Escherichia coli K-12 substr. MG1655 Enzyme: GMP synthetase

Gene: guaA Accession Numbers: EG10420 (EcoCyc), b2507, ECK2503

Regulation Summary Diagram: ?

Regulation summary diagram for guaA

Subunit composition of GMP synthetase = [GuaA]2
         GMP synthetase = GuaA

GMP synthetase catalyzes the glutamine- or ammonia-dependent synthesis of GMP from XMP [Patel75].

GMP synthetase contains an N-terminal glutamine amide transfer (GAT) domain [Zalkin85], a central ATP-pyrophosphatase (ATPP) domain, and a C-terminal dimerization domain [Tesmer96]. The GAT domain utilizes glutamine to generate ammonia, which is then transferred to the ATPP domain containing the adenylylated XMP intermediate via a substrate-protective channel or tunnel.

A protein consisting only of the ATPP and dimerization domains of GMP synthetase dimerizes in solution and has similar Km values for ATP, XMP, and ammonia as the full-length protein. However, the rate of catalysis using ammonia as a substrate is dramatically increased, possibly due to enhanced access to the active site [Abbott06]. Rapid kinetics studies of the full-length enzyme allowed analysis of the catalytic cycle, including observation of the enzyme-bound adenylylated XMP intermediate and the functional role of a substrate-induced conformational change [Oliver13]. A model of a catalytically active structure of the enzyme showed the presence of an ammonia tunnel and an interdomain salt bridge at its edge, between H186 in the glutaminase domain and E383 in the synthetase domain. Mutagenesis of these residues resulted in significantly reduced glutaminase activity and uncoupling of the two half reactions [Oliver14].

A crystal stucture of GMP synthetase has been solved at 2.2 Å resolution [Tesmer94, Tesmer96]. The enzyme from E. coli B is a dimer in solution [Sakamoto72], while the crystal structure of GMP synthetase contained a dimer of dimers.

Strains containing a guaA null mutation are auxotrophic for guanine [Lambden73, Joyce06]. A guaA null mutant contains modestly higher levels of xanthine in RNA, potentially interfering with RNA function [Pang12].

A guaA-disrupted mutant was used to shut down the XMP to GMP reaction in an inosine-producing strain of E. coli [Shimaoka06].

Reviews: [Raushel99, Zalkin98]

Citations: [Udaka63]

Gene Citations: [TesfaSelase92, Vales79, Shimada76, Hutchings00]

Locations: cytosol

Map Position: [2,628,980 <- 2,630,557] (56.66 centisomes, 204°)
Length: 1578 bp / 525 aa

Molecular Weight of Polypeptide: 58.679 kD (from nucleotide sequence)

Molecular Weight of Multimer: 126 kD (experimental) [Sakamoto72]

pI: 5.46

Unification Links: ASAP:ABE-0008254 , CGSC:657 , DIP:DIP-9852N , EchoBASE:EB0415 , EcoGene:EG10420 , EcoliWiki:b2507 , ModBase:P04079 , OU-Microarray:b2507 , PortEco:guaA , Pride:P04079 , Protein Model Portal:P04079 , RefSeq:NP_417002 , RegulonDB:EG10420 , SMR:P04079 , String:511145.b2507 , UniProt:P04079

Relationship Links: InterPro:IN-FAMILY:IPR001674 , InterPro:IN-FAMILY:IPR004739 , InterPro:IN-FAMILY:IPR014729 , InterPro:IN-FAMILY:IPR017926 , InterPro:IN-FAMILY:IPR022310 , InterPro:IN-FAMILY:IPR022955 , InterPro:IN-FAMILY:IPR025777 , InterPro:IN-FAMILY:IPR029062 , PDB:Structure:1GPM , Pfam:IN-FAMILY:PF00117 , Pfam:IN-FAMILY:PF00958 , Pfam:IN-FAMILY:PF02540 , Prosite:IN-FAMILY:PS51273 , Prosite:IN-FAMILY:PS51553

In Paralogous Gene Group: 9 (5 members)

Gene-Reaction Schematic: ?

Gene-Reaction Schematic

Genetic Regulation Schematic: ?

Genetic regulation schematic for guaA

GO Terms:

Biological Process: GO:0006177 - GMP biosynthetic process Inferred from experiment Inferred by computational analysis [UniProtGOA12, UniProtGOA11a, GOA06, GOA01a, Lambden73]
GO:0006164 - purine nucleotide biosynthetic process Inferred by computational analysis [UniProtGOA11a, GOA01a]
GO:0006541 - glutamine metabolic process Inferred by computational analysis [UniProtGOA11a, GOA06]
Molecular Function: GO:0003921 - GMP synthase activity Inferred from experiment [Abbott06]
GO:0003922 - GMP synthase (glutamine-hydrolyzing) activity Inferred from experiment Inferred by computational analysis [GOA06, GOA01, GOA01a, Deras99]
GO:0042802 - identical protein binding Inferred from experiment [Lasserre06]
GO:0042803 - protein homodimerization activity Inferred from experiment [Sakamoto72]
GO:0000166 - nucleotide binding Inferred by computational analysis [UniProtGOA11a]
GO:0005524 - ATP binding Inferred by computational analysis [UniProtGOA11a, GOA06, GOA01a]
GO:0016462 - pyrophosphatase activity Inferred by computational analysis [GOA01a]
GO:0016874 - ligase activity Inferred by computational analysis [UniProtGOA11a]
Cellular Component: GO:0005829 - cytosol Inferred from experiment Inferred by computational analysis [DiazMejia09, Ishihama08, Lasserre06]

MultiFun Terms: metabolism biosynthesis of building blocks nucleotides purine biosynthesis
metabolism central intermediary metabolism nucleotide and nucleoside conversions

Essentiality data for guaA 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 0.4% glucose No 37 Aerobic 7.2 0.27 No [Patrick07, Comment 3]
M9 medium with 1% glycerol No 37 Aerobic 7.2 0.35 No [Joyce06]
MOPS medium with 0.4% glucose No 37 Aerobic 7.2 0.22 No [Baba06, Comment 2]
No [Feist07, Comment 4]

Curated 04-Jan-2012 by Fulcher C , SRI International
Last-Curated ? 03-Apr-2015 by Keseler I , SRI International

Enzymatic reaction of: xanthosine-5'-phosphate:L-glutamine amido-ligase (AMP-forming) (GMP synthetase)

Synonyms: GMP synthetase (glutamine-hydrolysing), glutamine amidotransferase, GMP synthetase, XMP aminase, GMPS

EC Number:

L-glutamine + XMP + ATP + H2O <=> L-glutamate + GMP + AMP + diphosphate + 2 H+

The reaction direction shown, that is, A + B ↔ C + D versus C + D ↔ A + B, is in accordance with the direction in which it was curated.

The reaction is physiologically favored in the direction shown.

In Pathways: L-glutamine degradation I , superpathway of histidine, purine, and pyrimidine biosynthesis , superpathway of purine nucleotides de novo biosynthesis II , superpathway of guanosine nucleotides de novo biosynthesis II , guanosine ribonucleotides de novo biosynthesis

Early assays of the enzymatic activity (e.g. [Spector74, Patel77, Patel75, Zalkin77, Spector75, Sakamoto72, vonderSaal85]) were performed with enzyme purified from E. coli B 96, a purine-requiring derivative of E. coli B.

Cofactors or Prosthetic Groups: Mg2+ [vonderSaal85]

Inhibitors (Competitive): diphosphate [Spector75, Patel77] , N2-hydroxyguanosine 5'-monophosphate [Deras99]

Inhibitors (Irreversible): psicofuranine [Patel75, Patel77, vonderSaal85] , 6-diazo-5-oxonorleucine [Patel77, Chittur01]

Inhibitors (Unknown Mechanism): 3-bromopyruvate [Patel77] , 5,5'-dithio-bis-2-nitrobenzoate [Patel77] , tris [Patel77] , p-chloromercuribenzoate [Patel77] , acivicin [Chittur01]

Kinetic Parameters:

Km (μM)
kcat (sec-1)
kcat/Km (sec-1 μM-1)

pH(opt): 8.3 [Patel75]

Enzymatic reaction of: xanthosine-5'-phosphate:ammonia ligase (AMP-forming) (GMP synthetase)

Synonyms: xanthosine-5'-phosphate ammonia ligase, GMP synthetase, XMP aminase, GMP synthetase (ammonia dependent), xanthosine 5'-phosphate aminase

XMP + ammonium + ATP <=> GMP + AMP + diphosphate + 2 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.

Early assays of the enzymatic activity (e.g. [Spector74, Patel77, Patel75, Zalkin77, Spector75, Sakamoto72, vonderSaal85]) were performed with enzyme purified from E. coli B 96, a purine-requiring derivative of E. coli B. A variety of inhibitors of the enzyme were tested in [Spector75].

Inhibitors (Competitive): diphosphate [Spector75]

Inhibitors (Irreversible): 6-diazo-5-oxonorleucine [Patel77]

Inhibitors (Unknown Mechanism): decoyenine [Spector75] , adenosine [Spector75] , psicofuranine [Udaka63]

Kinetic Parameters:

Km (μM)
kcat (sec-1)
kcat/Km (sec-1 μM-1)

Sequence Features

Protein sequence of GMP synthetase with features indicated

Feature Class Location Attached Group Citations Comment
Sequence-Conflict 3  
[Tiedeman85, UniProt10a]
UniProt: (in Ref. 1; AA sequence);
Conserved-Region 9 -> 207  
UniProt: Glutamine amidotransferase type-1;
Active-Site 86  
UniProt: Nucleophile.
Active-Site 181  
Active-Site 183  
Conserved-Region 208 -> 400  
UniProt: GMPS ATP-PPase.
Nucleotide-Phosphate-Binding-Region 235 -> 241 ATP
UniProt: ATP.

Gene Local Context (not to scale): ?

Gene local context diagram

Transcription Units:

Transcription-unit diagram

Transcription-unit diagram


10/20/97 Gene b2507 from Blattner lab Genbank (v. M52) entry merged into EcoCyc gene EG10420; confirmed by SwissProt match.


Abbott06: Abbott JL, Newell JM, Lightcap CM, Olanich ME, Loughlin DT, Weller MA, Lam G, Pollack S, Patton WA (2006). "The Effects of Removing the GAT Domain from E. coli GMP Synthetase." Protein J 25;483-491. PMID: 17103135

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

Chittur01: Chittur SV, Klem TJ, Shafer CM, Davisson VJ (2001). "Mechanism for acivicin inactivation of triad glutamine amidotransferases." Biochemistry 40(4);876-87. PMID: 11170408

Deras99: Deras ML, Chittur SV, Davisson VJ (1999). "N2-hydroxyguanosine 5'-monophosphate is a time-dependent inhibitor of Escherichia coli guanosine monophosphate synthetase." Biochemistry 38(1);303-10. PMID: 9890911

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

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

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, 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."

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

Hutchings00: Hutchings MI, Drabble WT (2000). "Regulation of the divergent guaBA and xseA promoters of Escherichia coli by the cyclic AMP receptor protein." FEMS Microbiol Lett 187(2);115-22. PMID: 10856643

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

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

Lambden73: Lambden PR, Drabble WT (1973). "The gua operon of Escherichia coli K-12: evidence for polarity from guaB to guaA." J Bacteriol 115(3);992-1002. PMID: 4353875

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

Oliver13: Oliver JC, Linger RS, Chittur SV, Davisson VJ (2013). "Substrate activation and conformational dynamics of guanosine 5'-monophosphate synthetase." Biochemistry 52(31);5225-35. PMID: 23841499

Oliver14: Oliver JC, Gudihal R, Burgner JW, Pedley AM, Zwierko AT, Davisson VJ, Linger RS (2014). "Conformational changes involving ammonia tunnel formation and allosteric control in GMP synthetase." Arch Biochem Biophys 545;22-32. PMID: 24434004

Pang12: Pang B, McFaline JL, Burgis NE, Dong M, Taghizadeh K, Sullivan MR, Elmquist CE, Cunningham RP, Dedon PC (2012). "Defects in purine nucleotide metabolism lead to substantial incorporation of xanthine and hypoxanthine into DNA and RNA." Proc Natl Acad Sci U S A 109(7);2319-24. PMID: 22308425

Patel75: Patel N, Moyed HS, Kane JF (1975). "Xanthosine-5'-phosphate amidotransferase from Escherichia coli." J Biol Chem 250(7);2609-13. PMID: 235520

Patel77: Patel N, Moyed HS, Kane JF (1977). "Properties of xanthosine 5'-monophosphate-amidotransferase from Escherichia coli." Arch Biochem Biophys 1977;178(2);652-61. PMID: 189701

Patrick07: Patrick WM, Quandt EM, Swartzlander DB, Matsumura I (2007). "Multicopy suppression underpins metabolic evolvability." Mol Biol Evol 24(12);2716-22. PMID: 17884825

Raushel99: Raushel FM, Thoden JB, Holden HM (1999). "The amidotransferase family of enzymes: molecular machines for the production and delivery of ammonia." Biochemistry 38(25);7891-9. PMID: 10387030

Sakamoto72: Sakamoto N, Hatfield GW, Moyed HS (1972). "Physical properties and subunit structure of xanthosine 5'-phosphate aminase." J Biol Chem 247(18);5880-7. PMID: 4560421

Shimada76: Shimada K, Fukumaki Y, Takagi Y (1976). "Expression of the guanine operon of Escherichia coli as analyzed by bacteriophage lambda induced mutations." Mol Gen Genet 147(2);203-8. PMID: 787758

Shimaoka06: Shimaoka M, Takenaka Y, Mihara Y, Kurahashi O, Kawasaki H, Matsui H (2006). "Effects of xapA and guaA disruption on inosine accumulation in Escherichia coli." Biosci Biotechnol Biochem 70(12);3069-72. PMID: 17151449

Spector74: Spector T, Miller RL, Fyfe JA, Krenitsky TA (1974). "GMP synthetase from Escherichia coli B-96. Interactions with substrate analogs." Biochim Biophys Acta 370(2);585-91. PMID: 4613384

Spector75: Spector T, Beacham LM (1975). "Guanosine monophosphate synthetase from Escherichia coli B-96. Inhibition by nucleosides." J Biol Chem 250(8);3101-7. PMID: 164459

TesfaSelase92: Tesfa-Selase F, Drabble WT (1992). "Regulation of the gua operon of Escherichia coli by the DnaA protein." Mol Gen Genet 1992;231(2);256-64. PMID: 1736096

Tesmer94: Tesmer JJ, Stemmler TL, Penner-Hahn JE, Davisson VJ, Smith JL (1994). "Preliminary X-ray analysis of Escherichia coli GMP synthetase: determination of anomalous scattering factors for a cysteinyl mercury derivative." Proteins 18(4);394-403. PMID: 8208731

Tesmer96: Tesmer JJ, Klem TJ, Deras ML, Davisson VJ, Smith JL (1996). "The crystal structure of GMP synthetase reveals a novel catalytic triad and is a structural paradigm for two enzyme families." Nat Struct Biol 3(1);74-86. PMID: 8548458

Tiedeman85: Tiedeman AA, Smith JM, Zalkin H (1985). "Nucleotide sequence of the guaA gene encoding GMP synthetase of Escherichia coli K12." J Biol Chem 1985;260(15);8676-9. PMID: 3894345

Udaka63: Udaka S, Moyed HS (1963). "Inhibition of parental and mutant xanthosine 5'-phosphate aminases by psicofuranine." J Biol Chem 238;2797-803. PMID: 14063305

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

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

UniProt12a: UniProt Consortium (2012). "UniProt version 2012-02 released on 2012-02-29 00:00:00." Database.

UniProt15: UniProt Consortium (2015). "UniProt version 2015-01 released on 2015-01-16 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."

Vales79: Vales LD, Chase JW, Murphy JB (1979). "Orientation of the guanine operon of Escherichia coli K-12 by utilizing strains containing guaB-xse and guaB-upp deletions." J Bacteriol 139(1);320-2. PMID: 222730

vonderSaal85: von der Saal W, Crysler CS, Villafranca JJ (1985). "Positional isotope exchange and kinetic experiments with Escherichia coli guanosine-5'-monophosphate synthetase." Biochemistry 24(20);5343-50. PMID: 3907701

Zalkin77: Zalkin H, Truitt CD (1977). "Characterization of the glutamine site of Escherichia coli guanosine 5'-monophosphate synthetase." J Biol Chem 252(15);5431-6. PMID: 18463

Zalkin85: Zalkin H, Argos P, Narayana SV, Tiedeman AA, Smith JM (1985). "Identification of a trpG-related glutamine amide transfer domain in Escherichia coli GMP synthetase." J Biol Chem 260(6);3350-4. PMID: 2982857

Zalkin98: Zalkin H, Smith JL (1998). "Enzymes utilizing glutamine as an amide donor." Adv Enzymol Relat Areas Mol Biol 72;87-144. PMID: 9559052

Other References Related to Gene Regulation

Andrews88: Andrews SC, Guest JR (1988). "Nucleotide sequence of the gene encoding the GMP reductase of Escherichia coli K12." Biochem J 1988;255(1);35-43. PMID: 2904262

Cho11a: Cho BK, Federowicz SA, Embree M, Park YS, Kim D, Palsson BO (2011). "The PurR regulon in Escherichia coli K-12 MG1655." Nucleic Acids Res 39(15);6456-64. PMID: 21572102

Davies96: Davies IJ, Drabble WT (1996). "Stringent and growth-rate-dependent control of the gua operon of Escherichia coli K-12." Microbiology 1996;142 ( Pt 9);2429-37. PMID: 8828209

Fukumaki77: Fukumaki Y, Shimada K, Takagi Y (1977). "Secondary promoter of the guanine operon of Escherichia coli K-12." J Bacteriol 131(2);685-8. PMID: 328493

Husnain08: Husnain SI, Thomas MS (2008). "The UP element is necessary but not sufficient for growth rate-dependent control of the Escherichia coli guaB promoter." J Bacteriol 190(7);2450-7. PMID: 18203835

Husnain08a: Husnain SI, Thomas MS (2008). "Downregulation of the Escherichia coli guaB promoter by FIS." Microbiology 154(Pt 6);1729-38. PMID: 18524927

Husnain09: Husnain SI, Busby SJ, Thomas MS (2009). "Downregulation of the Escherichia coli guaB promoter by upstream-bound cyclic AMP receptor protein." J Bacteriol 191(19);6094-104. PMID: 19633076

Kim13: Kim HJ, Hou BK, Lee SG, Kim JS, Lee DW, Lee SJ (2013). "Genome-wide analysis of redox reactions reveals metabolic engineering targets for D-lactate overproduction in Escherichia coli." Metab Eng 18;44-52. PMID: 23563322

Meng90: Meng LM, Kilstrup M, Nygaard P (1990). "Autoregulation of PurR repressor synthesis and involvement of purR in the regulation of purB, purC, purL, purMN and guaBA expression in Escherichia coli." Eur J Biochem 1990;187(2);373-9. PMID: 2404765

TesfaSelase96: Tesfa-Selase F, Drabble WT (1996). "Specific binding of DnaA protein to a DnaA box in the guaB gene of Escherichia coli K12." Eur J Biochem 241(2);411-6. PMID: 8917437

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Please cite the following article in publications resulting from the use of EcoCyc: Nucleic Acids Research 41:D605-12 2013
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