Information on EC 6.3.5.3 - phosphoribosylformylglycinamidine synthase

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The expected taxonomic range for this enzyme is: Eukaryota, Bacteria, Archaea

EC NUMBER
COMMENTARY hide
6.3.5.3
-
RECOMMENDED NAME
GeneOntology No.
phosphoribosylformylglycinamidine synthase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
ATP + N2-formyl-N1-(5-phospho-D-ribosyl)glycinamide + L-glutamine + H2O = ADP + phosphate + 2-(formamido)-N1-(5-phospho-D-ribosyl)acetamidine + L-glutamate
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
amide group transfer
-
-
-
-
amination
-
-
-
-
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
5-aminoimidazole ribonucleotide biosynthesis I
-
-
5-aminoimidazole ribonucleotide biosynthesis II
-
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Biosynthesis of antibiotics
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Biosynthesis of secondary metabolites
-
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Metabolic pathways
-
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Purine metabolism
-
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superpathway of 5-aminoimidazole ribonucleotide biosynthesis
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purine metabolism
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SYSTEMATIC NAME
IUBMB Comments
N2-formyl-N1-(5-phospho-D-ribosyl)glycinamide:L-glutamine amido-ligase (ADP-forming)
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CAS REGISTRY NUMBER
COMMENTARY hide
9032-84-2
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
thermophilic archaeon
-
-
Manually annotated by BRENDA team
Murine gammaherpesvirus-68
virion tegument protein ORF75c
UniProt
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
Pigeon
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
5'-phosphoribosylformylglycinamide + ATP + L-glutamine + H2O
phosphoribosylformylglycinamidine + L-glutamate + ADP + phosphate
show the reaction diagram
5'-phosphoribosylformylglycinamide + L-glutamine + ATP
phosphoribosylformylglycinamidine + L-glutamate + ADP + phosphate
show the reaction diagram
ATP + 5'-phosphoribosylformylglycinamide + L-Gln
?
show the reaction diagram
ATP + 5'-phosphoribosylformylglycinamide + L-Gln + H2O
ADP + phosphate + 5'-phosphoribosylformylglycinamidine + L-Glu
show the reaction diagram
ATP + 5'-phosphoribosylformylglycinamide + NH4+ + H2O
ADP + phosphate + 5'-phosphoribosylformylglycinamidine
show the reaction diagram
ATP + carbocyclic-5'-phosphoribosylformylglycinamide + L-Gln + H2O
ADP + phosphate + carbocyclic-5'-phosphoribosylformylglycinamidine + L-Glu
show the reaction diagram
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(+/-)carbocyclic-5'-phosphoribosylformylglycinamide and (-)5'-phosphoribosylformylglycinamide at 10% of the activity relative to 5'-phosphoribosylformylglycinamide
-
-
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ATP + N2-formyl-N1-(5-phospho-D-ribosyl)glycinamide + L-glutamine + H2O
ADP + phosphate + 2-(formamido)-N1-(5-phospho-D-ribosyl)acetamidine + L-glutamate
show the reaction diagram
formylglycinamide ribonucleotide + ATP + L-glutamine + H2O
formylglycinamidine ribonucleotide + ADP + phosphate + L-glutamate
show the reaction diagram
formylglycinamide ribonucleotide + ATP + NH3 + H2O
formylglycinamidine ribonucleotide + ADP + phosphate
show the reaction diagram
-
-
-
-
?
additional information
?
-
NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
5'-phosphoribosylformylglycinamide + L-glutamine + ATP
phosphoribosylformylglycinamidine + L-glutamate + ADP + phosphate
show the reaction diagram
-
fourth step in the de novo purine biosynthetic pathway
-
r
ATP + 5'-phosphoribosylformylglycinamide + L-Gln
?
show the reaction diagram
ATP + N2-formyl-N1-(5-phospho-D-ribosyl)glycinamide + L-glutamine + H2O
ADP + phosphate + 2-(formamido)-N1-(5-phospho-D-ribosyl)acetamidine + L-glutamate
show the reaction diagram
formylglycinamide ribonucleotide + ATP + L-glutamine + H2O
formylglycinamidine ribonucleotide + ADP + phosphate + L-glutamate
show the reaction diagram
-
fourth step of purine biosynthetic pathway
-
-
?
additional information
?
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COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Co2+
-
can replace Mg2+ in activation
Mn2+
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can replace Mg2+ in activation
NH4+
-
can replace K+ as activator
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
6-diazo-5-oxo-L-norleucine
albizziin
beta-gamma-5'-Adenylyl methylene diphosphate
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Cyanate
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hydroxylamine
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-
iodoacetamide
iodoacetate
L-2-Amino-4-oxo-5-chloropentanoic acid
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O-diazoacetyl-L-Ser
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0468
(-)carbocyclic-5'-phosphoribosylformylglycinamidine
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-
0.06 - 0.1
5'-phosphoribosylformylglycinamide
0.18
alpha,beta-5'-phosphoribosylformylglycinamide
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ATPase activity
0.181 - 1.5
ATP
0.23
beta-5'-phosphoribosylformylglycinamide
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ATPase activity
0.507
formylglycinamide ribonucleotide
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formylglycinamide ribonucleotide amidotransferase complex
0.11 - 0.2
Gln
0.03
L-Gln
1.3
L-glutamine
-
formylglycinamide ribonucleotide amidotransferase complex
0.4
MgATP2-
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-
1 - 100
NH4+
additional information
additional information
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TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.05
NH4+
Escherichia coli
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-
additional information
additional information
Bacillus subtilis
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-
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SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.16
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-
additional information
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7
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activity with glutamine
7.4
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liver and hepatoma 3924A
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6.2 - 8
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about 50% of maximal activity at pH 6.2 and 8.0
6.5 - 7.5
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pH 6.5: about 40% of maximal activity, pH 7.5: about 35% of maximal activity, reaction with glutamine
6.8 - 7.5
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pH 6.8: about 50% of maximal activity, pH 7.5: about 65% of maximal activity, reaction with NH4+
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
-
enzyme activity is essential for male gametophytes
Manually annotated by BRENDA team
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16, 20, 44, 7787, 7777, 3924A, 5123tc
Manually annotated by BRENDA team
additional information
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
PDB
SCOP
CATH
ORGANISM
UNIPROT
Bacillus subtilis (strain 168)
Bacillus subtilis (strain 168)
Salmonella typhimurium (strain LT2 / SGSC1412 / ATCC 700720)
Salmonella typhimurium (strain LT2 / SGSC1412 / ATCC 700720)
Salmonella typhimurium (strain LT2 / SGSC1412 / ATCC 700720)
Salmonella typhimurium (strain LT2 / SGSC1412 / ATCC 700720)
Salmonella typhimurium (strain LT2 / SGSC1412 / ATCC 700720)
Salmonella typhimurium (strain LT2 / SGSC1412 / ATCC 700720)
Salmonella typhimurium (strain LT2 / SGSC1412 / ATCC 700720)
Salmonella typhimurium (strain LT2 / SGSC1412 / ATCC 700720)
Sulfolobus tokodaii (strain DSM 16993 / JCM 10545 / NBRC 100140 / 7)
Thermotoga maritima (strain ATCC 43589 / MSB8 / DSM 3109 / JCM 10099)
Thermotoga maritima (strain ATCC 43589 / MSB8 / DSM 3109 / JCM 10099)
Thermotoga maritima (strain ATCC 43589 / MSB8 / DSM 3109 / JCM 10099)
Thermotoga maritima (strain ATCC 43589 / MSB8 / DSM 3109 / JCM 10099)
Thermotoga maritima (strain ATCC 43589 / MSB8 / DSM 3109 / JCM 10099)
Thermotoga maritima (strain ATCC 43589 / MSB8 / DSM 3109 / JCM 10099)
Thermotoga maritima (strain ATCC 43589 / MSB8 / DSM 3109 / JCM 10099)
Thermotoga maritima (strain ATCC 43589 / MSB8 / DSM 3109 / JCM 10099)
Thermus thermophilus (strain HB8 / ATCC 27634 / DSM 579)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
24755
-
x * 24755, glutamine amide transfer subunit, + x * 80300, aminator subunit
80300
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x * 24755, glutamine amide transfer subunit, + x * 80300, aminator subunit
133000 - 135000
133000
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1 * 133000, SDS-PAGE. When stored in absence of a reducing agent, e.g. DTT or 2-mercaptoethanol, the enzyme undergoes polymerization
141418
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x * 141418, calculation from nucleotide sequence
additional information
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PurL exists in two forms: large PurL (lgPurL) is a single chain, multidomain enzyme of about 1300 amino acids, whereas small PurL (smPurL) contains about 800 amino acids but requires two additional gene products, PurS and PurQ, for activity
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
monomer
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1 * 133000, SDS-PAGE. When stored in absence of a reducing agent, e.g. DTT or 2-mercaptoethanol, the enzyme undergoes polymerization
additional information
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FGAR-AT is formed from three proteins: PurS, PurQ and smPurL. The stoichiometry of PurS/smPurL/PurQ is 2:1:1
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
determination of structure of PurS in two different crystal forms P2(1) and C2 at 2.5 and 2.0 A resolution, respectively. PurS forms a tight dimer with a central six-stranded beta-sheet flanked by four helices. In both the P2(1) and the C2 crystal forms, the quaternary structure of PurS is a tetramer. Native PurS is crystallized from 2.0 M ammonium sulfate, 6% PEG 400, 100 mM HEPES, pH 7.3. The crystals are grown at room temperature using the hanging-drop vapor diffusion technique. Drops containing a 2:1 mixture of protein and reservoir solution are optimal for crystal growth. Crystals grow over a period of 4-7 days. Under these conditions, PurS crystallizes in the monoclinic space group P2(1) with unit cell dimensions a = 42.91 A, b = 87.96 A, c = 52.68 A, and beta = 94.97°. The SeMet-PurS crystals grow in 20% PEG 4K, 5% glycerol, 15% 2-propanol, 100 mM sodium citrate, pH 5.6. Optimal drop size is 0.004 mL using a 1:1 mixture of protein and reservoir. Under these conditions, the protein crystallizes in the space group C2 with unit cell dimensions a = 89.13 A, b = 42.21 A, c = 47.03 A, and beta = 118.22°
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crucial component MTH169, space group P4(3)22, unit cell parameters a : 53.8 A, c : 142.7 A
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crystals of the pure SeMet-enzyme are grown at room temperature using hanging-drop vapor diffusion method. StPurL crystallizes in the hexagonal space group P6(5) with unit cell dimensions of a = 145.3 and c = 140.9 A. The asymmetric unit contains one monomer, corresponding to calculated solvent content of 57%. Domain organization revealed by X-ray crystallography
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mutants R1263A and F209W, vapor diffusion method
crystal structure of phosphoribosylformyl-glycinamidine synthase II, PurS subunit (TM1244) at 1.90 A resolution
hanging drop method, enzyme in complex with formylglycinamide ribonucleotide, mutant enzyme H72A in complex with beta,gamma-methylene adenosine 5'-triphosphate, mutant enzyme H72A in complex with ADP, enzyme in complex with formylglycinamide ribonucleotide and beta,gamma-methylene adenosine 5'-triphosphate, enzyme in complex with ATP
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in complex with gene products PurQ and PurS, to 3.5 A resolution. The complex has a stoichiometry of 2:1:1 for components PurS:PurQ:PurL, respectively. The flexibility of the PurS dimer is involved in the activation of the complex and the formation of an ammonia channel
nanodroplet vapor diffusion method, crystal structure of phosphoribosylformylglycinamidine synthase II (smPurL) from Thermotoga maritima at 2.15 A resolution
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pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6.5
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highest stability, rather unstable at pH 8
1667
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
42 - 80
the first unfolding transition at 42°C leads to a loss of 15-20% of the secondary structure content of the protein and 80% loss in activity. The second unfolding transition at 80°C represents unfolding of the bulk of the protein
50
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5 min, up to 98% loss of activity. A combination of Gln, MgCl2, and KCl protects against thermal inactivation
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
a combination of Gln, MgCl2, and KCl protects against thermal inactivation
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K+ required for stabilization
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loss of activity during freezing and thawing
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STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-30°C, 20 mM phosphate, pH 6.5, 1 mM Gln, 10% glycerol, stable for 2 months
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
attempts to purify the complex resulted in separation of the constituent proteins. PurS, smPurL, and PurQ are therefore separately expressed and purified to homogeneity
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wild-type and mutant enzymes expression in Escherichia coli
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expressed in Escherichia coli BL21DE3-834 (gold) cells
expression in Escherichia coli
expression in Escherichia coli B834(DE3)
MTH169 gene amplified by PCR using Mth genomic DNA as template, cloned into pET15b vector and expressed in Escherichia coli BL21 (DE3)
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P-ADE5 gene cloned from a genomic library of Pichia methanolica by the ade6 mutation complementation in the recipient Saccharomyces cerevisiae strain
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purL gene encoded aminator subunit, purQ gene encoded glutamine amide transfer subunit
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PurS, smPurL, and PurQ are separately expressed in Escherichia coli
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yexA gene cloned and expressed in Escherichia coli
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
F209W
both the secondary structure content and activity of the mutant enzyme is very similar to the wild type protein
L1181F
the mutation causes a 60% loss in activity
L1181W
the mutant has secondary structure content and enzyme at par with the wild type protein and exhibits almost no destabilization
L1181Y
the mutation results in failure to yield soluble protein
R1263A
the mutant is structurally identical to the the native protein
T683W
the mutation causes a 70% loss in activity
H32A
-
no synthesis of formylglycinamidine ribonucleotide detected
H32Q
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no synthesis of formylglycinamidine ribonucleotide detected
H72A
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mutant enzyme displays 1/20 of the wild type activity with a strongly increased Km for formylglycinamidine ribonucleotide
H72Q
-
mutant enzyme displayed 1/200 of the wild type activity
additional information
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