|Gene:||thyA||Accession Numbers: EG11002 (MetaCyc), b2827, ECK2823|
Species: Escherichia coli K-12 substr. MG1655
Subunit composition of
thymidylate synthase = [ThyA]2
thymidylate synthase = ThyA
Thymidylate synthase plays a key role in DNA synthesis. The conversion of dUMP to dTMP is the main pathway of de novo dTMP synthesis in the cell [Haertle79, Belfort83]. The enzyme undergoes conformational changes during the initial binding of dUMP, with even larger changes during the binding of the cofactor. The catalytic mechanism has been studied [Agrawal04].
Detailed studies of the overall mechanism of thymidylate synthase [Kanaan07], and of the proton abstraction and hydride transfer steps using purified wild-type and mutant enzymes [Hong07], have provided new insight into the reaction mechanism. The nature of the dimeric state has also been studied using wild-type and mutant enzymes [Saxl07].
In Escherichia coli K-12 the thyA gene encoding this enzyme contains a natural hotspot for T to A transversion mutations at base 131 of the coding sequence. This hotspot occurs within a 17 base pair quasi-palindrome and a template-switch mechanism of mutagenesis was confirmed and studied in detail [Dutra06].
Immunofluorescence microscopy studies showed E. coli thymidylate synthase to be located throughout the cell, which does not support the existence of a dNTP-synthesizing complex exclusively located near the replication factory [denBlaauwen06].
Inhibitors of the E. coli and human enzymes have been synthesized and studied with the aim of developing antitumor agents. These antifolate inhibitors of thymidylate synthase were novel 2-amino-4-oxo-5-arylthio-substituted-pyrrolo[2,3-d]pyrimidines [Gangjee05].
thyA expression may be directly or indirectly repressed by OmpR [Brombacher03].
|Map Position: [2,962,383 <- 2,963,177]|
Molecular Weight of Polypeptide: 30.48 kD (from nucleotide sequence)
Unification Links: ASAP:ABE-0009268 , CGSC:104 , DIP:DIP-48261N , EchoBASE:EB0995 , EcoGene:EG11002 , EcoliWiki:b2827 , Entrez-gene:949035 , Mint:MINT-1265510 , ModBase:P0A884 , OU-Microarray:b2827 , PortEco:thyA , Pride:P0A884 , Protein Model Portal:P0A884 , RefSeq:NP_417304 , RegulonDB:EG11002 , SMR:P0A884 , String:511145.b2827 , UniProt:P0A884
Relationship Links: InterPro:IN-FAMILY:IPR000398 , InterPro:IN-FAMILY:IPR020940 , InterPro:IN-FAMILY:IPR023451 , PDB:Structure:1AIQ , PDB:Structure:1AJM , PDB:Structure:1AN5 , PDB:Structure:1AOB , PDB:Structure:1AXW , PDB:Structure:1BDU , PDB:Structure:1BID , PDB:Structure:1BJG , PDB:Structure:1BQ1 , PDB:Structure:1BQ2 , PDB:Structure:1DDU , PDB:Structure:1DNA , PDB:Structure:1EV5 , PDB:Structure:1EV8 , PDB:Structure:1EVF , PDB:Structure:1EVG , PDB:Structure:1F4B , PDB:Structure:1F4C , PDB:Structure:1F4D , PDB:Structure:1F4E , PDB:Structure:1F4F , PDB:Structure:1F4G , PDB:Structure:1FFL , PDB:Structure:1FWM , PDB:Structure:1JG0 , PDB:Structure:1JTQ , PDB:Structure:1JTU , PDB:Structure:1JUT , PDB:Structure:1KCE , PDB:Structure:1KZI , PDB:Structure:1KZJ , PDB:Structure:1NCE , PDB:Structure:1QQQ , PDB:Structure:1SYN , PDB:Structure:1TDU , PDB:Structure:1TJS , PDB:Structure:1TLC , PDB:Structure:1TLS , PDB:Structure:1TRG , PDB:Structure:1TSD , PDB:Structure:1TSN , PDB:Structure:1TYS , PDB:Structure:1ZPR , PDB:Structure:2A9W , PDB:Structure:2BBQ , PDB:Structure:2FTN , PDB:Structure:2FTO , PDB:Structure:2FTQ , PDB:Structure:2g8x , PDB:Structure:2KCE , PDB:Structure:2TSC , PDB:Structure:2vet , PDB:Structure:2vf0 , PDB:Structure:3B5B , PDB:Structure:3B9H , PDB:Structure:3BFI , PDB:Structure:3BGX , PDB:Structure:3BHL , PDB:Structure:3BHR , PDB:Structure:3TMS , PDB:Structure:4F2V , PDB:Structure:4GEV , PDB:Structure:4ISK , PDB:Structure:4KNZ , Pfam:IN-FAMILY:PF00303 , Prints:IN-FAMILY:PR00108 , Prosite:IN-FAMILY:PS00091
|Biological Process:||GO:0006231 - dTMP biosynthetic process
GO:0006235 - dTTP biosynthetic process [UniProtGOA12]
GO:0006417 - regulation of translation [UniProtGOA11]
GO:0009165 - nucleotide biosynthetic process [UniProtGOA11]
GO:0032259 - methylation [UniProtGOA11]
|Molecular Function:||GO:0004799 - thymidylate synthase activity
[GOA06, GOA01a, GOA01, Haertle79]
GO:0003723 - RNA binding [UniProtGOA11]
GO:0008168 - methyltransferase activity [UniProtGOA11]
GO:0016740 - transferase activity [UniProtGOA11]
|Cellular Component:||GO:0005829 - cytosol
GO:0005737 - cytoplasm [UniProtGOA11a, UniProtGOA11, GOA06]
|MultiFun Terms:||metabolism → central intermediary metabolism → formyl-THF biosynthesis|
|metabolism → central intermediary metabolism → nucleotide and nucleoside conversions|
Enzymatic reaction of: thymidylate synthase
Synonyms: 5,10-methylenetetrahydrofolate:dUMP C-methyltransferase
EC Number: 188.8.131.52
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 irreversible in the direction shown. [Kanaan09]
In Pathways: superpathway of pyrimidine deoxyribonucleotides de novo biosynthesis (E. coli) , superpathway of pyrimidine deoxyribonucleotides de novo biosynthesis , pyrimidine deoxyribonucleotides de novo biosynthesis II , pyrimidine deoxyribonucleotides de novo biosynthesis I , N10-formyl-tetrahydrofolate biosynthesis
Thymidylate synthase catalyzes the reductive methylation of 2'-deoxyuridine-5'-monophosphate (dTMP) using the cofactor N5,N10-methylenetetrahydrofolate as a donor of both methylene and hydride, and producing 7,8-dihydrofolate. Hydride transfer from the 6S position of tetrahydrofolate is the rate limiting step of the overall reaction [Haertle79, SoteloMundo06, Newby06, Kanaan07, Hong07, Kanaan09].
Inhibitors (Competitive): stereoisomer (-)CH2H4folate [Comment 1]
10/20/97 Gene b2827 from Blattner lab Genbank (v. M52) entry merged into EcoCyc gene EG11002; confirmed by SwissProt match.
Agrawal04: Agrawal N, Hong B, Mihai C, Kohen A (2004). "Vibrationally enhanced hydrogen tunneling in the Escherichia coli thymidylate synthase catalyzed reaction." Biochemistry 43(7);1998-2006. PMID: 14967040
Belfort83: Belfort M, Maley G, Pedersen-Lane J, Maley F (1983). "Primary structure of the Escherichia coli thyA gene and its thymidylate synthase product." Proc Natl Acad Sci U S A 1983;80(16);4914-8. PMID: 6308660
Brombacher03: Brombacher E, Dorel C, Zehnder AJ, Landini P (2003). "The curli biosynthesis regulator CsgD co-ordinates the expression of both positive and negative determinants for biofilm formation in Escherichia coli." Microbiology 149(Pt 10);2847-57. PMID: 14523117
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
Galova92: Galova M, Koptidesova D, Rusznakova D, Racay P, Kollarova M (1992). "Characteristics of NADPH-dependent thymidylate synthetase purified from Streptomyces aureofaciens." Arch Biochem Biophys 296(1);81-7. PMID: 1605647
Gangjee05: Gangjee A, Jain HD, Kisliuk RL (2005). "Novel 2-amino-4-oxo-5-arylthio-substituted-pyrrolo[2,3-d]pyrimidines as nonclassical antifolate inhibitors of thymidylate synthase." Bioorg Med Chem Lett 15(9);2225-30. PMID: 15837298
Haertle79: Haertle T, Wohlrab F, Guschlbauer W (1979). "Thymidylate synthetase from Escherichia coli K12. Purification, and dependence of kinetic properties on sugar conformation and size of the 2' substituent." Eur J Biochem 1979;102(1);223-30. PMID: 42538
Kanaan07: Kanaan N, Marti S, Moliner V, Kohen A (2007). "A quantum mechanics/molecular mechanics study of the catalytic mechanism of the thymidylate synthase." Biochemistry 46(12);3704-13. PMID: 17328531
Kanaan09: Kanaan N, Marti S, Moliner V, Kohen A (2009). "QM/MM study of thymidylate synthase: enzymatic motions and the temperature dependence of the rate limiting step." J Phys Chem A 113(10);2176-82. PMID: 19182971
Montfort90: Montfort WR, Perry KM, Fauman EB, Finer-Moore JS, Maley GF, Hardy L, Maley F, Stroud RM (1990). "Structure, multiple site binding, and segmental accommodation in thymidylate synthase on binding dUMP and an anti-folate." Biochemistry 1990;29(30);6964-77. PMID: 2223754
Newby06: Newby Z, Lee TT, Morse RJ, Liu Y, Liu L, Venkatraman P, Santi DV, Finer-Moore JS, Stroud RM (2006). "The role of protein dynamics in thymidylate synthase catalysis: variants of conserved 2'-deoxyuridine 5'-monophosphate (dUMP)-binding Tyr-261." Biochemistry 45(24);7415-28. PMID: 16768437
Roberts06: Roberts SA, Hyatt DC, Honts JE, Changchien L, Maley GF, Maley F, Montfort WR (2006). "Structure of the Y94F mutant of Escherichia coli thymidylate synthase." Acta Crystallogr Sect F Struct Biol Cryst Commun 62(Pt 9);840-3. PMID: 16946460
Saxl07: Saxl RL, Maley GF, Hauer CR, Maccoll R, Changchien L, Maley F (2007). "Significance of mutations on the structural perturbation of thymidylate synthase: implications for their involvement in subunit exchange." Protein Sci 16(7);1439-48. PMID: 17586776
SoteloMundo06: Sotelo-Mundo RR, Changchien L, Maley F, Montfort WR (2006). "Crystal structures of thymidylate synthase mutant R166Q: structural basis for the nearly complete loss of catalytic activity." J Biochem Mol Toxicol 20(2);88-92. PMID: 16615077
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