DB code: T00114

RLCP classification 1.13.30000.42 : Hydrolysis
3.123.90030.312 : Transfer
CATH domain 3.40.50.880 : Rossmann fold Catalytic domain
3.40.50.620 : Rossmann fold Catalytic domain
3.30.300.10 : GMP Synthetase; Chain A, domain 3
E.C. 6.3.5.2
CSA 1gpm
M-CSA 1gpm
MACiE M0234

CATH domain Related DB codes (homologues)
3.40.50.620 : Rossmann fold S00314 S00549 S00316 S00317 S00318 S00315 T00085 T00249 D00300 M00177 M00178 T00106
3.40.50.880 : Rossmann fold D00526 T00021 M00215

Uniprot Enzyme Name
UniprotKB Protein name Synonyms RefSeq MEROPS Pfam
P04079 GMP synthase {glutamine-hydrolyzing}
EC 6.3.5.2
Glutamine amidotransferase
GMP synthetase
GMPS
NP_417002.1 (Protein)
NC_000913.2 (DNA/RNA sequence)
YP_490735.1 (Protein)
NC_007779.1 (DNA/RNA sequence)
C26.957 (Cysteine)
PF00117 (GATase)
PF00958 (GMP_synt_C)
PF02540 (NAD_synthase)
[Graphical View]

KEGG enzyme name
GMP synthase (glutamine-hydrolysing)
GMP synthetase (glutamine-hydrolysing)
guanylate synthetase (glutamine-hydrolyzing)
guanosine monophosphate synthetase (glutamine-hydrolyzing)
xanthosine 5'-phosphate amidotransferase
guanosine 5'-monophosphate synthetase

UniprotKB: Accession Number Entry name Activity Subunit Subcellular location Cofactor
P04079 GUAA_ECOLI ATP + xanthosine 5''-phosphate + L-glutamine + H(2)O = AMP + diphosphate + GMP + L-glutamate. Homodimer.

KEGG Pathways
Map code Pathways E.C.
MAP00230 Purine metabolism
MAP00251 Glutamate metabolism
MAP00983 Drug metabolism - other enzymes

Compound table
Cofactors Substrates Products Intermediates
KEGG-id C00305 C00002 C00655 C00064 C00001 C00020 C00013 C00144 C00025
E.C.
Compound Magnesium ATP Xanthosine 5'-phosphate L-Glutamine H2O AMP Pyrophosphate GMP L-Glutamate
Type divalent metal (Ca2+, Mg2+) amine group,nucleotide amide group,nucleotide amino acids,amide group H2O amine group,nucleotide phosphate group/phosphate ion amide group,amine group,nucleotide amino acids,carboxyl group
ChEBI 18420
 18420
 		15422
 15422
 		15652
 15652
 		18050
 58359
 18050
 58359
 		15377
 15377
 		16027
 16027
 		29888
 29888
 		17345
 17345
 		16015
 16015
PubChem 888
 888
 		5957
 5957
 		73323
 73323
 		5961
 6992086
 5961
 6992086
 		22247451
 962
 22247451
 962
 		6083
 6083
 		1023
 21961011
 1023
 21961011
 		6804
 6804
 		33032
 44272391
 88747398
 33032
 44272391
 88747398
1gpmA01 Unbound Unbound Unbound Unbound Unbound Unbound Unbound Unbound
1gpmB01 Unbound Unbound Unbound Unbound Unbound Unbound Unbound Unbound
1gpmC01 Unbound Unbound Unbound Unbound Unbound Unbound Unbound Unbound
1gpmD01 Unbound Unbound Unbound Unbound Unbound Unbound Unbound Unbound
1gpmA02 Unbound Unbound Unbound Unbound Bound:AMP Bound:POP Unbound Unbound
1gpmB02 Bound:_MG Unbound Unbound Unbound Bound:AMP Bound:POP Unbound Unbound
1gpmC02 Bound:_MG Unbound Unbound Unbound Bound:AMP Bound:POP Unbound Unbound
1gpmD02 Bound:_MG Unbound Unbound Unbound Bound:AMP Bound:POP Unbound Unbound
1gpmA03 Unbound Unbound Unbound Unbound Unbound Analogue:PO4 Unbound Unbound
1gpmB03 Unbound Unbound Unbound Unbound Unbound Analogue:PO4 Unbound Unbound
1gpmC03 Unbound Unbound Unbound Unbound Unbound Analogue:PO4 Unbound Unbound
1gpmD03 Unbound Unbound Unbound Unbound Unbound Analogue:PO4 Unbound Unbound

Reference for Active-site residues
resource references E.C.
Swiss-prot;P04079, literature [6]

Active-site residues
PDB Catalytic residues Cofactor-binding residues Modified residues Main-chain involved in catalysis Comment
1gpmA01 CYS 86;HIS 181;GLU 183 GLY 59;TYR 87;GLY 88
1gpmB01 CYS 86;HIS 181;GLU 183 GLY 59;TYR 87;GLY 88
1gpmC01 CYS 86;HIS 181;GLU 183 GLY 59;TYR 87;GLY 88
1gpmD01 CYS 86;HIS 181;GLU 183 GLY 59;TYR 87;GLY 88
1gpmA02 SER 235;SER 240;LYS 381 ASP 239(Magnesium binding) GLY 236;GLY 237
1gpmB02 SER 235;SER 240;LYS 381 ASP 239(Magnesium binding) GLY 236;GLY 237
1gpmC02 SER 235;SER 240;LYS 381 ASP 239(Magnesium binding) GLY 236;GLY 237
1gpmD02 SER 235;SER 240;LYS 381 ASP 239(Magnesium binding) GLY 236;GLY 237
1gpmA03
1gpmB03
1gpmC03
1gpmD03

References for Catalytic Mechanism
References Sections No. of steps in catalysis
[1]
Fig.11, p.2646 2
[6]
Fig.1, p.75 2
[8]
Scheme 3

References
[1]
Resource
Comments
Medline ID
PubMed ID 6378670
Journal Fed Proc
Year 1984
Volume 43
Pages 2640-7
Authors Villafranca JJ
Title Use of 31P and 13C NMR to study enzyme mechanisms.
Related PDB
Related UniProtKB
[2]
Resource
Comments GATASE DOMAIN.
Medline ID 85131126
PubMed ID 2982857
Journal J Biol Chem
Year 1985
Volume 260
Pages 3350-4
Authors Zalkin H, Argos P, Narayana SV, Tiedeman AA, Smith JM
Title Identification of a trpG-related glutamine amide transfer domain in Escherichia coli GMP synthetase.
Related PDB
Related UniProtKB P04079
[3]
Resource
Comments
Medline ID
PubMed ID 3911001
Journal Methods Enzymol
Year 1985
Volume 113
Pages 273-8
Authors Zalkin H
Title GMP synthetase.
Related PDB
Related UniProtKB
[4]
Resource
Comments
Medline ID
PubMed ID 8208731
Journal Proteins
Year 1994
Volume 18
Pages 394-403
Authors Tesmer JJ, Stemmler TL, Penner-Hahn JE, Davisson VJ, Smith JL
Title Preliminary X-ray analysis of Escherichia coli GMP synthetase: determination of anomalous scattering factors for a cysteinyl mercury derivative.
Related PDB
Related UniProtKB
[5]
Resource
Comments
Medline ID
PubMed ID 8895556
Journal EMBO J
Year 1996
Volume 15
Pages 5125-34
Authors Rizzi M, Nessi C, Mattevi A, Coda A, Bolognesi M, Galizzi A
Title Crystal structure of NH3-dependent NAD+ synthetase from Bacillus subtilis.
Related PDB
Related UniProtKB
[6]
Resource
Comments X-RAY CRYSTALLOGRAPHY (2.2 ANGSTROMS).
Medline ID 96133732
PubMed ID 8548458
Journal Nat Struct Biol
Year 1996
Volume 3
Pages 74-86
Authors Tesmer JJ, Klem TJ, Deras ML, Davisson VJ, Smith JL
Title The crystal structure of GMP synthetase reveals a novel catalytic triad and is a structural paradigm for two enzyme families.
Related PDB 1gpm
Related UniProtKB P04079
[7]
Resource
Comments
Medline ID
PubMed ID 11395405
Journal Annu Rev Biochem
Year 2001
Volume 70
Pages 149-80
Authors Huang X, Holden HM, Raushel FM
Title Channeling of substrates and intermediates in enzyme-catalyzed reactions.
Related PDB
Related UniProtKB
[8]
Resource
Comments
Medline ID
PubMed ID 11170408
Journal Biochemistry
Year 2001
Volume 40
Pages 876-87
Authors Chittur SV, Klem TJ, Shafer CM, Davisson VJ
Title Mechanism for acivicin inactivation of triad glutamine amidotransferases.
Related PDB
Related UniProtKB

Comments
According to the literature [6], [7], separate reactions occur at the N-terminal glutaminase domain (CATH 3.40.50.880) and at the ATP pyrophosphatase domain (CATH 3.40.50.620).
At the N-terminal glutaminase domain, which contains a catalytic triad (Cys86/His181/Glu183) and an oxyanion hole (composed of mainchain amide of Gly59/Tyr88/Gly88), hydrolysis of the sidechain amide of glutamine occurs by the trypsin-like reaction mechanism (see [6] & [8]).
(1) Cys86 acts as a nucleophile to make an attack on the carbonyl carbon. His181 and Glu183 assist this reaction.
(2) The transient negative charge on the intermediate will be stabilized by the oxyanion hole.
(3) A water molecule, activated by His181, may hydrolyze the intermediate.
At the ATP pyrophosphatase domain, two successive transfer reactions, transfer of AMP to carbonyl oxygen (O2 atom) of the XMP base, and transfer of the purine nucleotide to amine of ammonia molecule, released by the N-terminal domain.
According to the active-site structure and the data of the homologous enzyme (S00315 in EzCatDB), the transfer reaction of AMP to the carbonyl oxygen of XMP proceeds, probably as follows:
(A1) Mg2+ ion, Ser235, Gly236, Gly237 and Ser240 from P-loop and Lys381 stabilizes the leaving pyrophosphate, by neutralizing the negative charges, and also activate the transferred group, alpha-phosphate of ATP, by enhancing the electrophilicity of the phosphate group through polarization.
(A2) The acceptor group, the carbonyl oxygen (O2) atom of the XMP base, makes a nucleophilic attack on the transferred group, the alpha-phosphate of ATP.
(A3) The Mg2+ ion stabilize the pentacovalent transition-state.
(A4) O2-Adenyl-XMP intermediate is formed, releasing the pyrophosphate.
The detailed mechanism of the transfer of the nucleotide to amine of the ammonia molecule has not been elucidated. However, it will probably proceeds as follows:
(B1) Some group must act as a general base, to deprotonate the ammonium ion. However, it is not clear which group will activate the ammonium ion. (Probably, the remaining pyrophosphate or Asp340, considering the structure)
(B2) The activated ammonium makes a nucleophilic attack on the C2 atom of the O2-adenyl-XMP intermediate, forming a tetrahedral transition-state adduct.
(B3) Some group must stabilize the tetrahedral transition-state. However, it is not clear which group is involved in the stabilization. Mg2+ ion will stabilize the leaving phosphate of AMP.
(B4) Finally, GMP and AMP are formed.
More biochemical data will be required, to elucidate the mechanism.

Created Updated
2004-09-27 2009-02-26