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Information on EC 5.4.99.B22 - multisite-specific tRNA pseudouridine synthase and Organism(s) Homo sapiens and UniProt Accession Q9Y606
for references in articles please use BRENDA:EC5.4.99.B22preliminary BRENDA-supplied EC number
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EC Tree
5.4.99.B22 multisite-specific tRNA pseudouridine synthaseSpecify your search results
Word Map
- 5.4.99.B22
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pseudouridylation
- anemia
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sideroblastic
- myopathy
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dyskerin
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anticodon
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mlasa
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spliceosomal
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nuclear-encoded
- agriculture
The taxonomic range for the selected organisms is: Homo sapiens
The expected taxonomic range for this enzyme is: Eukaryota, Bacteria, Archaea
The expected taxonomic range for this enzyme is: Eukaryota, Bacteria, Archaea
Reaction Schemes
hide(3 overall reactions are displayed. Show all (9)>>)
tRNA uridine65
=
tRNA pseudouridine65
tRNA uridine26
=
tRNA pseudouridine26
tRNA uridine27
=
tRNA pseudouridine27
Synonyms
pus1p, mpus1p, pseudouridine synthase 1, rpusd4, hpus1p, scpus1p, sppus1p, tgpus1, 
more
topSYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
SYSTEMATIC NAME
IUBMB Comments
multisite-specific tRNA-uridine uracil mutase
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SUBSTRATE
PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
yeast pre-tRNAIle uridine
yeast pre-tRNAIle pseudouridine
possible modification sites are uridine 27, uridine30, uridine34 or uridine36
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mitochondrial tRNAPhe uridine39
mitochondrial tRNAPhe pseudouridine39
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?
additional information
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Pus1p modifies uridines at positions 1, 26, 27, 28, 30, 34, 36, 65, and 67 depending on the source of the enzyme (Saccharomyces cerevisiae or mouse), the substrates (intron-containing or not), or whether the activity is monitored in vivo or in vitro. Out of all the positions that Pus1p modifies, the modification of uridines at positions 27 and 28 is by far the most common in tRNAs, with pseudouridine infrequently found at the other positions that the Pus1p enzymes recognize
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additional information
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Pus1p modifies uridines at positions 1, 26, 27, 28, 30, 34, 36, 65, and 67 depending on the source of the enzyme (Saccharomyces cerevisiae or mouse), the substrates (intron-containing or not), or whether the activity is monitored in vivo or in vitro. Out of all the positions that Pus1p modifies, the modification of uridines at positions 27 and 28 is by far the most common in tRNAs, with pseudouridine infrequently found at the other positions that the Pus1p enzymes recognize
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additional information
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pseudouridine synthase assays of hPus1p enzymes tRNA and HP7 SRA RNA substrates, i.e. RNA of the minimal RNA fragment within steroid receptor RNA activator (SRA), termed HP7. Mouse mitochondrial tRNAAsp is used as a positive control for hPus1p activity
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?
additional information
?
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pseudouridine synthase assays of hPus1p enzymes tRNA and HP7 SRA RNA substrates, i.e. RNA of the minimal RNA fragment within steroid receptor RNA activator (SRA), termed HP7. Mouse mitochondrial tRNAAsp is used as a positive control for hPus1p activity
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NATURAL SUBSTRATE
NATURAL PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
mitochondrial tRNAPhe uridine39
mitochondrial tRNAPhe pseudouridine39
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?
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.000032
yeast pre-tRNAIle uridine
possible modification sites are uridine 27, uridine30, uridine34 or uridine36, wild-type enzyme, pH and temperature not specified in the publication
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0.000168
yeast pre-tRNAIle uridine
possible modification sites are uridine 27, uridine30, uridine34 or uridine36, mutant enzyme R116K, pH and temperature not specified in the publication
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ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
SOURCE
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY
GeneOntology No.
LITERATURE
SOURCE
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
malfunction
missense mutation in PUS1 causes mitochondrial myopathy and sideroblastic anemia (a rare, autosomal recessive oxidative phosphorylation disorder specific to skeletal muscle and bone marrow)
metabolism
the enzyme modulate class I and class II nuclear receptor responses through its ability to modify the steroid receptor RNA activator
evolution
eukaryal Pus10 genes share a conserved catalytic domain with archaeal Pus10 genes. Pus10 is found in earlier evolutionary branches of fungi (such as chytrid Batrachochytrium) but is absent in all dikaryon fungi surveyed (Ascomycetes and Basidiomycetes). Orthologs of Pus10, TrmA, and TruB are present in all the animals, plants, and protozoa surveyed. This indicates that the common eukaryotic ancestor possesses all the three genes. Pus10 exists as a single copy gene in all the surveyed genomes despite ancestral whole genome duplications has occurred. This indicates a possible deleterious gene dosage effect. Functional redundancy results in gene loss or neofunctionalization in different evolutionary lineages
malfunction
depletion of RPUSD4 leads to a severe reduction of the steady-state level of the 16S mitochondrial rRNA with defects in the biogenesis of the mitoribosome large subunit and consequently in mitochondrial translation
metabolism
additional information
metabolism
human Pus10 participates in apoptosis induced by the tumor necrosis factor-related apoptosis-inducing ligand
metabolism
pseudouridine synthase RPUSD4 is an essential component of mitochondrial RNA granules. Several steps of mitochondrial RNA processing and maturation, including RNA post-transcriptional modification, are spatially organized into mitochondrial RNA granules
additional information
hPus1p active site architecture analysis, overview
UNIPROT
ENTRY NAME
ORGANISM
NO. OF AA
NO. OF TRANSM. HELICES
MOLECULAR WEIGHT[Da]
SOURCE
SEQUENCE
LOCALIZATION PREDICTION
The reliability class of the localization prediction ranges from 1 (very reliable) to 5 (not reliable).
The reliability class of the localization prediction ranges from 1 (very reliable) to 5 (not reliable). SUBUNIT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
additional information
additional information
the hPus1p enzyme binds to its RNA substrate as a monomer. The C-terminal end of hPus1p folds into two alpha-helices and these helices pack against the back of the hPus1p molecule, which prevents the dimerization observed in the bacterial homologue, TruA. This C-terminal region is responsible for the atypical RNA substrate binding and activity of hPus1p. Enzyme crystal structure analysis, overview
additional information
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the hPus1p enzyme binds to its RNA substrate as a monomer. The C-terminal end of hPus1p folds into two alpha-helices and these helices pack against the back of the hPus1p molecule, which prevents the dimerization observed in the bacterial homologue, TruA. This C-terminal region is responsible for the atypical RNA substrate binding and activity of hPus1p. Enzyme crystal structure analysis, overview
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
purified recombinant detagged truncated enzyme mutant . DELTAhPus1p and mutant D146A DELTAhPus1p, sitting drop technique, mixing of 7 mg/ml protein in 20 mM HEPES, pH 7.0, 150 mM NaCl, 5 mM MgCl2, and 5 mM TCEP, with an equal amount of reservoir solution containing for DELTAhPus1p crystals 28% PEG 3350/glycerol, 0.1 M Bicine/Tris, pH 8.5, 10% amino acids mixture (with 0.02 M sodium L-glutamate, 0.02 M DL-alanine, 0.02 M glycine, 0.02 M DL-lysine HCl, 0.02 M DL-serine), and for D146A DELTAhPus1p crystals 17% PEG 8000, and 0.1 M HEPES, pH 8.0, both at 4°C, for 2-3 days, X-ray diffraction structure determination and analysis at 2.0-2.7 A resolution, respectively, molecular replacement using Pus1p structure, PDB ID 4ITS, as a model
sitting drop technique. The crystal structure of the catalytic domain of hPus1p and the D146A mutant of the enzyme is determined at 2.0 A resolution, alone and in a complex with several molecules present during crystallisation
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
D146A
site-directed mutagenesis of truncated enzyme mutant DELTAhPus1p, differences between structures of DELTAhPus1p and DELTAhPus1pD146A are limited to external, poorly conserved loops, which confirm the flexibility of these regions
R116A
the mutant exhibits some activity with the yeast pre-tRNAIle substrate. Low activity with the mouse tRNAMet substrate
R116C
the activity of the mutant enzyme with the yeast pre-tRNAIle substrate is approximately one-third that of the wild-type levels. No activity with the mouse tRNAMet substrate
R116E
the mutant is inactive with with the yeast pre-tRNAIle substrate. No activity with the mouse tRNAMet substrate
R116G
the mutant exhibits some activity with the yeast pre-tRNAIle substrate. No activity with the mouse tRNAMet substrate
R116H
the mutant is inactive with with the yeast pre-tRNAIle substrate. Low activity with the mouse tRNAMet substrate
R116K
the activity of the mutant enzyme with the yeast pre-tRNAIle substrate is approximately one-third that of the wild-type levels
R116N
the mutant exhibits some activity with the yeast pre-tRNAIle substrate. No activity with the mouse tRNAMet substrate
R116Q
the mutant exhibits some activity with the yeast pre-tRNAIle substrate. No activity with the mouse tRNAMet substrate
R116S
the mutant exhibits some activity with the yeast pre-tRNAIle substrate. No activity with the mouse tRNAMet substrate
R116W
the mutant is inactive with with the yeast pre-tRNAIle substrate. Low activity with the mouse tRNAMet substrate
Y173C
mutant enzyme retains some activity with yeast pre-tRNAIle. The activity is relatively low
Y173F
Y173G
mutant enzyme retains some activity with yeast pre-tRNAIle. The activity is relatively low
Y173T
additional information
Y173F
mutant enzyme retains some activity with yeast pre-tRNAIle. The activity is relatively low
Y173T
mutant enzyme retains some activity with yeast pre-tRNAIle. The activity is relatively low
additional information
generation of a truncated hPus1p version encompassing residues 83-394, i.e. DELTAhPus1p. The deleted N- and C-termini are not essential for either the RNA binding or the activity of the hPus1p enzyme
additional information
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generation of a truncated hPus1p version encompassing residues 83-394, i.e. DELTAhPus1p. The deleted N- and C-termini are not essential for either the RNA binding or the activity of the hPus1p enzyme
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
recombinant His-tagged wild-type and mutant enzymes from Escherichia coli strain BL21(DE3) by nickel affinity chromatography, tag cleavage by TEV protease, followed by gel filtration, to hoomogeneity
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
gene PUS1, recombinant expression of His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3)
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Bykhovskaya, Y.; Casas, K.; Mengesha, E.; Inbal, A.; Fischel-Ghodsian, N.
Missense mutation in pseudouridine synthase 1 (PUS1) causes mitochondrial myopathy and sideroblastic anemia (MLASA)
Am. J. Hum. Genet.
74
1303-1308
2004
Homo sapiens (Q9Y606), Homo sapiens
Sibert, B.S.; Fischel-Ghodsian, N.; Patton, J.R.
Partial activity is seen with many substitutions of highly conserved active site residues in human Pseudouridine synthase 1
RNA
14
1895-1906
2008
Homo sapiens (Q9Y606), Homo sapiens
Zaganelli, S.; Rebelo-Guiomar, P.; Maundrell, K.; Rozanska, A.; Pierredon, S.; Powell, C.A.; Jourdain, A.A.; Hulo, N.; Lightowlers, R.N.; Chrzanowska-Lightowlers, Z.M.; Minczuk, M.; Martinou, J.C.
The pseudouridine synthase RPUSD4 is an essential component of mitochondrial RNA granules
J. Biol. Chem.
292
4519-4532
2017
Homo sapiens (Q96CM3)
Fitzek, E.; Joardar, A.; Gupta, R.; Geisler, M.
Evolution of eukaryal and archaeal pseudouridine synthase Pus10
J. Mol. Evol.
86
77-89
2018
Arabidopsis thaliana (F4HSS8), Homo sapiens (Q3MIT2), Methanocaldococcus jannaschii (Q60346), Pyrococcus furiosus (Q8U1R6)
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