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Escherichia coli K-12 substr. MG1655 Enzyme: tRNA pseudouridine synthase I



Gene: truA Accession Numbers: EG10454 (EcoCyc), b2318, ECK2312

Synonyms: asuC, hisT, leuK

Regulation Summary Diagram: ?

Subunit composition of tRNA pseudouridine synthase I = [TruA]2
         tRNA pseudouridine synthase I = TruA

Summary:
TruA is the tRNA pseudouridine synthase responsible for catalyzing pseudouridine formation in the anticodon loop of a subset of tRNAs [Kammen88]. TruA-mediated pseudouridylation contributes to structural stability of this region of the tRNA [Davis91] and reading frame maintenance [Urbonavicius01].

The reaction mechanism is discussed [Kammen88]. Results from site-directed mutagenesis indicate that the rearrangement mechanism likely does not involve a covalent cysteine intermediate [Zhao97]. Instead, the D60 residue was shown to be essential for catalytic activity [Huang98a], forming a covalent adduct that can undergo O-acyl hydrolytic cleavage to form the pseudouridine product [Gu99].

Crystal structures of the enzyme have been solved [Foster00, Hur07]. TruA is a dimer in the crystal as well as in solution [Foster00]. Modeling of the substrate recognition pathway indicates that the enzyme utilizes the intrinsic flexibility of the tRNA anticodon loop [Hur07].

A truA mutant exhibits a tRNA pseudouridine synthase defect [Bruni77] and increased translation errors under conditions of low histidine [Parker82]. A mutant exhibits a growth defect that is rescued by addition of adenine, uracil, and isoleucine to the growth medium [Bruni77]. A truA insertion mutant has a severe growth defect in minimal medium which can be rescued by addition of uracil [Tsui91a].

truA is encoded in an operon downstream of usg, a gene of unknown function [Arps85, Arps87]. Expression of truA is growth rate-regulated and increases in stationary phase [Tsui91a].

AsuC: "antisuppressor" [Sullivan85]

Reviews: [Hamma06, Hur06]

Gene Citations: [Schoenlein89, Arps87a, Pease02]

Locations: cytosol

Map Position: [2,432,846 <- 2,433,658] (52.44 centisomes)
Length: 813 bp / 270 aa

Molecular Weight of Polypeptide: 30.4 kD (from nucleotide sequence), 31 kD (experimental) [Marvel85 ]

Molecular Weight of Multimer: 66.0 kD (experimental) [Foster00]

Unification Links: ASAP:ABE-0007657 , CGSC:623 , DIP:DIP-11043N , EchoBASE:EB0449 , EcoGene:EG10454 , EcoliWiki:B2318 , Mint:MINT-1310146 , ModBase:P07649 , OU-Microarray:b2318 , PortEco:truA , Pride:P07649 , Protein Model Portal:P07649 , RefSeq:NP_416821 , RegulonDB:EG10454 , SMR:P07649 , String:511145.b2318 , UniProt:P07649

Relationship Links: InterPro:IN-FAMILY:IPR001406 , InterPro:IN-FAMILY:IPR020094 , InterPro:IN-FAMILY:IPR020095 , InterPro:IN-FAMILY:IPR020097 , InterPro:IN-FAMILY:IPR020103 , Panther:IN-FAMILY:PTHR11142 , PDB:Structure:1DJ0 , PDB:Structure:2NQP , PDB:Structure:2NR0 , PDB:Structure:2NRE , Pfam:IN-FAMILY:PF01416

Gene-Reaction Schematic: ?

GO Terms:

Biological Process: GO:0031119 - tRNA pseudouridine synthesis Inferred from experiment Inferred by computational analysis [GOA06, Lawther77]
GO:0001522 - pseudouridine synthesis Inferred by computational analysis [GOA01]
GO:0008033 - tRNA processing Inferred by computational analysis [UniProtGOA11]
Molecular Function: GO:0000049 - tRNA binding Inferred from experiment [Kammen88]
GO:0009982 - pseudouridine synthase activity Inferred from experiment Inferred by computational analysis [GOA06, GOA01a, GOA01, Kammen88]
GO:0003723 - RNA binding Inferred by computational analysis [GOA01]
GO:0016853 - isomerase activity Inferred by computational analysis [UniProtGOA11]
Cellular Component: GO:0005737 - cytoplasm
GO:0005829 - cytosol Inferred by computational analysis [DiazMejia09]

MultiFun Terms: information transfer RNA related RNA modification

Essentiality data for truA knockouts: ?

Growth Medium Growth? T (°C) O2 pH Osm/L Growth Observations
LB enriched Yes 37 Aerobic 6.95   Yes [Gerdes03, Comment 1]
LB Lennox Yes 37 Aerobic 7   Yes [Baba06, Comment 2]
M9 medium with 1% glycerol Yes 37 Aerobic 7.2 0.35 Yes [Joyce06, Comment 3]
MOPS medium with 0.4% glucose Yes 37 Aerobic 7.2 0.22 Yes [Baba06, Comment 2]

Credits:
Created 09-Oct-2008 by Keseler I , SRI International
Last-Curated ? 09-Oct-2008 by Keseler I , SRI International


Enzymatic reaction of: tRNA pseudouridine synthase

EC Number: 5.4.99.12

a tRNA uridine38-40 <=> a pseudouridine38-40 in tRNA

The reaction direction shown, that is, A + B ↔ C + D versus C + D ↔ A + B, is in accordance with the direction of enzyme catalysis.

The reaction is physiologically favored in the direction shown.

Inhibitors (Competitive): tRNA [Kammen88]


Sequence Features

Feature Class Location Common Name Citations Comment
Cleavage-of-Initial-Methionine 1  
[Kammen88]
 
Mutagenesis-Variant 58  
[Hur07, UniProt11]
Alternate sequence: R → A; UniProt: Loss of activity.
Amino-Acid-Site 58  
[UniProt11]
UniProt: Interaction with tRNA; Important for base-flipping; Sequence Annotation Type: site.
Active-Site 60 catalytic aspartate residue
[Gu99, Huang98a]
 
Amino-Acid-Site 78  
[UniProt11]
UniProt: Interaction with tRNA; Sequence Annotation Type: site.
Protein-Segment 107 -> 111  
[UniProt10a]
UniProt: RNA binding; Sequence Annotation Type: region of interest; Non-Experimental Qualifier: probable;
Amino-Acid-Site 110  
[UniProt11]
UniProt: Interaction with tRNA; Sequence Annotation Type: site.
Amino-Acid-Sites-That-Bind 118  
[UniProt11]
UniProt: Substrate.
Amino-Acid-Site 126  
[UniProt11]
UniProt: Interaction with tRNA; Sequence Annotation Type: site.
Amino-Acid-Site 139  
[UniProt11]
UniProt: Interaction with tRNA; Sequence Annotation Type: site.
Protein-Segment 168 -> 172  
[UniProt11]
UniProt: Interaction with tRNA; Sequence Annotation Type: region of interest.


Gene Local Context (not to scale): ?

Transcription Units:

Notes:

History:
10/20/97 Gene b2318 from Blattner lab Genbank (v. M52) entry merged into EcoCyc gene EG10454; confirmed by SwissProt match.


References

Arps85: Arps PJ, Marvel CC, Rubin BC, Tolan DA, Penhoet EE, Winkler ME (1985). "Structural features of the hisT operon of Escherichia coli K-12." Nucleic Acids Res 13(14);5297-315. PMID: 2991861

Arps87: Arps PJ, Winkler ME (1987). "Structural analysis of the Escherichia coli K-12 hisT operon by using a kanamycin resistance cassette." J Bacteriol 169(3);1061-70. PMID: 3029016

Arps87a: Arps PJ, Winkler ME (1987). "An unusual genetic link between vitamin B6 biosynthesis and tRNA pseudouridine modification in Escherichia coli K-12." J Bacteriol 1987;169(3);1071-9. PMID: 3029017

Baba06: Baba T, Ara T, Hasegawa M, Takai Y, Okumura Y, Baba M, Datsenko KA, Tomita M, Wanner BL, Mori H (2006). "Construction of Escherichia coli K-12 in-frame, single-gene knockout mutants: the Keio collection." Mol Syst Biol 2;2006.0008. PMID: 16738554

Bruni77: Bruni CB, Colantuoni V, Sbordone L, Cortese R, Blasi F (1977). "Biochemical and regulatory properties of Escherichia coli K-12 hisT mutants." J Bacteriol 130(1);4-10. PMID: 323237

Davis91: Davis DR, Poulter CD (1991). "1H-15N NMR studies of Escherichia coli tRNA(Phe) from hisT mutants: a structural role for pseudouridine." Biochemistry 30(17);4223-31. PMID: 2021615

DiazMejia09: Diaz-Mejia JJ, Babu M, Emili A (2009). "Computational and experimental approaches to chart the Escherichia coli cell-envelope-associated proteome and interactome." FEMS Microbiol Rev 33(1);66-97. PMID: 19054114

Foster00: Foster PG, Huang L, Santi DV, Stroud RM (2000). "The structural basis for tRNA recognition and pseudouridine formation by pseudouridine synthase I." Nat Struct Biol 7(1);23-7. PMID: 10625422

Gerdes03: Gerdes SY, Scholle MD, Campbell JW, Balazsi G, Ravasz E, Daugherty MD, Somera AL, Kyrpides NC, Anderson I, Gelfand MS, Bhattacharya A, Kapatral V, D'Souza M, Baev MV, Grechkin Y, Mseeh F, Fonstein MY, Overbeek R, Barabasi AL, Oltvai ZN, Osterman AL (2003). "Experimental determination and system level analysis of essential genes in Escherichia coli MG1655." J Bacteriol 185(19);5673-84. PMID: 13129938

GOA01: GOA, DDB, FB, MGI, ZFIN (2001). "Gene Ontology annotation through association of InterPro records with GO terms."

GOA01a: GOA, MGI (2001). "Gene Ontology annotation based on Enzyme Commission mapping." Genomics 74;121-128.

GOA06: GOA, SIB (2006). "Electronic Gene Ontology annotations created by transferring manual GO annotations between orthologous microbial proteins."

Gu99: Gu X, Liu Y, Santi DV (1999). "The mechanism of pseudouridine synthase I as deduced from its interaction with 5-fluorouracil-tRNA." Proc Natl Acad Sci U S A 96(25);14270-5. PMID: 10588695

Hamma06: Hamma T, Ferre-D'Amare AR (2006). "Pseudouridine synthases." Chem Biol 13(11);1125-35. PMID: 17113994

Huang98a: Huang L, Pookanjanatavip M, Gu X, Santi DV (1998). "A conserved aspartate of tRNA pseudouridine synthase is essential for activity and a probable nucleophilic catalyst." Biochemistry 37(1);344-51. PMID: 9425056

Hur06: Hur S, Stroud RM, Finer-Moore J (2006). "Substrate recognition by RNA 5-methyluridine methyltransferases and pseudouridine synthases: a structural perspective." J Biol Chem 281(51);38969-73. PMID: 17085441

Hur07: Hur S, Stroud RM (2007). "How U38, 39, and 40 of many tRNAs become the targets for pseudouridylation by TruA." Mol Cell 26(2);189-203. PMID: 17466622

Joyce06: Joyce AR, Reed JL, White A, Edwards R, Osterman A, Baba T, Mori H, Lesely SA, Palsson BO, Agarwalla S (2006). "Experimental and computational assessment of conditionally essential genes in Escherichia coli." J Bacteriol 188(23);8259-71. PMID: 17012394

Kammen88: Kammen HO, Marvel CC, Hardy L, Penhoet EE (1988). "Purification, structure, and properties of Escherichia coli tRNA pseudouridine synthase I." J Biol Chem 263(5);2255-63. PMID: 3276686

Lawther77: Lawther RP, Hatfield W (1977). "Biochemical characterization of an Escherichia coli hisT strain." J Bacteriol 130(1);552-7. PMID: 192717

Marvel85: Marvel CC, Arps PJ, Rubin BC, Kammen HO, Penhoet EE, Winkler ME (1985). "hisT is part of a multigene operon in Escherichia coli K-12." J Bacteriol 161(1);60-71. PMID: 2981810

Parker82: Parker J (1982). "Specific mistranslation in hisT mutants of Escherichia coli." Mol Gen Genet 187(3);405-9. PMID: 6757659

Pease02: Pease AJ, Roa BR, Luo W, Winkler ME (2002). "Positive growth rate-dependent regulation of the pdxA, ksgA, and pdxB genes of Escherichia coli K-12." J Bacteriol 184(5);1359-69. PMID: 11844765

Schoenlein89: Schoenlein PV, Roa BB, Winkler ME (1989). "Divergent transcription of pdxB and homology between the pdxB and serA gene products in Escherichia coli K-12." J Bacteriol 171(11);6084-92. PMID: 2681152

Searles86: Searles LL, Jones JW, Fournier MJ, Grambow N, Tyler B, Calvo JM (1986). "Escherichia coli B/r leuK mutant lacking pseudouridine synthase I activity." J Bacteriol 166(1);341-5. PMID: 3514581

Sullivan85: Sullivan MA, Bock RM (1985). "Isolation and characterization of antisuppressor mutations in Escherichia coli." J Bacteriol 161(1);377-84. PMID: 3918006

Tsui91a: Tsui HC, Arps PJ, Connolly DM, Winkler ME (1991). "Absence of hisT-mediated tRNA pseudouridylation results in a uracil requirement that interferes with Escherichia coli K-12 cell division." J Bacteriol 173(22);7395-400. PMID: 1938930

UniProt10a: UniProt Consortium (2010). "UniProt version 2010-07 released on 2010-06-15 00:00:00." Database.

UniProt11: UniProt Consortium (2011). "UniProt version 2011-06 released on 2011-06-30 00:00:00." Database.

UniProtGOA11: UniProt-GOA (2011). "Gene Ontology annotation based on manual assignment of UniProtKB keywords in UniProtKB/Swiss-Prot entries."

Urbonavicius01: Urbonavicius J, Qian Q, Durand JM, Hagervall TG, Bjork GR (2001). "Improvement of reading frame maintenance is a common function for several tRNA modifications." EMBO J 20(17);4863-73. PMID: 11532950

Zhao97: Zhao X, Horne DA (1997). "The role of cysteine residues in the rearrangement of uridine to pseudouridine catalyzed by pseudouridine synthase I." J Biol Chem 272(3);1950-5. PMID: 8999885


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Please cite the following article in publications resulting from the use of EcoCyc: Nucleic Acids Research 41:D605-12 2013
Page generated by SRI International Pathway Tools version 18.5 on Sun Nov 23, 2014, biocyc13.