.A.; Klumpp, K.
A sensitive, single-tube assay to measure the enzymatic activities of influenza RNA polymerase and other poly(A) polymerases: application to kinetic and inhibitor analysis
Nucleic Acids Res.
29
2691-2698
2001
Saccharomyces cerevisiae
Manually annotated by BRENDA team
Zhelkovsky, A.M.; Kessler, M.M.; Moore, C.L.
Structure-function relationships in the Saccharomyces cerevisiae poly(A) polymerase. Identification of a novel RNA binding site and a domain that interacts with specificity factor(s)
J. Biol. Chem.
270
26715-26720
1995
Saccharomyces cerevisiae
Manually annotated by BRENDA team
Sillero, M.A.; De Diego, A.; Osorio, H.; Sillero, A.
Dinucleoside polyphosphates stimulate the primer independent synthesis of poly(A) catalyzed by yeast poly(A) polymerase
Eur. J. Biochem.
269
5323-5329
2002
Saccharomyces cerevisiae
Manually annotated by BRENDA team
Balbo, P.B.; Meinke, G.; Bohm, A.
Kinetic studies of yeast polyA polymerase indicate an induced fit mechanism for nucleotide specificity
Biochemistry
44
7777-7786
2005
Saccharomyces cerevisiae
Manually annotated by BRENDA team
Chen, L.S.; Sheppard, T.L.
Chain termination and inhibition of Saccharomyces cerevisiae poly(A) polymerase by C-8-modified ATP analogs
J. Biol. Chem.
279
40405-40411
2004
Saccharomyces cerevisiae
Manually annotated by BRENDA team
Zhelkovsky, A.; Helmling, S.; Bohm, A.; Moore, C.
Mutations in the middle domain of yeast poly(A) polymerase affect interactions with RNA but not ATP
RNA
10
558-564
2004
Saccharomyces cerevisiae
Manually annotated by BRENDA team
Houseley, J.; Tollervey, D.
Yeast Trf5p is a nuclear poly(A) polymerase
EMBO Rep.
7
205-211
2006
Saccharomyces cerevisiae
Manually annotated by BRENDA team
Balbo, P.B.; Bohm, A.
Mechanism of poly(A) polymerase: Structure of the enzyme-MgATP-RNA ternary complex and kinetic analysis
Structure
15
1117-1131
2007
Saccharomyces cerevisiae (P29468), Saccharomyces cerevisiae
Manually annotated by BRENDA team
Meinke, G.; Ezeokonkwo, C.; Balbo, P.; Stafford, W.; Moore, C.; Bohm, A.
Structure of yeast poly(A) polymerase in complex with a peptide from Fip1, an intrinsically disordered protein
Biochemistry
47
6859-6869
2008
Saccharomyces cerevisiae (P29468), Saccharomyces cerevisiae
Manually annotated by BRENDA team
Balbo, P.B.; Toth, J.; Bohm, A.
X-ray crystallographic and steady state fluorescence characterization of the protein dynamics of yeast polyadenylate polymerase
J. Mol. Biol.
366
1401-1415
2007
Saccharomyces cerevisiae (P29468), Saccharomyces cerevisiae
Manually annotated by BRENDA team
Mellman, D.L.; Gonzales, M.L.; Song, C.; Barlow, C.A.; Wang, P.; Kendziorski, C.; Anderson, R.A.
A PtdIns4,5P2-regulated nuclear poly(A) polymerase controls expression of select mRNAs
Nature
451
1013-1017
2008
Saccharomyces cerevisiae
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
Ezeokonkwo, C.; Ghazy, M.A.; Zhelkovsky, A.; Yeh, P.C.; Moore, C.
Novel interactions at the essential N-terminus of poly(A) polymerase that could regulate poly(A) addition in Saccharomyces cerevisiae
FEBS Lett.
586
1173-1178
2012
Saccharomyces cerevisiae (P29468), Saccharomyces cerevisiae
Manually annotated by BRENDA team
Losh, J.S.; King, A.K.; Bakelar, J.; Taylor, L.; Loomis, J.; Rosenzweig, J.A.; Johnson, S.J.; van Hoof, A.
Interaction between the RNA-dependent ATPase and poly(A) polymerase subunits of the TRAMP complex is mediated by short peptides and important for snoRNA processing
Nucleic Acids Res.
43
1848-1858
2015
Saccharomyces cerevisiae (P48561)
Manually annotated by BRENDA team