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MetaCyc Enzyme: dTDP-glucose pyrophosphorylase

Gene: rfbA Accession Numbers: EG11978 (MetaCyc), b2039, ECK2033

Synonyms: rmlA, som

Species: Escherichia coli K-12 substr. MG1655

Subunit composition of dTDP-glucose pyrophosphorylase = [RfbA]4

Summary:
dTDP-glucose pyrophosphorylase is involved in the biosynthesis of dTDP-L-rhamnose. dTDP-L-rhamnose is a precursor of L-rhamnose which is an essential component of surface antigens such as the O-lipopolysaccharide. The enzyme catalyzes the formation of dTDP-glucose from dTTP and glucose-1-phosphate, as well as its pyrophosphorolysis. A divalent cation is essential for catalysis [Zuccotti01].

dTDP-glucose pyrophosphorylase is encoded by the rfbA gene which is paralogous to rffH. The product of rffH is another dTDP-glucose pyrophosphorylase which catalyzes the same reaction, but functions in the enterobacterial common antigen biosynthesis pathway. There is 68% amino acid sequence identity between the two paralogs. [Sivaraman02, Marolda95, Stevenson94]

The crystal structure of the homotetrameric molecule in complex with dDTP-glucose was refined at 1.9 Å resolution and shows conserved substrate and inhibitor binding modes [Zuccotti01].

Mutant cells deficient in dTDP-glucose pyrophosphorylase activity lose viability faster, form longer filaments, lose thymidine nucleotides and dTDP-sugar pools at a faster rate than wild-type cells [Ohkawa75].

Locations: cytosol

Map Position: [2,108,162 <- 2,109,043]

Molecular Weight of Polypeptide: 32.694 kD (from nucleotide sequence)

Unification Links: ASAP:ABE-0006762 , CGSC:297 , EchoBASE:EB1921 , EcoGene:EG11978 , EcoliWiki:b2039 , ModBase:P37744 , OU-Microarray:b2039 , PortEco:rfbA , PR:PRO_000023783 , Pride:P37744 , Protein Model Portal:P37744 , RefSeq:NP_416543 , RegulonDB:EG11978 , SMR:P37744 , String:511145.b2039 , UniProt:P37744

Relationship Links: InterPro:IN-FAMILY:IPR005835 , InterPro:IN-FAMILY:IPR005907 , InterPro:IN-FAMILY:IPR029044 , Panther:IN-FAMILY:PTHR22572:SF13 , PDB:Structure:1H5R , PDB:Structure:1H5S , PDB:Structure:1H5T , Pfam:IN-FAMILY:PF00483

Gene-Reaction Schematic: ?

Gene-Reaction Schematic

GO Terms:

Biological Process: GO:0009058 - biosynthetic process Inferred by computational analysis [GOA01a]
GO:0009103 - lipopolysaccharide biosynthetic process Inferred by computational analysis [UniProtGOA11a]
GO:0009243 - O antigen biosynthetic process Inferred by computational analysis [UniProtGOA12]
GO:0019305 - dTDP-rhamnose biosynthetic process Inferred by computational analysis [UniProtGOA12]
GO:0045226 - extracellular polysaccharide biosynthetic process Inferred by computational analysis [GOA01a]
Molecular Function: GO:0008879 - glucose-1-phosphate thymidylyltransferase activity Inferred from experiment Inferred by computational analysis [GOA01, GOA01a, Zuccotti01]
GO:0042802 - identical protein binding Inferred from experiment [Zuccotti01]
GO:0046872 - metal ion binding Inferred from experiment Inferred by computational analysis [UniProtGOA11a, Zuccotti01]
GO:0016740 - transferase activity Inferred by computational analysis [UniProtGOA11a]
GO:0016779 - nucleotidyltransferase activity Inferred by computational analysis [UniProtGOA11a, GOA01a]
Cellular Component: GO:0005829 - cytosol Inferred from experiment Inferred by computational analysis [DiazMejia09, Ishihama08, Lasserre06]

MultiFun Terms: cell structure surface antigens (ECA, O antigen of LPS)
metabolism biosynthesis of macromolecules (cellular constituents) lipopolysaccharide O antigen
metabolism central intermediary metabolism sugar nucleotide biosynthesis, conversions

Credits:
Created in EcoCyc 30-Aug-2013 by Kubo A , SRI International
Imported from EcoCyc 02-Jun-2015 by Paley S , SRI International


Enzymatic reaction of: dTDP-glucose pyrophosphorylase

Synonyms: glucose-1-phosphate thymidylyltransferase, dTDP-glucose synthase, dTTP:α-D-glucose-1-phosphate thymidylyltransferase

EC Number: 2.7.7.24

α-D-glucopyranose 1-phosphate + dTTP + H+ <=> dTDP-α-D-glucose + 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.

This reaction is reversible. [Zuccotti01]

In Pathways: superpathway of dTDP-glucose-derived O-antigen building blocks biosynthesis , O-antigen building blocks biosynthesis (E. coli) , enterobacterial common antigen biosynthesis , dTDP-L-rhamnose biosynthesis I , dTDP-N-acetylthomosamine biosynthesis

Credits:
Imported from EcoCyc 02-Jun-2015 by Paley S , SRI International

Cofactors or Prosthetic Groups: Mg2+ [Comment 1, Zuccotti01]

Inhibitors (Competitive): UDP-D-glucose [Bernstein65]

Inhibitors (Unknown Mechanism): dTDP-α-D-glucose [Zuccotti01] , dTDP-β-L-rhamnose , diphosphate [Zuccotti01, Comment 2]

Kinetic Parameters:

Substrate
Km (μM)
kcat (sec-1)
kcat/Km (sec-1 μM-1)
Citations
α-D-glucopyranose 1-phosphate
34.0
[Zuccotti01, BRENDA14]
dTDP-α-D-glucose
95.0
95.0
1.0
[Zuccotti01, BRENDA14]
dTTP
20.5
[Zuccotti01]
diphosphate
154.0
154.0
1.0
[Zuccotti01, BRENDA14]

pH(opt): 8.0 [BRENDA14, Zuccotti01]


Sequence Features

Feature Class Location Citations Comment
Metal-Binding-Site 111
[UniProt10]
UniProt: Magnesium; Non-Experimental Qualifier: by similarity;
Metal-Binding-Site 226
[UniProt10]
UniProt: Magnesium; Non-Experimental Qualifier: by similarity;
Sequence-Conflict 247
[Yao94, UniProt10a]
UniProt: (in Ref. 6; AAC31629);

History:
10/20/97 Gene b2039 from Blattner lab Genbank (v. M52) entry merged into EcoCyc gene EG11978; confirmed by SwissProt match.


References

Bernstein65: Bernstein R, Robbins P "Control aspects of uridine 5'-diphosphate glucose and thymidine 5'-diphosphate glucose synthesis by microbial enzymes." J Biol Chem 1965;240(1):391-397.

BRENDA14: BRENDA team (2014). "Imported from BRENDA version existing on Aug 2014." http://www.brenda-enzymes.org.

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

GOA01: GOA, MGI (2001). "Gene Ontology annotation based on Enzyme Commission mapping." Genomics 74;121-128.

GOA01a: GOA, DDB, FB, MGI, ZFIN (2001). "Gene Ontology annotation through association of InterPro records with GO terms."

Ishihama08: Ishihama Y, Schmidt T, Rappsilber J, Mann M, Hartl FU, Kerner MJ, Frishman D (2008). "Protein abundance profiling of the Escherichia coli cytosol." BMC Genomics 9;102. PMID: 18304323

Lasserre06: Lasserre JP, Beyne E, Pyndiah S, Lapaillerie D, Claverol S, Bonneu M (2006). "A complexomic study of Escherichia coli using two-dimensional blue native/SDS polyacrylamide gel electrophoresis." Electrophoresis 27(16);3306-21. PMID: 16858726

Marolda95: Marolda CL, Valvano MA (1995). "Genetic analysis of the dTDP-rhamnose biosynthesis region of the Escherichia coli VW187 (O7:K1) rfb gene cluster: identification of functional homologs of rfbB and rfbA in the rff cluster and correct location of the rffE gene." J Bacteriol 1995;177(19);5539-46. PMID: 7559340

Ohkawa75: Ohkawa T (1975). "Studies of intracellular thymidine nucleotides. Thymineless death and the recovery after re-addition of thymine in Escherichia coli K 12." Eur J Biochem 60(1);57-66. PMID: 1107038

Sivaraman02: Sivaraman J, Sauve V, Matte A, Cygler M (2002). "Crystal structure of Escherichia coli glucose-1-phosphate thymidylyltransferase (RffH) complexed with dTTP and Mg2+." J Biol Chem 277(46);44214-9. PMID: 12171937

Stevenson94: Stevenson G, Neal B, Liu D, Hobbs M, Packer NH, Batley M, Redmond JW, Lindquist L, Reeves P (1994). "Structure of the O antigen of Escherichia coli K-12 and the sequence of its rfb gene cluster." J Bacteriol 1994;176(13);4144-56. PMID: 7517391

UniProt10: UniProt Consortium (2010). "UniProt version 2010-07 released on 2010-06-15 00:00:00." Database.

UniProt10a: UniProt Consortium (2010). "UniProt version 2010-11 released on 2010-11-02 00:00:00." Database.

UniProtGOA11a: UniProt-GOA (2011). "Gene Ontology annotation based on manual assignment of UniProtKB keywords in UniProtKB/Swiss-Prot entries."

UniProtGOA12: UniProt-GOA (2012). "Gene Ontology annotation based on UniPathway vocabulary mapping."

Yao94: Yao Z, Valvano MA (1994). "Genetic analysis of the O-specific lipopolysaccharide biosynthesis region (rfb) of Escherichia coli K-12 W3110: identification of genes that confer group 6 specificity to Shigella flexneri serotypes Y and 4a." J Bacteriol 1994;176(13);4133-43. PMID: 7517390

Zuccotti01: Zuccotti S, Zanardi D, Rosano C, Sturla L, Tonetti M, Bolognesi M (2001). "Kinetic and crystallographic analyses support a sequential-ordered bi bi catalytic mechanism for Escherichia coli glucose-1-phosphate thymidylyltransferase." J Mol Biol 313(4);831-43. PMID: 11697907


Report Errors or Provide Feedback
Please cite the following article in publications resulting from the use of MetaCyc: Caspi et al, Nucleic Acids Research 42:D459-D471 2014
Page generated by SRI International Pathway Tools version 19.0 on Mon Aug 3, 2015, BIOCYC14B.