Information on EC 5.4.99.27 - tRNA pseudouridine13 synthase

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

EC NUMBER
COMMENTARY hide
5.4.99.27
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RECOMMENDED NAME
GeneOntology No.
tRNA pseudouridine13 synthase
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REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
tRNA uridine13 = tRNA pseudouridine13
show the reaction diagram
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].
CAS REGISTRY NUMBER
COMMENTARY hide
430429-15-5
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61506-89-6
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
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Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
evolution
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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
metabolism
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Pus7p that catalyses U2 pseudouridylation at position 35, also catalyses pseudouridylation at position 56 during nutrient deprivation or heat shock
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
Pyrococcus abyssi tRNAAsp(GUA) uridine13
Pyrococcus abyssi tRNAAsp(GUA) pseudouridine13
show the reaction diagram
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?
Pyrococcus abyssi tRNATyr(GUA) uridine13
Pyrococcus abyssi tRNATyr(GUA) pseudouridine13
show the reaction diagram
Pyrococcus abyssi tRNATyr(GUA) uridine35
Pyrococcus abyssi tRNATyr(GUA) pseudouridine35
show the reaction diagram
tRNA uridine13
tRNA pseudouridine13
show the reaction diagram
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
Pyrococcus abyssi tRNATyr(GUA) uridine35
Pyrococcus abyssi tRNATyr(GUA) pseudouridine35
show the reaction diagram
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residue 35 reinforces the stability of the anticodon stemloop by interaction with residue U33 (13). This structural stabilization may increase the aminoacylation efficiency of tRNATyr(GUA) by its cognate aminoacyl-tRNA synthetase
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?
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.00038
tRNA uridine13
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TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.00097
tRNA uridine13
Escherichia coli
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kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.255
tRNA uridine13
Escherichia coli
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27242
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
PDB
SCOP
CATH
ORGANISM
UNIPROT
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
?
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x * 39100, calculated from sequence, SDS-PAGE
Crystallization/COMMENTARY
ORGANISM
UNIPROT
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
LITERATURE
wild-type and mutant enzymes produced in Escherichia coli as His6-tagged protein fusions
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
wild-type and mutant enzymes are produced in Escherichia coli as His6-tagged protein fusions
EXPRESSION
ORGANISM
UNIPROT
LITERATURE
the mutated enzyme is not able to form residue pseudouridine13 in tRNAAsp and tRNAGlu in vivo
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
E31D
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27% of wild-type activity
E31Q
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no activity
E31Q/F131Y
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no activity
F131Y
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68% of wild-type activity
K79L
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5% of wild-type activity; no activity
K79L/F131Y
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no activity
K79L/N129K/F131Y
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no activity
K79R
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10% of wild-type activity
N129K
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no activity
Q87E
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29% of wild-type activity
K19I
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mutation decreases the activity at position 13 in the Pyrococcus abyssi tRNAAsp at 55C. At 80C the mutation has almost no negative effect on its activity towards the Pyrococcus abyssi tRNATyr(GUA)
K19I/R78A/H79N
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mutation completely abolishes the activity at position 13 in the Pyrococcus abyssi tRNAAsp at 55C. At 80C the mutation nearly abolishes activity towards the Pyrococcus abyssi tRNATyr(GUA)
R78A/H79N
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mutation decreases the activity at position 13 in the Pyrococcus abyssi tRNAAsp at 55C. At 80C the mutation has almost no negative effect on its activity towards the Pyrococcus abyssitRNATyr(GUA)
A78R/N79H
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mutation has no positive effect on activity at 80C on Pyrococcus abyssi tRNAAsp(GUA)
I27K
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mutation increases activity at 80C on Pyrococcus abyssi tRNAAsp(GUA) 1.6fold
N79H
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mutation has no positive effect on activity at 80C on Pyrococcus abyssi tRNAAsp(GUA)
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