Information on EC 6.3.5.5 - carbamoyl-phosphate synthase (glutamine-hydrolysing)

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The enzyme appears in viruses and cellular organisms

EC NUMBER
COMMENTARY hide
6.3.5.5
-
RECOMMENDED NAME
GeneOntology No.
carbamoyl-phosphate synthase (glutamine-hydrolysing)
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
2 ATP + L-glutamine + hydrogencarbonate + H2O = 2 ADP + phosphate + L-glutamate + carbamoyl phosphate
show the reaction diagram
ATP + carbamate = ADP + carbamoyl phosphate
show the reaction diagram
(1d)
-
-
-
ATP + hydrogencarbonate = ADP + carboxyphosphate
show the reaction diagram
(1b)
-
-
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L-glutamine + H2O = L-glutamate + NH3
show the reaction diagram
(1a)
-
-
-
NH3 + carboxyphosphate = carbamate + phosphate
show the reaction diagram
(1c)
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-
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REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
amide group transfer
-
-
-
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amination
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-
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Phosphorylation
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PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
Alanine, aspartate and glutamate metabolism
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arginine metabolism
-
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L-arginine biosynthesis I (via L-ornithine)
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L-arginine biosynthesis II (acetyl cycle)
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L-arginine biosynthesis III (via N-acetyl-L-citrulline)
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L-arginine biosynthesis IV (archaebacteria)
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Metabolic pathways
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Pyrimidine metabolism
-
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UMP biosynthesis
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SYSTEMATIC NAME
IUBMB Comments
hydrogen-carbonate:L-glutamine amido-ligase (ADP-forming, carbamate-phosphorylating)
The product carbamoyl phosphate is an intermediate in the biosynthesis of arginine and the pyrimidine nucleotides [4]. The enzyme from Escherichia coli has three separate active sites, which are connected by a molecular tunnel that is almost 100 A in length [8]. The amidotransferase domain within the small subunit of the enzyme hydrolyses glutamine to ammonia via a thioester intermediate. The ammonia migrates through the interior of the protein, where it reacts with carboxyphosphate to produce the carbamate intermediate. The carboxyphosphate intermediate is formed by the phosphorylation of hydrogencarbonate by ATP at a site contained within the N-terminal half of the large subunit. The carbamate intermediate is transported through the interior of the protein to a second site within the C-terminal half of the large subunit, where it is phosphorylated by another ATP to yield the final product, carbamoyl phosphate [6]. cf. EC 6.3.4.16, carbamoyl-phosphate synthase (ammonia).
CAS REGISTRY NUMBER
COMMENTARY hide
37233-48-0
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
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Manually annotated by BRENDA team
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Manually annotated by BRENDA team
genes carA and carB encoding the small and large subunit of the CPSase
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-
Manually annotated by BRENDA team
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Manually annotated by BRENDA team
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Manually annotated by BRENDA team
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Manually annotated by BRENDA team
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Manually annotated by BRENDA team
NCIMB 10648
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Manually annotated by BRENDA team
NCIMB 10648
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Manually annotated by BRENDA team
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Manually annotated by BRENDA team
L814
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Manually annotated by BRENDA team
possess 2 carbamoyl-phosphate synthases, one belonging to the arginine pathway, CPSase-A, and another belonging to the pyrimidine pathway, CPSase-P
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-
Manually annotated by BRENDA team
Fresh-water teleost
-
-
-
Manually annotated by BRENDA team
Blackmous catshark
-
-
Manually annotated by BRENDA team
wild-type and mutant strain
Q8RSS4 and Q8RSS3
UniProt
Manually annotated by BRENDA team
HCV, gene CAD
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-
Manually annotated by BRENDA team
atlantic halibut, yolk-sac larvae and adult, two enzymes: CPS II and CPS III
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-
Manually annotated by BRENDA team
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-
-
Manually annotated by BRENDA team
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SwissProt
Manually annotated by BRENDA team
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Manually annotated by BRENDA team
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Manually annotated by BRENDA team
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Manually annotated by BRENDA team
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Manually annotated by BRENDA team
carB; DSM20617, gene carB
UniProt
Manually annotated by BRENDA team
MTCC1
-
-
Manually annotated by BRENDA team
MTCC1
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-
Manually annotated by BRENDA team
-
SwissProt
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
metabolism
-
the enzyme is organized as CAD, i.e. carbamoyl-phosphate synthetase 2, aspartate transcarbamylase, and dihydroorotase, a tripartite enzyme that catalyzes the first three steps of pyrimidine biosynthesis
physiological function
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
2 ATP + L-Gln + HCO3- + H+ + H2O
2 ADP + phosphate + L-glutamate + carbamoyl phosphate
show the reaction diagram
2 ATP + L-glutamine + HCO3- + H2O
2 ADP + phosphate + L-glutamate + carbamoyl phosphate
show the reaction diagram
2 ATP + L-glutamine + HCO3- + H2O + H+
2 ADP + phosphate + L-glutamate + carbamoyl phosphate
show the reaction diagram
ATP + gamma-glutamyl hydrazide + HCO3-
?
show the reaction diagram
-
-
-
?
ATP + gamma-glutamyl hydroxamate + HCO3-
?
show the reaction diagram
-
-
-
?
ATP + hydrazine + HCO3-
ATP + phosphate + N-amino carbamoyl phosphate
show the reaction diagram
-
-
-
?
ATP + hydroxylamine + HCO3-
ATP + phosphate + N-hydroxy carbamoyl phosphate
show the reaction diagram
-
-
-
?
ATP + L-Gln + HCO3-
?
show the reaction diagram
ATP + L-Gln + HCO3-
ADP + phosphate + L-Glu + carbamoyl phosphate
show the reaction diagram
ATP + L-glutamine + HCO3- + H2O
2 ADP + phosphate + L-glutamate + carbamoyl phosphate
show the reaction diagram
-
there is no preferential partitioning of carbamoyl phosphate between the arginine and pyrimidine biosynthetic pathways. Channeling must occur during the dynamic association of coupled enzymes pairs. The interaction of carbamoyl-phosphate synthetase/aspartate transcarbamoylase is demonstrated by the unexpectedly weak inhibition of the coupled reaction by the bisubstrate analog, N-(phosphonacetyl)-L-aspartate. In the coupled reaction, the effective concentration of carbamoyl phosphate in the vicinity of the aspartate transcarbamoylase active site is 96fold higher than the concentration in the bulk phase. Channeling probably plays an essential role in protecting this very unstable intermediate of metabolic pathways performing at extreme temperatures
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-
?
ATP + L-glutamine + HCO3- + H2O
ADP + phosphate + L-glutamate + carbamoyl phosphate
show the reaction diagram
ATP + NH4+ + HCO3-
ADP + phosphate + carbamoyl phosphate
show the reaction diagram
additional information
?
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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
2 ATP + L-Gln + HCO3- + H+ + H2O
2 ADP + phosphate + L-glutamate + carbamoyl phosphate
show the reaction diagram
-
the product carbamoyl phosphate is utilized in the pyrimidine and arginine biosynthetic pathways
-
?
2 ATP + L-glutamine + HCO3- + H2O
2 ADP + phosphate + L-glutamate + carbamoyl phosphate
show the reaction diagram
2 ATP + L-glutamine + HCO3- + H2O + H+
2 ADP + phosphate + L-glutamate + carbamoyl phosphate
show the reaction diagram
ATP + L-Gln + HCO3-
?
show the reaction diagram
ATP + L-Gln + HCO3-
ADP + phosphate + L-Glu + carbamoyl phosphate
show the reaction diagram
ATP + L-glutamine + HCO3- + H2O
2 ADP + phosphate + L-glutamate + carbamoyl phosphate
show the reaction diagram
-
there is no preferential partitioning of carbamoyl phosphate between the arginine and pyrimidine biosynthetic pathways. Channeling must occur during the dynamic association of coupled enzymes pairs. The interaction of carbamoyl-phosphate synthetase/aspartate transcarbamoylase is demonstrated by the unexpectedly weak inhibition of the coupled reaction by the bisubstrate analog, N-(phosphonacetyl)-L-aspartate. In the coupled reaction, the effective concentration of carbamoyl phosphate in the vicinity of the aspartate transcarbamoylase active site is 96fold higher than the concentration in the bulk phase. Channeling probably plays an essential role in protecting this very unstable intermediate of metabolic pathways performing at extreme temperatures
-
-
?
ATP + L-glutamine + HCO3- + H2O
ADP + phosphate + L-glutamate + carbamoyl phosphate
show the reaction diagram
additional information
?
-
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
AMP
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enhances activity
GDP
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enhances activity
GMP
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enhances activity
IDP
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enhances activity
MgUTP2-
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in presence of high concentrations of dimethyl sulfoxide, MgUTP2- stimulates at low concentrations, while it is inhibitory at higher concentrations
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Mn2+
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maximal activity at concentration of Mn2+ approximately equal to the ATP concentration
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
6-diazo-5-oxonorleucine
acivicin
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selective time-dependent inhibition of L-Gln-dependent activity, L-Gln protects the enzyme from inactivation. Stimulation of NH4+-dependent activity
ADP
-
competitive with MgATP2-
Ala
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inhibits glutamine hydrolysis and Gln-dependent carbamoyl phosphate synthesis
Alkyl hydrazines
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inhibits Gln-dependent activity, but not NH4+-dependent activity
Arg
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arginine-specific carbamoyl-phosphate synthase is repressed, pyrimidine-specific enzyme not
arginine
Q8RSS4 and Q8RSS3
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azaserine
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selective inactivation of Gln-dependent activity
Carbamoyl phosphate
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guanidine hydrochloride
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H2O2
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0.2 mM, inhibits Gln-dependent activity. No effect on the activity with NH4+ in carbamoyl-phosphate synthase reaction
HCO3-
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20 mM
hydroxylamine
Hydroxymercuribenzoate
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L-2-Amino-4-oxo-5-chloropentanoate
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selective inactivation of Gln-dependent activity
L-Cyanate
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inhibits Gln-dependent activity, but not NH4+-dependent activity
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leflunomide
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Mersalyl
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MgATP2-
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product inhibition
MgUTP2-
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in presence of high concentrations of dimethyl sulfoxide, MgUTP2- at low concentrations stimulates, while it is inhibitory at higher concentrations
N-(phosphonacetyl)-L-aspartate
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there is no preferential partitioning of carbamoyl phosphate between the arginine and pyrimidine biosynthetic pathways. Channeling must occur during the dynamic association of coupled enzymes pairs. The interaction of carbamoyl-phosphate synthetase/aspartate transcarbamoylase is demonstrated by the unexpectedly weak inhibition of the coupled reaction by the bisubstrate analog, N-(phosphonacetyl)-L-aspartate
NEM
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irreversible inactivation of synthetase activity. Increase of glutaminase activity
P1,P5-di(adenosine 5')-pentaphosphate
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inhibits two partial reactions catalyzed by the enzyme: bicarbonate-dependent ATPase and ATP synthesis from carbamoyl phosphate
phosphate
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Polyamines
putrescine
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spermidine
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spermine
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UDPglucose
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Uracil
Q8RSS4 and Q8RSS3
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additional information
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
5-phospho-alpha-D-ribose 1-diphosphate
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activates wild-type enzyme
5-phospho-alpha-D-ribosyl 1-diphosphate
acetylornithine
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activates
acivicin
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selective time-dependent inhibition of L-Gln-dependent activity, L-Gln protects the enzyme from inactivation. Stimulation of NH4+-dependent activity
ammonia
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-
GTP
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enhances activity
human Rad9 checkpoint protein
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stimulates the carbamoyl phosphate synthetase activity of the multifunctional protein CAD. Rad9 binds to the Cpsase domain, and, moreover, this binding results in a 2fold stimulation of the CPSase activity of CAD
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ITP
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enhances activity
L-ornithine
N-acetyl-L-glutamate
N-acetylglutamate
NEM
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250fold activation of glutaminase activity. Irreversible inactivation of synthetase activity
ornithine
phosphate
phosphoribosyl 5'-diphosphate
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activation of Escherichia coli and chimeric enzyme, the presence of UTP reduces the PRPP binding
thiol
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required for Gln-dependent activity
XMP
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enhances activity
additional information
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.022 - 0.53
ADP
2.2 - 111
ammonia
0.004 - 8.36
ATP
0.11 - 0.15
Gln
0.64 - 32.5
HCO3-
0.0119 - 22
L-Gln
0.04 - 18.87
L-glutamine
1.7
MgATP2-
9.8 - 160
NH4+
additional information
additional information
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.01 - 0.32
ADP
5.1
ammonia
Escherichia coli
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-
0.006 - 2500
ATP
0.1 - 4.2
Gln
0.005 - 3.1
L-Gln
0.53 - 2660
L-glutamine
0.097 - 5.8
NH4+
additional information
additional information
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0018 - 0.005
UMP
IC50 VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.004 - 0.2
UMP
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.00035
Q8RSS4 and Q8RSS3
mutant starin S36K
0.001125
Q8RSS4 and Q8RSS3
wild-type strain
0.041
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recombinant enzyme
0.348
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recombinant mutant P909C/G919C
0.472
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recombinant mutant P909C
0.524
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recombinant mutant G919C
0.855
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recombinant wild-type enzyme
additional information
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6.6
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mutant enzyme E841K
6.7
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highest activity in the pH-range 6.7-8.8
6.8 - 7.2
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Gln-dependent activity
7.5
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with either 5 mM Gln or 100 mM NH4+ as substrate
7.8 - 8.2
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7.8
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NH4+-dependent activity
9.3
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wild-type enzyme
9.5
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and a second optimum at pH 4.2, glutaminase activity
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6.7 - 8.5
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6.7: highest activity in the pH-range 6.7-8.5, pH 7.2: 95% of the activity at pH 6.7, pH 8.5: 64% of the activity at pH 6.7
7.5 - 8.5
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7.5: about 65% of maximal activity, 8.5: about 40% of maximal activity
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
25 - 37
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assay at
25
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assay at
70 - 80
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CPS.A-CPS.B complex
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
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SV40-transformed
Manually annotated by BRENDA team
growth medium is bovine milk
Manually annotated by BRENDA team
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mammary carcinoma DMBA-1C, DMBA-5A, 205A, R3230AC, RNC-211
Manually annotated by BRENDA team
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about 10fold higher activity than in normal liver
Manually annotated by BRENDA team
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maintained in human foreskin fibroblasts
Manually annotated by BRENDA team
additional information
-
rapidly growing tissues, e.g. fetal liver, spleen, and testis have much more carbamoyl-phosphate synthase activity than other tissues testes, e.g. lung, uterus, heart, brain and muscle. Tumors are more active than most normal tissues
Manually annotated by BRENDA team
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
-
carbamoyl-phosphate-synthase/aspartate-transcarbamoylase complex
Manually annotated by BRENDA team
PDB
SCOP
CATH
ORGANISM
UNIPROT
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)