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Escherichia coli K-12 substr. MG1655 Enzyme: dihydroneopterin triphosphate pyrophosphohydrolase



Gene: nudB Accession Numbers: EG11138 (EcoCyc), b1865, ECK1866

Synonyms: yebD, ntpA, Orf17

Regulation Summary Diagram: ?

Summary:
Dihydroneopterin triphosphate pyrophosphohydrolase (DHNTPase) catalyzes the hydrolytic removal of pyrophosphate from dihydroneopterin triphosphate (DHNTP) to yield dihydroneopterin phosphate as a product. This is the committed step in the biosynthesis of folate [Suzuki74, Gabelli07].

DHNTPase was also shown to have nucleotide pyrophosphohydrolase activity with various substrates [OHandley96, Hori05], but the catalytic efficiency of the enzyme for DHNTP is over 7 times that for dATP. The severe reduction of folate levels in a nudB null mutant similarly argues that DHNTP is the physiological substrate of the enzyme [Gabelli07].

Crystal structures of NudB have been solved at 1.8 Å resolution [Gabelli07].

A nudB null mutant has no detectable growth defect in either M9 minimal medium or LB; however, mutant cells contain significantly reduced levels of folic acid [Gabelli07]. Deletion of nudB causes a "glycogen-excess" phenotype during entrance to stationary phase [Eydallin07].

Reviews: [McLennan06, McLennan07]

Gene Citations: [Takahagi91]

Locations: cytosol

Map Position: [1,946,204 <- 1,946,656] (41.95 centisomes)
Length: 453 bp / 150 aa

Molecular Weight of Polypeptide: 17.306 kD (from nucleotide sequence), 17.0 kD (experimental) [Suzuki74 ]

Unification Links: ASAP:ABE-0006223 , EchoBASE:EB1128 , EcoGene:EG11138 , EcoliWiki:b1865 , ModBase:P0AFC0 , OU-Microarray:b1865 , PortEco:nudB , PR:PRO_000023420 , Protein Model Portal:P0AFC0 , RefSeq:NP_416379 , RegulonDB:EG11138 , SMR:P0AFC0 , String:511145.b1865 , UniProt:P0AFC0

Relationship Links: InterPro:IN-FAMILY:IPR000086 , InterPro:IN-FAMILY:IPR003564 , InterPro:IN-FAMILY:IPR015797 , InterPro:IN-FAMILY:IPR020084 , PDB:Structure:2O1C , PDB:Structure:2O5W , Pfam:IN-FAMILY:PF00293 , Prints:IN-FAMILY:PR01404 , Prosite:IN-FAMILY:PS00893 , Prosite:IN-FAMILY:PS51462

Gene-Reaction Schematic: ?

Instance reactions of [a nucleoside triphosphate + H2O → a nucleoside 5'-monophosphate + diphosphate + H+] (3.6.1.19):
i1: CTP + H2O → CMP + diphosphate + H+ (3.6.1.65)

i2: dATP + H2O → dAMP + diphosphate + H+ (3.6.1.19)

i3: dUTP + H2O → dUMP + diphosphate + H+ (3.6.1.19/3.6.1.23)

i4: dGTP + H2O → dGMP + diphosphate + H+ (3.6.1.19)

i5: dITP + H2O → dIMP + diphosphate + H+ (3.6.1.66)

i6: ITP + H2O → IMP + diphosphate + H+ (3.6.1.19)

i7: XTP + H2O → XMP + diphosphate + H+ (3.6.1.66)

i8: ATP + H2O → AMP + diphosphate + H+ (3.6.1.8)

i9: dTTP + H2O → dTMP + diphosphate + H+ (3.6.1.19)

i10: UTP + H2O → UMP + diphosphate + H+ (3.6.1.19)

i11: dCTP + H2O → dCMP + diphosphate + H+ (3.6.1.65)

GO Terms:

Biological Process: GO:0046654 - tetrahydrofolate biosynthetic process Inferred from experiment [Gabelli07]
GO:0046656 - folic acid biosynthetic process Inferred from experiment Inferred by computational analysis [UniProtGOA11a, Gabelli07]
GO:0006281 - DNA repair Inferred by computational analysis [GOA01]
Molecular Function: GO:0000287 - magnesium ion binding Inferred from experiment [Suzuki74]
GO:0008828 - dATP pyrophosphohydrolase activity Inferred from experiment Inferred by computational analysis [GOA01, Gabelli07]
GO:0019177 - dihydroneopterin triphosphate pyrophosphohydrolase activity Inferred from experiment [Gabelli07]
GO:0016787 - hydrolase activity Inferred by computational analysis [UniProtGOA11a, GOA01]
GO:0046872 - metal ion binding Inferred by computational analysis [UniProtGOA11a]
Cellular Component: GO:0005829 - cytosol Inferred by computational analysis [DiazMejia09]

MultiFun Terms: metabolism biosynthesis of building blocks cofactors, small molecule carriers folic acid

Essentiality data for nudB knockouts: ?

Growth Medium Growth? T (°C) O2 pH Osm/L Growth Observations
LB Lennox Yes 37 Aerobic 7   Yes [Baba06, Comment 1]
M9 medium with 1% glycerol Yes 37 Aerobic 7.2 0.35 Yes [Joyce06, Comment 2]
MOPS medium with 0.4% glucose Yes 37 Aerobic 7.2 0.22 Yes [Baba06, Comment 1]
Yes [Feist07, Comment 3]

Credits:
Last-Curated ? 12-Jul-2011 by Keseler I , SRI International


Enzymatic reaction of: dihydroneopterin triphosphate pyrophosphohydrolase

Synonyms: dihydroneopterin-PPP pyrophosphohydrolase, H2-neopterin-PPP pyrophosphohydrolase, DHNTPase

EC Number: 3.6.1.-

7,8-dihydroneopterin 3'-triphosphate + H2O <=> 7,8-dihydroneopterin 3'-phosphate + diphosphate + H+

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.

Alternative Substrates [Comment 4]:

In Pathways: superpathway of chorismate metabolism , superpathway of tetrahydrofolate biosynthesis , 6-hydroxymethyl-dihydropterin diphosphate biosynthesis I

Cofactors or Prosthetic Groups [Comment 5]: Mg2+ [Suzuki74]

Inhibitors (Unknown Mechanism): 7,8-dihydroneopterin 3'-phosphate [Suzuki74] , dTTP [Suzuki74] , diphosphate [Suzuki74, Comment 6]

Kinetic Parameters:

Substrate
Km (μM)
kcat (sec-1)
kcat/Km (sec-1 μM-1)
Citations
7,8-dihydroneopterin 3'-triphosphate
270.0
11.6
[Gabelli07]

T(opt): 42 °C [Suzuki74]

pH(opt): 8.5 [Suzuki74]


Enzymatic reaction of: pyrophosphohydrolase

Synonyms: dATP pyrophosphohydrolase

dATP + H2O <=> dAMP + diphosphate + H+

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.

Alternative Substrates for dATP: 8-OH-dATP [Hori05 ] , dADP [Hori05 ] , 8-oxo-dADP [Hori05 ]

Summary:
The enzyme shows a preference for dATP over the other (deoxy)nucleoside triphosphates [OHandley96]; it was also found to be capable of hydrolyzing various damaged di- and trinucleotides. The Km for the best such substrate, 8-OH-dADP, is 69 µM [Hori05].

Cofactors or Prosthetic Groups [Comment 7]: Mg2+ [OHandley96]

Inhibitors (Competitive): dAMP [OHandley96, Comment 8] , diphosphate [OHandley96, Comment 9]

Kinetic Parameters:

Substrate
Km (μM)
kcat (sec-1)
kcat/Km (sec-1 μM-1)
Citations
dATP
800.0
[OHandley96]
dATP
4.59
[Gabelli07]

pH(opt): 8.6 [OHandley96]


Sequence Features

Feature Class Location Citations Comment
Conserved-Region 5 -> 146
[UniProt09]
UniProt: Nudix hydrolase;
Amino-Acid-Sites-That-Bind 7
[UniProt10]
UniProt: Substrate;
Amino-Acid-Sites-That-Bind 29
[UniProt10]
UniProt: Substrate;
Amino-Acid-Sites-That-Bind 40
[UniProt10]
UniProt: Substrate;
Protein-Segment 41 -> 62
[UniProt10]
UniProt: Nudix box; Sequence Annotation Type: short sequence motif;
Metal-Binding-Site 56
[UniProt10]
UniProt: Magnesium;
Metal-Binding-Site 60
[UniProt10]
UniProt: Magnesium;
Protein-Segment 81 -> 84
[UniProt10a]
UniProt: Substrate binding; Sequence Annotation Type: region of interest; Non-Experimental Qualifier: potential;
Metal-Binding-Site 117
[UniProt10]
UniProt: Magnesium;
Amino-Acid-Sites-That-Bind 135
[UniProt10a]
UniProt: Substrate; Non-Experimental Qualifier: potential;


Gene Local Context (not to scale): ?

Transcription Units:

Notes:

History:
10/20/97 Gene b1865 from Blattner lab Genbank (v. M52) entry merged into EcoCyc gene EG11138; 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

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

Eydallin07: Eydallin G, Viale AM, Moran-Zorzano MT, Munoz FJ, Montero M, Baroja-Fernandez E, Pozueta-Romero J (2007). "Genome-wide screening of genes affecting glycogen metabolism in Escherichia coli K-12." FEBS Lett 581(16);2947-53. PMID: 17543954

Feist07: Feist AM, Henry CS, Reed JL, Krummenacker M, Joyce AR, Karp PD, Broadbelt LJ, Hatzimanikatis V, Palsson BO (2007). "A genome-scale metabolic reconstruction for Escherichia coli K-12 MG1655 that accounts for 1260 ORFs and thermodynamic information." Mol Syst Biol 3;121. PMID: 17593909

Gabelli07: Gabelli SB, Bianchet MA, Xu W, Dunn CA, Niu ZD, Amzel LM, Bessman MJ (2007). "Structure and function of the E. coli dihydroneopterin triphosphate pyrophosphatase: a Nudix enzyme involved in folate biosynthesis." Structure 15(8);1014-22. PMID: 17698004

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

Hori05: Hori M, Fujikawa K, Kasai H, Harashima H, Kamiya H (2005). "Dual hydrolysis of diphosphate and triphosphate derivatives of oxidized deoxyadenosine by Orf17 (NtpA), a MutT-type enzyme." DNA Repair (Amst) 4(1);33-9. PMID: 15533835

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

McLennan06: McLennan AG (2006). "The Nudix hydrolase superfamily." Cell Mol Life Sci 63(2);123-43. PMID: 16378245

McLennan07: McLennan AG (2007). "Folate synthesis: an old enzyme identified." Structure 15(8);891-2. PMID: 17697994

OHandley96: O'Handley SF, Frick DN, Bullions LC, Mildvan AS, Bessman MJ (1996). "Escherichia coli orf17 codes for a nucleoside triphosphate pyrophosphohydrolase member of the MutT family of proteins. Cloning, purification, and characterization of the enzyme." J Biol Chem 1996;271(40);24649-54. PMID: 8798731

Suzuki74: Suzuki Y, Brown GM (1974). "The biosynthesis of folic acid. XII. Purification and properties of dihydroneopterin triphosphate pyrophosphohydrolase." J Biol Chem 1974;249(8);2405-10. PMID: 4362677

Takahagi91: Takahagi M, Iwasaki H, Nakata A, Shinagawa H (1991). "Molecular analysis of the Escherichia coli ruvC gene, which encodes a Holliday junction-specific endonuclease." J Bacteriol 173(18);5747-53. PMID: 1885548

UniProt09: UniProt Consortium (2009). "UniProt version 15.8 released on 2009-10-01 00:00:00." Database.

UniProt10: UniProt Consortium (2010). "UniProt version 2010-11 released on 2010-11-02 00:00:00." Database.

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

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


Report Errors or Provide Feedback
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 Thu Dec 18, 2014, biocyc13.