Information on EC 5.4.99.27 - tRNA pseudouridine13 synthase

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

EC NUMBER
COMMENTARY
5.4.99.27
-
RECOMMENDED NAME
GeneOntology No.
tRNA pseudouridine13 synthase
-
REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
tRNA uridine13 = tRNA pseudouridine13
show the reaction diagram
-
-
-
-
tRNA uridine13 = tRNA pseudouridine13
show the reaction diagram
a strictly conserved glutamate near the active site in TruD is likely to act as a general base for the proton abstraction at C5 of the uridine base
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SYSTEMATIC NAME
IUBMB Comments
tRNA-uridine13 uracil mutase
Pseudouridine synthase TruD from Escherichia coli specifically acts on uridine13 in tRNA [2,3]. The Pus7 protein from Saccharomyces cerevisiae is a multisite-multisubstrate pseudouridine synthase that is able to modify uridine13 in several yeast tRNAs, uridine35 in the pre-tRNATyr, uridine35 in U2 small nuclear RNA, and uridine50 in 5S rRNA [5].
SYNONYMS
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
Pus7 protein
Q08647
-
RNA:pseudouridine-synthase Pus7p
Q08647
-
RNA:PSI-synthase Pus7p
Q08647
-
tRNA PSI13 synthase
Q57261
-
YgbO
Q57261
-
CAS REGISTRY NUMBER
COMMENTARY
430429-15-5
-
61506-89-6
-
ORGANISM
COMMENTARY
LITERATURE
SEQUENCE CODE
SEQUENCE DB
SOURCE
GENERAL INFORMATION
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
evolution
-
TruD folds into a V-shaped molecule with a catalytic domain that is structurally very similar to the catalytic modules of the other known pseudouridine synthases despite its lack of sequence homology and likely arose by divergent evolution
malfunction
-
disruption of the PUS7 gene abolishes pseudouridine13 formation in tRNAs in vivo. Absence of pseudouridine13 formation in tRNAAsp and tRNAGlu extracted from the DELTAPUS7 strain
malfunction
-
deletion of the ygbO gene causes the loss of tRNAGlu pseudouridine13 and plasmid-borne restoration of the structural gene restores pseudouridine13. Growth competition does not show any effect of the deletion
SUBSTRATE
PRODUCT                      
REACTION DIAGRAM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
(Substrate)
LITERATURE
(Substrate)
COMMENTARY
(Product)
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
tRNA uridine13
tRNA pseudouridine13
show the reaction diagram
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-
-
-
?
tRNA uridine13
tRNA pseudouridine13
show the reaction diagram
-
Pus7p is a multisite-multisubstrate RNA:PSI-synthase. Recombinant Pus7p has tRNAAsp:pseudouridine13- and pre-RNATyr:pseudouridine35-synthase activities
-
-
?
tRNA uridine13
tRNA pseudouridine13
show the reaction diagram
Q08647, -
the multisite-specific enzyme Pus7p is able to modify uridine13 in several yeast tRNAs, uridine35 in the pre-tRNATyr, uridine35 in U2 small nuclear RNA, and uridine50 in 5S rRNA. Using site-directed mutagenesis, a set of RNA variants derived from the yeast tRNAAsp(GUC), pretRNA Tyr(G(pseudouridine)A), and U2 small nuclear RNA is designed and the ability to be modified by Pus7p in vitro is tested. Pseudouridine13 formation in tRNAAsp(GUC). Does not depend on correct tRNA three-dimensional structure but is increased by the presence of RNA helices
-
-
?
KM VALUE [mM]
KM VALUE [mM] Maximum
SUBSTRATE
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
IMAGE
0.00038
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tRNA uridine13
-
-
-
TURNOVER NUMBER [1/s]
TURNOVER NUMBER MAXIMUM[1/s]
SUBSTRATE
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
IMAGE
0.00097
-
tRNA uridine13
-
-
-
kcat/KM VALUE [1/mMs-1]
kcat/KM VALUE [1/mMs-1] Maximum
SUBSTRATE
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
IMAGE
0.255
-
tRNA uridine13
-
-
0
pH OPTIMUM
pH MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
PDB
SCOP
CATH
ORGANISM
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Methanosarcina mazei (strain ATCC BAA-159 / DSM 3647 / Goe1 / Go1 / JCM 11833 / OCM 88)
SUBUNITS
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
?
-
x * 39100, calculated from sequence, SDS-PAGE
Crystallization/COMMENTARY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
crystal structure of Escherichia coli TruD at 2.0 A resolution. The structure reveals an overall V-shaped molecule with an RNA binding cleft formed between two domains: a catalytic domain and an insertion domain
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crystals of TruD are grown at room temperature by the hanging drop vapor diffusion method. TruD folds into a V-shaped molecule with a catalytic domain that is structurally very similar to the catalytic modules of the other known pseudouridine synthases despite its lack of sequence homology and likely arose by divergent evolution
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hanging drop vapor diffusion method, 2.2 A resolution structure. TruD reveals a U-shaped molecule with a catalytic domain that superimposes closely on that of other pseudouridine synthases
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sitting-drop vapour-diffusion method, crystals diffract to a minimum Bragg spacing of 2.4 A and belong to space group P212121, with unit-cell parameters a = 63.4, b = 108.6, c = 111.7 A
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Purification/COMMENTARY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
Cloned/COMMENTARY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
EXPRESSION
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
the mutated enzyme is not able to form residue pseudouridine13 in tRNAAsp and tRNAGlu in vivo
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ENGINEERING
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
E31D
-
27% of wild-type activity
E31Q
-
no activity
E31Q/F131Y
-
no activity
F131Y
-
68% of wild-type activity
K79L
-
5% of wild-type activity; no activity
K79L/F131Y
-
no activity
K79L/N129K/F131Y
-
no activity
K79R
-
10% of wild-type activity
N129K
-
no activity
Q87E
-
29% of wild-type activity