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Escherichia coli K-12 substr. MG1655 Enzyme: fused tRNA nucleotidyltransferase / 2',3'-cyclic phosphodiesterase / 2' nucleotidase and phosphatase



Gene: cca Accession Numbers: EG10136 (EcoCyc), b3056, ECK3046

Synonyms: CCA-adding enzyme

Regulation Summary Diagram: ?

Summary:
tRNA nucleotidyltransferase catalyzes the addition of CCA to the 3' ends of tRNAs in a template-independent manner [Cudny86]. Phosphatase, nucleotidase, and phosphodiesterase activities of CCA were discovered in a high-throughput screen of purified proteins, and these activities may act together to repair the 3' end of tRNA [Kuznetsova05].

All E. coli tRNAs encode the 3' CCA nucleotides as part of the gene [Komine90].

Polymerization of the CCA sequence does not appear to involve translocation of the enzyme [Shi98]. A model for the specific addition of CCA involving binding sites for ATP and CTP has been proposed [Tomari00, Seth02].

The N-terminal domain of the enzyme contains the nucleotidyltransferase activity. The C-terminal domain carries an HD motif found in a family of metal-dpendent phosphohydrolases and displays a variety of phosphohydrolase activities in the absence of tRNA [Yakunin04]. The C terminus of the protein contains an "anchor domain" of 27 amino acids that defines the addition of CCA versus poly(A), the activity of poly(A) polymerase [Betat04].

tRNA nucleotidyltransferase is not essential for viability of E. coli; a cca null mutant has a slow growth phenotype [Zhu87a]. Poly(A) polymerase I and polynucleotide phosphorylase can partially substitute for tRNA nucleotidyltransferase in the repair of the CCA end of tRNAs [Reuven97].

A cca mutant that is not impaired in incorporation of C, but can not add the A nucleotide at the end of tRNAs has been isolated [McGann80]; the point mutation was shown to lead to substitution of glycine at position 70 by aspartate [Zhu86].

Review: [Hou10]

Citations: [Deutscher74a, Foulds74, Deutscher74, Cudny86a, Deutscher77, Hegg90, Reuven93]

Gene Citations: [Rhodius05]

Locations: cytosol

Map Position: [3,199,913 -> 3,201,151] (68.97 centisomes)
Length: 1239 bp / 412 aa

Molecular Weight of Polypeptide: 46.467 kD (from nucleotide sequence), 47.0 kD (experimental) [Cudny86 ]

Unification Links: ASAP:ABE-0010031 , CGSC:933 , DIP:DIP-9250N , EchoBASE:EB0134 , EcoGene:EG10136 , EcoliWiki:b3056 , Mint:MINT-1263446 , ModBase:P06961 , OU-Microarray:b3056 , PortEco:cca , PR:PRO_000022254 , Protein Model Portal:P06961 , RefSeq:NP_417528 , RegulonDB:EG10136 , SMR:P06961 , String:511145.b3056 , UniProt:P06961

Relationship Links: InterPro:IN-FAMILY:IPR002646 , InterPro:IN-FAMILY:IPR003607 , InterPro:IN-FAMILY:IPR006674 , InterPro:IN-FAMILY:IPR012006 , Pfam:IN-FAMILY:PF01743 , Pfam:IN-FAMILY:PF01966 , Smart:IN-FAMILY:SM00471

In Paralogous Gene Group: 47 (2 members)

Gene-Reaction Schematic: ?

Genetic Regulation Schematic: ?

GO Terms:

Biological Process: GO:0001680 - tRNA 3'-terminal CCA addition Inferred from experiment Inferred by computational analysis [GOA06, GOA01a, GOA01, Deutscher77a, Cudny86]
GO:0042245 - RNA repair Inferred from experiment Inferred by computational analysis [UniProtGOA11, Reuven93]
GO:0006396 - RNA processing Inferred by computational analysis [GOA01]
GO:0008033 - tRNA processing Inferred by computational analysis [UniProtGOA11]
GO:0008152 - metabolic process Inferred by computational analysis [UniProtGOA11]
GO:0016311 - dephosphorylation Inferred by computational analysis [GOA06]
Molecular Function: GO:0004810 - tRNA adenylyltransferase activity Inferred from experiment Inferred by computational analysis [GOA06, GOA01, Cudny86]
GO:0016437 - tRNA cytidylyltransferase activity Inferred from experiment Inferred by computational analysis [GOA06, Cudny86]
GO:0000049 - tRNA binding Inferred by computational analysis [GOA06]
GO:0000166 - nucleotide binding Inferred by computational analysis [UniProtGOA11]
GO:0000287 - magnesium ion binding Inferred by computational analysis [GOA06]
GO:0003723 - RNA binding Inferred by computational analysis [UniProtGOA11, GOA01]
GO:0003824 - catalytic activity Inferred by computational analysis [UniProtGOA11]
GO:0004112 - cyclic-nucleotide phosphodiesterase activity Inferred by computational analysis [GOA06]
GO:0005524 - ATP binding Inferred by computational analysis [UniProtGOA11, GOA06, GOA01]
GO:0016740 - transferase activity Inferred by computational analysis [UniProtGOA11]
GO:0016779 - nucleotidyltransferase activity Inferred by computational analysis [UniProtGOA11, GOA01]
GO:0016787 - hydrolase activity Inferred by computational analysis [UniProtGOA11]
GO:0016791 - phosphatase activity Inferred by computational analysis [GOA06]
GO:0046872 - metal ion binding Inferred by computational analysis [UniProtGOA11]
GO:0052927 - CTP:tRNA cytidylyltransferase activity Inferred by computational analysis [GOA01a]
GO:0052928 - CTP:3'-cytidine-tRNA cytidylyltransferase activity Inferred by computational analysis [GOA01a]
GO:0052929 - ATP:3'-cytidine-cytidine-tRNA adenylyltransferase activity Inferred by computational analysis [GOA01a]
Cellular Component: GO:0005737 - cytoplasm Inferred by curator
GO:0005829 - cytosol Inferred by computational analysis [DiazMejia09]

MultiFun Terms: information transfer RNA related RNA modification

Essentiality data for cca knockouts: ?

Growth Medium Growth? T (°C) O2 pH Osm/L Growth Observations
LB Lennox No 37 Aerobic 7   No [Baba06, Comment 1]

Enzymatic reaction of: tRNA adenylyl-/cytidylyl-transferase (fused tRNA nucleotidyltransferase / 2',3'-cyclic phosphodiesterase / 2' nucleotidase and phosphatase)

Synonyms: tRNA cytidylyltransferase, tRNA CCA-pyrophosphorylase, tRNA CCA-diphosphorylase, CCA-adding enzyme

EC Number: 2.7.7.72

a tRNA precursor + 2 CTP + ATP <=> a tRNA containing a 3' CCA end + 3 diphosphate

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.

Summary:
Initial purification and characterization of the enzyme was done in E. coli B, e.g. [Carre70, Miller71d, Miller71e, Best71, Best71a, Schofield77, Williams77], and MRE 600 [Gross70].

Kinetic Parameters:

Substrate
Km (μM)
Citations
a tRNA precursor
20.0
[Cudny86]
CTP
30.0
[Cudny86]
ATP
330.0
[Cudny86]

pH(opt): 9.4-10 [Cudny86]


Sequence Features

Feature Class Location Citations Comment
Amino-Acid-Sites-That-Bind 8
[UniProt10]
UniProt: ATP or CTP; via amide nitrogen; Non-Experimental Qualifier: by similarity;
Amino-Acid-Sites-That-Bind 11
[UniProt10]
UniProt: ATP or CTP; Non-Experimental Qualifier: by similarity;
Mutagenesis-Variant 21
[Yakunin04, UniProt11]
Alternate sequence: D → A; UniProt: No effect on phosphodiesterase and phosphatase activities.
Metal-Binding-Site 21
[UniProt10]
UniProt: Magnesium; Non-Experimental Qualifier: by similarity;
Mutagenesis-Variant 23
[Yakunin04, UniProt11]
Alternate sequence: D → A; UniProt: No effect on phosphodiesterase and phosphatase activities.
Metal-Binding-Site 23
[UniProt10]
UniProt: Magnesium; Non-Experimental Qualifier: by similarity;
Mutagenesis-Variant 70
[Zhu86, UniProt11]
Alternate sequence: G → D; UniProt: Lowered AMP incorporation.
Amino-Acid-Sites-That-Bind 91
[UniProt10]
UniProt: ATP or CTP; Non-Experimental Qualifier: by similarity;
Amino-Acid-Sites-That-Bind 137
[UniProt10]
UniProt: ATP or CTP; Non-Experimental Qualifier: by similarity;
Amino-Acid-Sites-That-Bind 140
[UniProt10]
UniProt: ATP or CTP; Non-Experimental Qualifier: by similarity;
Mutagenesis-Variant 255
[Yakunin04, UniProt11]
Alternate sequence: H → A; UniProt: Loss of phosphodiesterase and phosphatase activities.
Mutagenesis-Variant 256
[Yakunin04, UniProt11]
Alternate sequence: D → A; UniProt: Loss of phosphodiesterase and phosphatase activities.
Mutagenesis-Variant 305
[Yakunin04, UniProt11]
Alternate sequence: H → A; UniProt: Loss of phosphodiesterase and phosphatase activities.
Mutagenesis-Variant 306
[Yakunin04, UniProt11]
Alternate sequence: D → A; UniProt: Still possesses phosphodiesterase and phosphatase activities.


Gene Local Context (not to scale): ?

Transcription Units:

Notes:

History:
10/20/97 Gene b3056 from Blattner lab Genbank (v. M52) entry merged into EcoCyc gene EG10136; confirmed by SwissProt match.


References

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

Best71: Best AN, Novelli GD (1971). "Studies with tRNA adenylyl(cytidylyl)transferase from Escherichia coli B. I. Purification and kinetic properties." Arch Biochem Biophys 142(2);527-38. PMID: 4927556

Best71a: Best AN, Novelli GD (1971). "Studies with tRNA adenylyl(cytidylyl)transferase from Escherichia coli B. II. Regulation of AMP and CMP incorporation into tRNApCpC and tRNApC." Arch Biochem Biophys 142(2);539-47. PMID: 4927557

Betat04: Betat H, Rammelt C, Martin G, Morl M (2004). "Exchange of regions between bacterial poly(A) polymerase and the CCA-adding enzyme generates altered specificities." Mol Cell 15(3);389-98. PMID: 15304219

Carre70: Carre DS, Litvak S, Chapeville F (1970). "Purification and properties of Escherichia coli CTP (ATP)-tRNA nucleotidyltransferase." Biochim Biophys Acta 224(2);371-81. PMID: 5498070

Cudny86: Cudny H, Deutscher MP (1986). "High-level overexpression, rapid purification, and properties of Escherichia coli tRNA nucleotidyltransferase." J Biol Chem 261(14);6450-3. PMID: 3516995

Cudny86a: Cudny H, Lupski JR, Godson GN, Deutscher MP (1986). "Cloning, sequencing, and species relatedness of the Escherichia coli cca gene encoding the enzyme tRNA nucleotidyltransferase." J Biol Chem 261(14);6444-9. PMID: 3009457

Deutscher74: Deutscher MP (1974). "Preparation of cells permeable to macromolecules by treatment with toluene: studies of transfer ribonucleic acid nucleotidyltransferase." J Bacteriol 118(2);633-9. PMID: 4597454

Deutscher74a: Deutscher MP, Hilderman RH (1974). "Isolation and partial characterization of Escherichia coli mutants with low levels of transfer ribonucleic acid nucleotidyltransferase." J Bacteriol 118(2);621-7. PMID: 4597452

Deutscher77: Deutscher MP, Setlow P, Foulds J (1977). "relA overcomes the slow growth of cca mutants." J Mol Biol 117(4);1095-100. PMID: 342707

Deutscher77a: Deutscher MP, Lin JJ, Evans JA (1977). "Transfer RNA metabolism in Escherichia coli cells deficient in tRNA nucleotidyltransferase." J Mol Biol 117(4);1081-94. PMID: 342706

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

Foulds74: Foulds J, Hilderman RH, Deutscher MP (1974). "Mapping of the locus for Escherichia coli transfer ribonucleic acid nucleotidyltransferase." J Bacteriol 118(2);628-32. PMID: 4597453

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."

Gross70: Gross HJ, Duerinck FR, Fiers WC (1970). "The tRNA pyrophosphorylase activity of Escherichia coli. A study on substrate specificity." Eur J Biochem 17(1);116-23. PMID: 4922328

Hegg90: Hegg LA, Thurlow DL (1990). "Cytidines in tRNAs that are required intact by ATP/CTP:tRNA nucleotidyltransferases from Escherichia coli and Saccharomyces cerevisiae." Nucleic Acids Res 18(20);5975-9. PMID: 1700367

Hou10: Hou YM (2010). "CCA addition to tRNA: Implications for tRNA quality control." IUBMB Life. PMID: 20101632

Komine90: Komine Y, Adachi T, Inokuchi H, Ozeki H (1990). "Genomic organization and physical mapping of the transfer RNA genes in Escherichia coli K12." J Mol Biol 212(4);579-98. PMID: 2184240

Kuznetsova05: Kuznetsova E, Proudfoot M, Sanders SA, Reinking J, Savchenko A, Arrowsmith CH, Edwards AM, Yakunin AF (2005). "Enzyme genomics: Application of general enzymatic screens to discover new enzymes." FEMS Microbiol Rev 29(2);263-79. PMID: 15808744

McGann80: McGann RG, Deutscher MP (1980). "Purification and characterization of a mutant tRNA nucleotidyltransferase." Eur J Biochem 106(1);321-8. PMID: 6176445

Miller71d: Miller JP, Philipps GR (1971). "Transfer ribonucleic acid nucleotidyltransferase from Escherichia coli. II. Purification, physical properties, and substrate specificity." J Biol Chem 246(5);1274-9. PMID: 5545070

Miller71e: Miller JP, Philipps GR (1971). "Transfer ribonucleic acid nucleotidyltransferase from Escherichia coli. 3. Kinetic analysis." J Biol Chem 246(5);1280-4. PMID: 5545071

Reuven93: Reuven NB, Deutscher MP (1993). "Substitution of the 3' terminal adenosine residue of transfer RNA in vivo." Proc Natl Acad Sci U S A 90(10);4350-3. PMID: 7685099

Reuven97: Reuven NB, Zhou Z, Deutscher MP (1997). "Functional overlap of tRNA nucleotidyltransferase, poly(A) polymerase I, and polynucleotide phosphorylase." J Biol Chem 272(52);33255-9. PMID: 9407115

Rhodius05: Rhodius VA, Suh WC, Nonaka G, West J, Gross CA (2005). "Conserved and variable functions of the sigmaE stress response in related genomes." PLoS Biol 4(1);e2. PMID: 16336047

Schofield77: Schofield P, Williams KR (1977). "Purification and some properties of Escherichia coli tRNA nucleotidyltransferase." J Biol Chem 252(15);5584-8. PMID: 328503

Seth02: Seth M, Thurlow DL, Hou YM (2002). "Poly(C) synthesis by class I and class II CCA-adding enzymes." Biochemistry 41(14);4521-32. PMID: 11926813

Shi98: Shi PY, Maizels N, Weiner AM (1998). "CCA addition by tRNA nucleotidyltransferase: polymerization without translocation?." EMBO J 17(11);3197-206. PMID: 9606201

Tomari00: Tomari Y, Suzuki T, Watanabe K, Ueda T (2000). "The role of tightly bound ATP in Escherichia coli tRNA nucleotidyltransferase." Genes Cells 5(9);689-98. PMID: 10971651

UniProt10: 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."

Williams77: Williams KR, Schofield P (1977). "Kinetic mechanism of tRNA nucleotidyltransferase from Escherichia coli." J Biol Chem 252(15);5589-97. PMID: 18468

Yakunin04: Yakunin AF, Proudfoot M, Kuznetsova E, Savchenko A, Brown G, Arrowsmith CH, Edwards AM (2004). "The HD domain of the Escherichia coli tRNA nucleotidyltransferase has 2',3'-cyclic phosphodiesterase, 2'-nucleotidase, and phosphatase activities." J Biol Chem 279(35);36819-27. PMID: 15210699

Zhu86: Zhu LQ, Cudny H, Deutscher MP (1986). "A mutation in Escherichia coli tRNA nucleotidyltransferase that affects only AMP incorporation is in a sequence often associated with nucleotide-binding proteins." J Biol Chem 261(32);14875-7. PMID: 3533927

Zhu87a: Zhu L, Deutscher MP (1987). "tRNA nucleotidyltransferase is not essential for Escherichia coli viability." EMBO J 6(8);2473-7. PMID: 3311729

Other References Related to Gene Regulation

Gogol11: Gogol EB, Rhodius VA, Papenfort K, Vogel J, Gross CA (2011). "Small RNAs endow a transcriptional activator with essential repressor functions for single-tier control of a global stress regulon." Proc Natl Acad Sci U S A 108(31);12875-80. PMID: 21768388


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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 Fri Nov 28, 2014, biocyc14.