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Information on EC 5.4.99.23 - 23S rRNA pseudouridine1911/1915/1917 synthase and Organism(s) Escherichia coli and UniProt Accession P33643

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IUBMB Comments
Pseudouridine synthase RluD converts uridines at positions 1911, 1915, and 1917 of 23S rRNA to pseudouridines. These nucleotides are located in the functionally important helix-loop 69 of 23S rRNA .
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Escherichia coli
UNIPROT: P33643
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The taxonomic range for the selected organisms is: Escherichia coli
The expected taxonomic range for this enzyme is: Bacteria, Archaea, Eukaryota
Reaction Schemes
23S rRNA uridine1911/uridine1915/uridine1917
=
23S rRNA pseudouridine1911/pseudouridine1915/pseudouridine1917
Synonyms
pseudouridine synthase rlud, rlud pseudouridine synthase, more
SYNONYM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
pseudouridine synthases RluD
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ribosomal large subunit pseudouridine synthase D
UniProt
pseudouridine synthase RluD
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RluD pseudouridine synthase
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SYSTEMATIC NAME
IUBMB Comments
23S rRNA-uridine1911/1915/1917 uracil mutase
Pseudouridine synthase RluD converts uridines at positions 1911, 1915, and 1917 of 23S rRNA to pseudouridines. These nucleotides are located in the functionally important helix-loop 69 of 23S rRNA [1].
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
23S rRNA uridine1911/uridine1915/uridine1917
23S rRNA pseudouridine1911/pseudouridine1915/pseudouridine1917
show the reaction diagram
23S rRNA uridine1911/uridine1915/uridine1917
23S rRNA pseudouridine1911/pseudouridine1915/pseudouridine1917
show the reaction diagram
additional information
?
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the recombinant chimeric enzyme mutant RluCD isomerizes many uridines of rRNA in a non-specific manner
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-
?
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
23S rRNA uridine1911/uridine1915/uridine1917
23S rRNA pseudouridine1911/pseudouridine1915/pseudouridine1917
show the reaction diagram
23S rRNA uridine1911/uridine1915/uridine1917
23S rRNA pseudouridine1911/pseudouridine1915/pseudouridine1917
show the reaction diagram
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
evolution
the structure of RluD (a RluA family member) emphasizes that the RluA, RsuA, TruB, and TruA families of pseudouridine synthases arose by divergent evolution from a common ancestor
malfunction
physiological function
RluD is a ribosomal assembly factor that may be involved in the late stages of maturation of the large ribosomal subunit
malfunction
physiological function
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
44000
x * 44000, SDS-PAGE
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
?
x * 44000, SDS-PAGE
monomer
analytical ultracentrifugation sedimentation velocity experiments show that RluD is monomeric in solution
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
2.0 A structure of the catalytic domain of RluD (residues 77–326). The catalytic domain folds into a mainly antiparallel beta-sheet flanked by several loops and helices. A positively charged cleft that presumably binds RNA leads to the conserved Asp139. The RluD N-terminal S4 domain, connected by a flexible linker, is disordered in our structure. RluD is very similar in both catalytic domain structure and active site arrangement to the pseudouridine synthases RsuA, TruB, and TruA
crystals of full-length RluD are grown at 20°C using the hanging drop method. The S4 domain of RluD appears to be highly flexible or unfolded and is completely invisible in the electron density map
crystallization of selenomethionine-substituted RluD by the hanging-drop method, crystals diffract to 1.9 A and belong to space group P4(3)2(1)2, with unit cell parameters a = b = 75.14, c = 181.81 A
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crystals of SeMet-labeled DELTARluD are obtained under oil by the microbatch method, crystal structure of the catalytic module of RluD (residues 68–326, DELTARluD) refined at 1.8 A to a final R-factor of 21.8%. DELTARluD is a monomeric enzyme having an overall mixed alpha/beta fold. The DELTARluD molecule consists of two subdomains, a catalytic subdomain and C-terminal subdomain with the RNA-binding cleft formed by loops extending from the catalytic sub-domain. Comparison of the structure with other pseudouridine synthases
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
D139N
D139T
mutation completely blocks pseudouridine formation in vivo and in vitro. The mutant rluD gene produces a protein capable of complete reversal of the growth defect (of Escherichia coli mutant with a truncation in the rluD gene) without concomitant pseudouridine formation
additional information
construction of a chimeric pseudouridine synthase (RluCD) containing the N-terminal S4 domain of enzyme RluC (EC 5.4.99.24) and the C-terminal catalytic domain of enzyme RluD (EC 5.4.99.23). The chimeric mutant is able to introduce excessive pseudouridines into rRNA at non-native positions. The chimeric enzyme RluCD is used as a tool to study an effect of over-modification of rRNA on the ribosome biogenesis. Excessive pseudouridylation of 23S rRNA reduces progression of ribosome assembly during early or middle stages. A modification interference approach identifies the sites in 23S rRNA whose modification prevents ribosome assembly. It is plausible that pseudouridines can cause RNA misfolding when present at non-native positions. RluCD isomerizes many uridines of rRNA in a non-specific manner. Induction of the RluCD at the exponential growth phase leads to severe inhibition of translation while transcription is only slightly affected
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
recombinant expression of chimeric enzyme mutants RluCD in Escherichia coli strains M15 of CD204, expression of chimeric pseudouridine synthase RluCD, under control of arabinose inducible araBAD promoter, interferes with ribosome formation in wild-type M15 strain and strain CD204 lacking RluC and RluD, while cells transformed with catalytically inactive RluCD (RluCD D139N) and empty vector (pQE60), used as a controls, represent a normal ribosomal profile in sucrose density gradient. Strain CD204 contains a mutant form of RF-2 (D131Y) that suppresses ribosome assembly defect caused by the deletion of the RluD. Strain rluD114 where yfiI (RluD) gene has been disrupted with Km cassette is used as a parent strain to construct strain CD204
wild-type and mutant enzymes D139T and D139N are overexpressed from plasmids
overexpressed in Escherichia coli
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REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Del Campo, M.; Ofengand, J.; Malhotra, A.
Purification and crystallization of Escherichia coli pseudouridine synthase RluD
Acta Crystallogr. Sect. D
59
1871-1873
2003
Escherichia coli
Manually annotated by BRENDA team
Sivaraman, J.; Iannuzzi, P.; Cygler, M.; Matte, A.
Crystal structure of the RluD pseudouridine synthase catalytic module, an enzyme that modifies 23S rRNA and is essential for normal cell growth of Escherichia coli
J. Mol. Biol.
335
87-101
2004
Escherichia coli (Q8X9F0), Escherichia coli
Manually annotated by BRENDA team
Gutgsell, N.S.; Deutscher, M.P.; Ofengand, J.
The pseudouridine synthase RluD is required for normal ribosome assembly and function in Escherichia coli
RNA
11
1141-1152
2005
Escherichia coli
Manually annotated by BRENDA team
Leppik, M.; Peil, L.; Kipper, K.; Liiv, A.; Remme, J.
Substrate specificity of the pseudouridine synthase RluD in Escherichia coli
FEBS J.
274
5759-5766
2007
Escherichia coli
Manually annotated by BRENDA team
Huang, L.; Ku, J.; Pookanjanatavip, M.; Gu, X.; Wang, D.; Greene, P.J.; Santi, D.V.
Identification of two Escherichia coli pseudouridine synthases that show multisite specificity for 23S RNA
Biochemistry
37
15951-15957
1998
Escherichia coli
Manually annotated by BRENDA team
Mizutani, K.; Machida, Y.; Unzai, S.; Park, S.Y.; Tame, J.R.
Crystal structures of the catalytic domains of pseudouridine synthases RluC and RluD from Escherichia coli
Biochemistry
43
4454-4463
2004
Escherichia coli (P33643)
Manually annotated by BRENDA team
Wrzesinski, J.; Bakin, A.; Ofengand, J.; Lane, B.G.
Isolation and properties of Escherichia coli 23S-RNA pseudouridine 1911, 1915, 1917 synthase (RluD)
IUBMB Life
50
33-37
2000
Escherichia coli (P33643)
Manually annotated by BRENDA team
Ejby, M.; Sorensen, M.A.; Pedersen, S.
Pseudouridylation of helix 69 of 23S rRNA is necessary for an effective translation termination
Proc. Natl. Acad. Sci. USA
104
19410-19415
2007
Escherichia coli
Manually annotated by BRENDA team
Del Campo, M.; Ofengand, J.; Malhotra, A.
Crystal structure of the catalytic domain of RluD, the only rRNA pseudouridine synthase required for normal growth of Escherichia coli
RNA
10
231-239
2004
Escherichia coli (P33643), Escherichia coli
Manually annotated by BRENDA team
Vaidyanathan, P.P.; Deutscher, M.P.; Malhotra, A.
RluD, a highly conserved pseudouridine synthase, modifies 50S subunits more specifically and efficiently than free 23S rRNA
RNA
13
1868-1876
2007
Escherichia coli (P33643), Escherichia coli
Manually annotated by BRENDA team
Raychaudhuri, S.; Conrad, J.; Hall, B.G.; Ofengand, J.
A pseudouridine synthase required for the formation of two universally conserved pseudouridines in ribosomal RNA is essential for normal growth of Escherichia coli
RNA
4
1407-1417
1998
Escherichia coli (P33643)
Manually annotated by BRENDA team
Gutgsell, N.S.; Del Campo, M.; Raychaudhuri, S.; Ofengand, J.
A second function for pseudouridine synthases: A point mutant of RluD unable to form pseudouridines 1911, 1915, and 1917 in Escherichia coli 23S ribosomal RNA restores normal growth to an RluD-minus strain
RNA
7
990-998
2001
Escherichia coli (P33643)
Manually annotated by BRENDA team
Leppik, M.; Liiv, A.; Remme, J.
Random pseuoduridylation in vivo reveals critical region of Escherichia coli 23S rRNA for ribosome assembly
Nucleic Acids Res.
45
6098-6108
2017
Escherichia coli (P33643)
Manually annotated by BRENDA team