Information on EC 6.3.5.7 - glutaminyl-tRNA synthase (glutamine-hydrolysing)

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

EC NUMBER
COMMENTARY
6.3.5.7
-
RECOMMENDED NAME
GeneOntology No.
glutaminyl-tRNA synthase (glutamine-hydrolysing)
REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT
LITERATURE
ATP + L-glutamyl-tRNAGln + L-glutamine = ADP + phosphate + L-glutaminyl-tRNAGln + L-glutamate
show the reaction diagram
-
-
-
-
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
Aminoacyl-tRNA biosynthesis
-
-
glutamate and glutamine metabolism
-
-
L-glutamine biosynthesis II (tRNA-dependent)
-
-
Metabolic pathways
-
-
SYSTEMATIC NAME
IUBMB Comments
glutamyl-tRNAGln:L-glutamine amido-ligase (ADP-forming)
In systems lacking discernible glutamine---tRNA ligase (EC 6.1.1.18), glutaminyl-tRNAGln is formed by a two-enzyme system. In the first step, a nondiscriminating ligase (EC 6.1.1.24, glutamate---tRNAGln ligase) mischarges tRNAGln with glutamate, forming glutamyl-tRNAGln. The glutamyl-tRNAGln is not used in protein synthesis until the present enzyme converts it into glutaminyl-tRNAGln (glutamyl-tRNAGlu is not a substrate for this reaction). Ammonia or asparagine can substitute for the preferred substrate glutamine.
SYNONYMS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
amidotransferase B
-
-
-
-
amidotransferase C
-
-
-
-
amidotransferase, glutamyl-transfer ribonucleate (glutamine-specific)
-
-
-
-
Glu-AdT
-
-
-
-
Glu-tRNAGln amidotransferase
-
-
-
-
Glu-tRNAGlnAT
-
-
-
-
glutamyl-tRNA(Gln) amidotransferase
-
-
-
-
glutamyl-tRNAGln amidotransferase
-
-
-
-
CAS REGISTRY NUMBER
COMMENTARY
52232-48-1
-
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
physiological function
-
inactivation of any of the hGatCAB subunits by siRNA-mediated knock down in HeLa cells leads to accumulation of the Glu-charged form of tRNAGln and defects in respiration can be observed
physiological function
Q9X0Z9
using gel mobility shift assays it is shown that formation of the glutamine transamidosome from Thermotoga maritima, consists of tRNAGln, glutamyltRNA synthase (GluRS) and the heterotrimeric amidotransferase GatCAB. The tail body of GatCAB recognizes the outer corner of the L-shaped tRNAGln in a tRNAGln-specific manner. GatCAB is in the non-productive form: the catalytic body of GatCAB contacts that of GluRS and is located near the acceptor stem of tRNAGln, in an appropriate site to wait for the completion of Glu-tRNAGln formation by GluRS. Hinges are identified between the catalytic and anticodon-binding bodies of GluRS and between the catalytic and tail bodies of GatCAB, which allow both GluRS and GatCAB to adopt the productive and non-productive forms
SUBSTRATE
PRODUCT                      
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
LITERATURE
(Substrate)
COMMENTARY
(Product)
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
ATP + Asp-tRNAAsn + L-glutamine
ADP + phosphate + Asn-tRNAAsn + L-glutamate
show the reaction diagram
-
-
-
?
ATP + Asp-tRNAAsn + L-glutamine
ADP + phosphate + Asn-tRNAAsn + L-glutamate
show the reaction diagram
-
-
-
-
?
ATP + Asp-tRNAAsn + L-glutamine
ADP + phosphate + Asn-tRNAAsn + L-glutamate
show the reaction diagram
-
identity elements used by GatCAB to discriminate tRNAAsn from tRNAAsp. GatCAB specifically binds Asp-tRNAAsn. Therefore, modified nucleotides do not play an essential role in GatCAB discrimination of Asp-tRNAAsn from Asp-tRNAAsp
-
-
?
ATP + Asp-tRNAAsn + L-glutamine
ADP + phosphate + Asn-tRNAAsn + L-glutamate
show the reaction diagram
-
the enzyme transamidates Asp-tRNAAsn and Glu-tRNAGln with similar efficiency
-
-
?
ATP + Glu-tRNAGln + Asn
ADP + phosphate + Gln-tRNAGln + Asp
show the reaction diagram
-
-
-
-
?
ATP + Glu-tRNAGln + Asn
ADP + phosphate + Gln-tRNAGln + Asp
show the reaction diagram
-
Asn is much less effective as amide donor than glutamine
-
-
?
ATP + Glu-tRNAGln + L-asparagine
ADP + phosphate + Gln-tRNAGln + L-glutamate
show the reaction diagram
-
-
-
-
?
ATP + Glu-tRNAGln + L-glutamine
ADP + phosphate + Gln-tRNAGln + L-glutamate
show the reaction diagram
-
-
-
-
?
ATP + Glu-tRNAGln + L-glutamine
ADP + phosphate + Gln-tRNAGln + L-glutamate
show the reaction diagram
-
-
-
?
ATP + Glu-tRNAGln + L-glutamine
ADP + phosphate + Gln-tRNAGln + L-glutamate
show the reaction diagram
-
-
-
?
ATP + Glu-tRNAGln + L-glutamine
ADP + phosphate + Gln-tRNAGln + L-glutamate
show the reaction diagram
-
-
-
-
?
ATP + Glu-tRNAGln + L-glutamine
ADP + phosphate + Gln-tRNAGln + L-glutamate
show the reaction diagram
-
-
-
?
ATP + Glu-tRNAGln + L-glutamine
ADP + phosphate + Gln-tRNAGln + L-glutamate
show the reaction diagram
-
-
-
-
?
ATP + Glu-tRNAGln + L-glutamine
ADP + phosphate + Gln-tRNAGln + L-glutamate
show the reaction diagram
-
-
-
-
?
ATP + Glu-tRNAGln + L-glutamine
ADP + phosphate + Gln-tRNAGln + L-glutamate
show the reaction diagram
O43716, O75879, Q9H0R6
-
-
-
?
ATP + Glu-tRNAGln + L-glutamine
ADP + phosphate + Gln-tRNAGln + L-glutamate
show the reaction diagram
-
Ser176A is the active-site nucleophile for facilitating Gln hydrolysis by the enzyme
-
?
ATP + Glu-tRNAGln + L-glutamine
ADP + phosphate + Gln-tRNAGln + L-glutamate
show the reaction diagram
-
the amidation of Glu-tRNAGln proceeds via a gamma-phosphorylated intermediate
-
-
?
ATP + Glu-tRNAGln + L-glutamine
ADP + phosphate + Gln-tRNAGln + L-glutamate
show the reaction diagram
-
organisms lacking Gln-tRNA synthetase produce Gln-tRNAGln from misacylated Glu-tRNAGln through the transamidation activity of Glu-tRNAGln amidotransferase. The enzyme hydrolyzes Gln to Glu and NH3, using the latter product to transamidate Glu-tRNAGln in concert with ATP hydrolysis
-
?
ATP + Glu-tRNAGln + L-glutamine
ADP + phosphate + Gln-tRNAGln + L-glutamate
show the reaction diagram
-
the enzyme produces Gln-tRNAGln required for plastidal protein biosynthesis
-
-
?
ATP + Glu-tRNAGln + L-glutamine
ADP + phosphate + Gln-tRNAGln + L-glutamate
show the reaction diagram
-
disruption of this operon is lethal. Transamidation is the only pathway to Gln-tRNAGln in Bacillus subtilis. The enzyme furnishes a means for formation of correctly charged Gln-tRNAGln through the transamidation of misacylated Glu-tRNAGln, functionally replacing the lack of glutaminyl-tRNA synthetase activity in Gram-positive eubacteria, cyanobacteria, archaea and organelles
-
-
?
ATP + Glu-tRNAGln + L-glutamine
ADP + phosphate + Gln-tRNAGln + L-glutamate
show the reaction diagram
-
GatDE is a heterodimeric amidotransferase. GatD acts as a glutaminase but only in the presence of both Glu-tRNAGln and the other subunit, GatE. The fact that only Glu-tRNAGln but not tRNA Gln could activate the glutaminase activity of GatD suggests that glutamine hydrolysis is coupled tightly to transamidation. GatE is a Glu-tRNAGln kinase that activates Glu-tRNAGln via gamma-phosphorylation
-
-
?
ATP + Glu-tRNAGln + L-glutamine
ADP + phosphate + Gln-tRNAGln + L-glutamate
show the reaction diagram
-
the enzyme transamidates Asp-tRNAAsn and Glu-tRNAGln with similar efficiency. GatCAB uses the amide donor glutamine 129fold more efficiently than asparagine
-
-
?
ATP + Glu-tRNAGln + L-glutamine + H2O
ADP + phosphate + Gln-tRNAGln + L-glutamate
show the reaction diagram
-
-
-
-
?
ATP + Glu-tRNAGln + L-glutamine + H2O
ADP + phosphate + Gln-tRNAGln + L-glutamate
show the reaction diagram
-
-
-
-
?
ATP + Glu-tRNAGln + NH4Cl
ADP + phosphate + Gln-tRNAGln + ?
show the reaction diagram
-
-
-
-
?
ATP + Glu-tRNAGln + NH4Cl
ADP + phosphate + Gln-tRNAGln + ?
show the reaction diagram
-
NH4Cl is much less effective as amide donor than glutamine
-
-
?
ATP + L-glutamate + tRNAGln1(UUG)
AMP + diphosphate + L-glutamyl-tRNAGln1(UUG)
show the reaction diagram
-
the enzyme is active toward the two tRNAGln isoacceptors, but with a significant catalytic preference for tRNAGln2(CUG). The less active tRNAGln1(UUG) responds to the less common CAA codon for Gln
-
-
?
ATP + L-glutamate + tRNAGln2(CUG)
AMP + diphosphate + L-glutamyl-tRNAGln2(CUG)
show the reaction diagram
-
the enzyme is active toward the two tRNAGln isoacceptors, but with a significant catalytic preference for tRNAGln2(CUG). The less active tRNAGln1(UUG) responds to the less common CAA codon for Gln. The wild-type enzyme shows a 24fold catalytic preference for tRNAGln2 over tRNAGlu
-
-
?
ATP + L-glutamate + tRNAGlu
AMP + diphosphate + L-glutamyl-tRNAGlu
show the reaction diagram
-
-
-
-
?
ATP-gammaS + Glu-tRNAGln + L-glutamine
? + phosphate + Gln-tRNAGln + L-glutamate
show the reaction diagram
-
-
-
?
additional information
?
-
-
in absence of the amido acceptor, Glu-tRNAGln, the enzyme has basal glutaminase activity that is unaffected by ATP
-
-
-
additional information
?
-
-
the enzyme possesses low glutaminase activity
-
-
-
additional information
?
-
-
the association of archaeal glutamyl-tRNA synthetase (ND-GluRS) with GatDE sequesters the tRNA synthetase for Gln-tRNAGln formation, with GatDE reducing the affinity of glutamyl-tRNA synthetase (ND-GluRS) for tRNAGlu 13fold
-
-
-
NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
LITERATURE
(Substrate)
COMMENTARY
(Product)
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
ATP + Asp-tRNAAsn + L-glutamine
ADP + phosphate + Asn-tRNAAsn + L-glutamate
show the reaction diagram
-
-
-
-
?
ATP + Glu-tRNAGln + L-glutamine
ADP + phosphate + Gln-tRNAGln + L-glutamate
show the reaction diagram
-
-
-
-
?
ATP + Glu-tRNAGln + L-glutamine
ADP + phosphate + Gln-tRNAGln + L-glutamate
show the reaction diagram
-
organisms lacking Gln-tRNA synthetase produce Gln-tRNAGln from misacylated Glu-tRNAGln through the transamidation activity of Glu-tRNAGln amidotransferase. The enzyme hydrolyzes Gln to Glu and NH3, using the latter product to transamidate Glu-tRNAGln in concert with ATP hydrolysis
-
-
?
ATP + Glu-tRNAGln + L-glutamine
ADP + phosphate + Gln-tRNAGln + L-glutamate
show the reaction diagram
-
the enzyme produces Gln-tRNAGln required for plastidal protein biosynthesis
-
-
?
ATP + Glu-tRNAGln + L-glutamine
ADP + phosphate + Gln-tRNAGln + L-glutamate
show the reaction diagram
-
disruption of this operon is lethal. Transamidation is the only pathway to Gln-tRNAGln in Bacillus subtilis. The enzyme furnishes a means for formation of correctly charged Gln-tRNAGln through the transamidation of misacylated Glu-tRNAGln, functionally replacing the lack of glutaminyl-tRNA synthetase activity in Gram-positive eubacteria, cyanobacteria, archaea and organelles
-
-
?
ATP + Glu-tRNAGln + L-glutamine + H2O
ADP + phosphate + Gln-tRNAGln + L-glutamate
show the reaction diagram
-
-
-
-
?
ATP + Glu-tRNAGln + L-glutamine + H2O
ADP + phosphate + Gln-tRNAGln + L-glutamate
show the reaction diagram
-
-
-
-
?
COFACTOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
Mg2+
-
required
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
(1R,2R)-1-(4-methylsulfonylphenyl)-2-(S-dioxo-L-methioneamido)-1,3-propanediol
-
-
(1R,2R)-1-(4-nitrophenyl)-2-(S-dioxo-L-methioneamido)-1,3-propanediol
-
-
(1R,2R)-1-phenyl-2-(S-dioxo-L-methioneamido)-1,3-propanediol
-
-
(1R,2S)-1-(4-nitrophenyl)-2-(S-dioxo-L-methioneamido)-1,3-propanediol
-
-
(1S,2R)-1-(4-nitrophenyl)-2-(S-dioxo-L-methioneamido)-1,3-propanediol
-
-
(1S,2S)-1-(4-nitrophenyl)-2-(S-dioxo-L-methioneamido)-1,3-propanediol
-
-
2'-O-(trinitrophenyl)adenosine 5'-triphosphate
-
IC50: 2.4 mM
3'-(L-alpha-aspartylamino)-3'-deoxy-N,N-dimethyladenosine
-
puromycin analogue
3'-deoxy-3'-(L-alpha-glutamylamino)-N,N-dimethyladenosine
-
puromycin analogue
3'-deoxy-3'-(L-glutaminylamino)-N,N-dimethyladenosine
-
puromycin analogue
3'-O-(trinitrophenyl)adenosine 5'-triphosphate
-
IC50: 2.4 mM
3'-[[(2S)-2-amino-4-(methylsulfinyl)butanoyl]amino]-3'-deoxy-N,N-dimethyladenosine
-
puromycin analogue
3'-[[(2S)-2-amino-4-(methylsulfonyl)butanoyl]amino]-3'-deoxy-N,N-dimethyladenosine
-
puromycin analogue
3'-[[(2S)-4-carboxy-2-hydroxybutanoyl]amino]-3'-deoxy-N,N-dimethyladenosine
-
puromycin analogue
3'-[[2-amino-4-(methylphosphinato)butanoyl]amino]-3'-deoxy-N,N-dimethyladenosine
-
puromycin analogue
6-Diazo-5-oxonorleucine
-
blocking of glutamine-dependent reaction, no inhibition of ammonia-dependent reaction
adenosine 5'-[beta,gamma-methylene]triphosphate
-
IC50: 2.3 mM
ADP
-
IC50: 0.026 mM
ATP-gammaS
-
IC50: 0.19 mM
Chloramphenicol
-
-
gamma-Glu boronic acid
-
IC50: 0.0016 mM
puromycin
-
aminonucleoside antibiotic produced by Streptomyces alboniger, very weak inhibitor of AdT
L-methionine-S-sulfoximine
-
at 1 mM, 3 mM or 5 mM, 20% inhibition
additional information
-
no inhibition by 6-diazo-5-oxonorleucine
-
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
additional information
-
activation of glutaminase activity by ATP or ATP-gammaS together with Glu-tRNAGln, results either from an allosteric effect due simply to binding of these analogues to the enzyme or from some structural changes that attend ATP or ATP-gammaS hydrolysis
-
additional information
-
the presence of 0.0066 mM GluRS2 increases the KM of the enzyme for Glu-tRNAGln nearly 4fold
-
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.0224
Asn
-
37C, pH 7.2, amidotransferase activity
0.1 - 0.2
Asn
-
-
0.00095
Asp-tRNAAsn
-
37C, pH 7.2, amidotransferase activity
0.00118
Asp-tRNAGln
-
37C, pH 7.2, amidotransferase activity
0.2068
ATP
-
37C, pH 7.2, amidotransferase activity
0.01
Gln
-
-
0.0207
Gln
-
37C, pH 7.2, amidotransferase activity
0.0402
Gln
-
37C, cosubstrate Asp-tRNAAsn + ATP, glutaminase activity
0.0509
Gln
-
37C, cosubstrate Glu-tRNAGln + ATP, glutaminase activity
0.00042
Glu-tRNAGln
-
in the absence of GluRS2, in 100 mM Na-HEPES pH 7.2, 30 mM KCl, 12 mM MgCl2, at 37C
0.00097
Glu-tRNAGln
-
in the presence of equimolar amounts of GluRS2, in 100 mM Na-HEPES pH 7.2, 30 mM KCl, 12 mM MgCl2, at 37C
0.00134
Glu-tRNAGln
-
in the presence of 440fold excess of GluRS2, in 100 mM Na-HEPES pH 7.2, 30 mM KCl, 12 mM MgCl2, at 37C
0.0017
Glu-tRNAGln
-
37C, pH not specified in the publication
0.0023
Glu-tRNAGln
-
37C, pH not specified in the publication, in the presence of glutamyl-tRNA synthetase (0.002 mM)
0.0024
Glu-tRNAGln
-
37C, pH not specified in the publication, in the presence of bovine serum albumin (0.002 mM)
0.00539
tRNAGln1(UUG)
-
pH 7.2, 37C
-
0.00133
tRNAGln2(CUG)
-
pH 7.2, 37C
-
0.00361
tRNAGlu
-
pH 7.2, 37C
0.0002
L-glutamyl-tRNAGlu
-
-
additional information
additional information
-
aminoacylation kinetics of enzyme variants
-
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.027
Asn
-
37C, pH 7.2, amidotransferase activity
1.3
Asp-tRNAAsn
-
37C, pH 7.2, amidotransferase activity
3.61
Asp-tRNAGln
-
37C, pH 7.2, amidotransferase activity
6.1
ATP
-
37C, pH 7.2, amidotransferase activity
0.052 - 2.1
Gln
-
37C, pH 7.2, cosubstrate: Glu-tRNAGln
3 - 6
Gln
-
37C, pH 7.2, amidotransferase activity
3.49
Gln
-
37C, pH 7.2, amidotransferase activity
10.29
Gln
-
37C, pH 7.2, cosubstrate: Glu-tRNAGln
11.8
Gln
-
37C, pH 7.2, cosubstrate Asp-tRNAAsn + ATP, glutaminase activity
0.21
Glu-tRNAGln
-
in the absence of GluRS2, in 100 mM Na-HEPES pH 7.2, 30 mM KCl, 12 mM MgCl2, at 37C; in the presence of 440fold excess of GluRS2, in 100 mM Na-HEPES pH 7.2, 30 mM KCl, 12 mM MgCl2, at 37C
0.23
Glu-tRNAGln
-
in the presence of equimolar amounts of GluRS2, in 100 mM Na-HEPES pH 7.2, 30 mM KCl, 12 mM MgCl2, at 37C
0.9
Glu-tRNAGln
-
37C, pH not specified in the publication; 37C, pH not specified in the publication, in the presence of bovine serum albumin (0.002 mM)
0.006
tRNAGln1(UUG)
-
pH 7.2, 37C
-
0.41
tRNAGln2(CUG)
-
pH 7.2, 37C
-
0.04
tRNAGlu
-
pH 7.2, 37C
1.2
Glu-tRNAGln
-
37C, pH not specified in the publication, in the presence of glutamyl-tRNA synthetase (0.002 mM)
additional information
additional information
-
aminoacylation kinetics of enzyme variants
-
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
400
Glu-tRNAGln
-
37C, pH not specified in the publication, in the presence of bovine serum albumin (0.002 mM)
4658
500
Glu-tRNAGln
-
37C, pH not specified in the publication
4658
1.1
tRNAGln1(UUG)
-
pH 7.2, 37C
0
308
tRNAGln2(CUG)
-
pH 7.2, 37C
0
11
tRNAGlu
-
pH 7.2, 37C
1988
600
Glu-tRNAGln
-
37C, pH not specified in the publication, in the presence of glutamyl-tRNA synthetase (0.002 mM)
4658
additional information
additional information
-
aminoacylation kinetics of enzyme variants
0
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.12
(1R,2R)-1-(4-methylsulfonylphenyl)-2-(S-dioxo-L-methioneamido)-1,3-propanediol
-
pH 7, temperature not specified in the publication
0.027
(1R,2R)-1-(4-nitrophenyl)-2-(S-dioxo-L-methioneamido)-1,3-propanediol
-
pH 7, temperature not specified in the publication
0.4
(1R,2R)-1-phenyl-2-(S-dioxo-L-methioneamido)-1,3-propanediol
-
pH 7, temperature not specified in the publication
0.37
(1R,2S)-1-(4-nitrophenyl)-2-(S-dioxo-L-methioneamido)-1,3-propanediol
-
pH 7, temperature not specified in the publication
2.8
(1S,2R)-1-(4-nitrophenyl)-2-(S-dioxo-L-methioneamido)-1,3-propanediol
-
pH 7, temperature not specified in the publication
0.16
(1S,2S)-1-(4-nitrophenyl)-2-(S-dioxo-L-methioneamido)-1,3-propanediol
-
pH 7, temperature not specified in the publication
0.134
3'-(L-alpha-aspartylamino)-3'-deoxy-N,N-dimethyladenosine
-
-
0.105
3'-deoxy-3'-(L-alpha-glutamylamino)-N,N-dimethyladenosine
-
-
0.045
3'-deoxy-3'-(L-glutaminylamino)-N,N-dimethyladenosine
-
-
0.011
3'-[[(2S)-2-amino-4-(methylsulfinyl)butanoyl]amino]-3'-deoxy-N,N-dimethyladenosine
-
-
0.004
3'-[[(2S)-2-amino-4-(methylsulfonyl)butanoyl]amino]-3'-deoxy-N,N-dimethyladenosine
-
-
0.13
3'-[[(2S)-4-carboxy-2-hydroxybutanoyl]amino]-3'-deoxy-N,N-dimethyladenosine
-
-
0.033
3'-[[2-amino-4-(methylphosphinato)butanoyl]amino]-3'-deoxy-N,N-dimethyladenosine
-
-
1.85
Chloramphenicol
-
pH 7, temperature not specified in the publication
4.1
puromycin
-
very weak inhibitor of AdT
IC50 VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
2.4
2'-O-(trinitrophenyl)adenosine 5'-triphosphate
-
IC50: 2.4 mM
2.4
3'-O-(trinitrophenyl)adenosine 5'-triphosphate
-
IC50: 2.4 mM
2.3
adenosine 5'-[beta,gamma-methylene]triphosphate
-
IC50: 2.3 mM
0.026
ADP
-
IC50: 0.026 mM
0.19
ATP-gammaS
-
IC50: 0.19 mM
0.0016
gamma-Glu boronic acid
-
IC50: 0.0016 mM
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY
GeneOntology No.
LITERATURE
SOURCE
PDB
SCOP
CATH
ORGANISM
Aquifex aeolicus (strain VF5)
Aquifex aeolicus (strain VF5)
Aquifex aeolicus (strain VF5)
Pseudomonas aeruginosa (strain ATCC 15692 / PAO1 / 1C / PRS 101 / LMG 12228)
Saccharomyces cerevisiae (strain ATCC 204508 / S288c)
Saccharomyces cerevisiae (strain ATCC 204508 / S288c)
Staphylococcus aureus (strain Mu50 / ATCC 700699)
Staphylococcus aureus (strain Mu50 / ATCC 700699)
Staphylococcus aureus (strain Mu50 / ATCC 700699)
Staphylococcus aureus (strain Mu50 / ATCC 700699)
Staphylococcus aureus (strain Mu50 / ATCC 700699)
Staphylococcus aureus (strain Mu50 / ATCC 700699)
Thermotoga maritima (strain ATCC 43589 / MSB8 / DSM 3109 / JCM 10099)
Thermotoga maritima (strain ATCC 43589 / MSB8 / DSM 3109 / JCM 10099)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
120000
-
gel filtration, glycerol density gradient sedimentation
492306
132000
O43716, O75879, Q9H0R6
gel filtration, GatABC complex; gel filtration, GatABC complex; gel filtration, GatABC complex
706514
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
dimer
-
2 * 63000, SDS-PAGE
heterotrimer
-
-
homodimer
-
gel filtration, GatDE forms a homodimer with the binding interface between the D-subunits
trimer
-
1 * 53000 + 1 * 53500 + 1 * 10900, SDS-PAGE
trimer
-
1 * subunit A plus 1 * subunit B plus 1 * subunit C
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
the glutamine transamidosome complex consisting of tRNAGln, glutamyltRNA synthase (GluRS) and the heterotrimeric amidotransferase GatCAB is crystalyzed at 3.35 A. Crystals are grown by the sitting-drop vapour-diffusion method at 20C
Q9X0Z9
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
overexpressed with pTrcgatCABBST in E. coli
-
DEAE-cellulose column chromatography, Ni-NTA resin column chromatography, and Superdex 200 gel filtration
-
using Ni-NTA chromatography
-
using heat treatment and column chromatography
Q9X0Z9
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expression in Escherichia coli
-
cloning of the three genes, gatC, gatA, and gatB, which constitute the transcriptional unit of the enzyme
-
expressed in Escherichia coli as a His-tagged fusion protein
-
overexpressed in Escherichia coli
Q9X0Z9
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
S128T
-
mutant protein retains significant glutaminase activity and transamidase activity in the presence of Gln
S152A
-
mutant is glutaminase inactive
S152T
-
mutant is glutaminase inactive
D178E
-
glutamine hydrolysis is negligible, Gln-tRNAGln formation is undetectable
D178N
-
glutamine hydrolysis is negligible, Gln-tRNAGln formation is undetectable
T101A
-
glutamine hydrolysis is negligible, Gln-tRNAGln formation is undetectable
T101S
-
hydrolyzes about 10% of glutamine compared to wild-type enzyme. Compared to wild-type enzyme, the mutant enzyme converts approximately half as much mischarged tRNA substrate to product
T177S
-
mutant enzyme hydrolyzes the same amount of glutamine as the wild-type enzyme. As the wild-type enzyme, the mutant enzyme transforms most of Glu-tRNAGln to Gln-tRNAGln
T177V
-
glutamine hydrolysis is negligible. Gln-tRNAGln formation is undetectable
K254E
-
glutamine hydrolysis is negligible, Gln-tRNAGln formation is undetectable
additional information
-
activity of the engineered enzyme variants indicates that the acceptor stem loop is the principle discrimination element because insertion of this loop alone enhances the specificity of the archaeal enzyme toward tRNAGln2
APPLICATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
analysis
-
simple system for monitoring the inhibition of GatCAB activity using Escherichia coli Top10 co-expressing the non-discriminating glutamyl-tRNA synthetase ndGluRS and GatCAB genes from Staphylococcus aureus Mu50. Growth repression is confirmed by introducing ndgluRS from Staphylococcus aureus Mu50 into Escherichia coli. Co-expression of the gatCAB operon alleviates growth repression in the host Escherichia coli. The screening system consists of these two transformants and non-expressing Escherichia coli Top10. The transformant harbors both ndGluRS gene and GatCAB operon can be co-expressed in the presence and in the absence of chemical compounds of interest. There is no inhibitor that inactivates GatCAB activity, but upon expression of two inactive GatCAB deletion variants, GatCAB-10 and GatCAB-CHD, together with ndGluRS in Escherichia coli Top10, the cells show repressed growth as well as ndGluRS is expressed