Information on EC 2.7.2.2 - carbamate kinase

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

EC NUMBER
COMMENTARY hide
2.7.2.2
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RECOMMENDED NAME
GeneOntology No.
carbamate kinase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
ATP + NH3 + CO2 = ADP + carbamoyl phosphate
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
phospho group transfer
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
allantoin degradation IV (anaerobic)
-
-
Arginine biosynthesis
-
-
L-citrulline degradation
-
-
Metabolic pathways
-
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Microbial metabolism in diverse environments
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-
Nitrogen metabolism
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Purine metabolism
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-
urea cycle
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-
SYSTEMATIC NAME
IUBMB Comments
ATP:carbamate phosphotransferase
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CAS REGISTRY NUMBER
COMMENTARY hide
9026-69-1
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
strain ATCC 700683
-
-
Manually annotated by BRENDA team
SD10 strain
-
-
Manually annotated by BRENDA team
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-
-
Manually annotated by BRENDA team
enzyme has inherent activities of agmatine iminohydrolase, putrescine transcarbamylase, ornithine transcarbamylase and carbamate kinase
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-
Manually annotated by BRENDA team
type II strain 07
-
-
Manually annotated by BRENDA team
no activity in Homo sapiens
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
strain ATCC 700122
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-
Manually annotated by BRENDA team
serotype M49, strain 591, gene arcC
UniProt
Manually annotated by BRENDA team
serotype 2
-
-
Manually annotated by BRENDA team
strain RH
-
-
Manually annotated by BRENDA team
strain RH
-
-
Manually annotated by BRENDA team
-
SwissProt
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
metabolism
physiological function
additional information
mdelling of the environment of Cys242 superposed in three conformational states, the AMP-PNP bound structure, the citric acid bound structure, and the thiocarbamoylated structure, overview. Modification of Cys242 interferes with the conformational transition that accompanies nucleotide binding, whereas the citric acid binding remains unaltered and the auxiliary domain is in the closed conformation
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
ADP + carbamoyl phosphate
ATP + NH3 + CO2
show the reaction diagram
ATP + carbamate
ADP + carbamoyl phosphate
show the reaction diagram
-
-
-
?
ATP + NH3 + acetate
ADP + ?
show the reaction diagram
ATP + NH3 + CO2
ADP + carbamoyl phosphate
show the reaction diagram
ATP + NH3 + formate
ADP + ?
show the reaction diagram
poor substrate
-
-
r
ATP + NH3 + propionate
ADP + ?
show the reaction diagram
poor substrate
-
-
r
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
ADP + carbamoyl phosphate
ATP + NH3 + CO2
show the reaction diagram
ATP + NH3 + CO2
ADP + carbamoyl phosphate
show the reaction diagram
additional information
?
-
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
additional information
-
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Bivalent cation
additional information
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
2,3-Butanedione
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in borate buffer, inactivation, implying the presence of an essential arginine
adenosine(5')hexaphospho(5')adenosine
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-
adenosine(5')pentaphospho(5')adenosine
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-
bicarbonate
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slight substrate inhibition
Disulfiram
the anti-alcoholism drug kills the trophozoites and inhibits the enzyme. Disulfiram acts by modifying Cys-242 adjacent to the active site and cures giardiasis in mice. The compound thiocarbamoylates Cys242, a residue located at the edge of the active site. The modified Cys242 prevents a conformational transition of a loop adjacent to the ADP/ATP binding site, which is required for the stacking of Tyr245 side chain against the adenine moiety
iodoacetamide
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weak
L-arginine
slight inhibition
metronidazole
irreversible inactivation mechanism, the thiuram compounds can circumvent the resistance mechanism that renders metronidazole ineffectiveness in drug resistance cases of giardiasis
MgADP-
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competitive to ATP
p-hydroxymercuribenzoate
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-
Phosphonoacetate
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; competitive to carbamoyl phosphate
Silver-Tris
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-
-
additional information
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
1.7
acetate
0.001 - 0.72
ADP
0.017 - 2
ATP
4
bicarbonate
-
-
3.8 - 43
Carbamate
0.085 - 1.63
Carbamoyl phosphate
0.57 - 0.71
MgADP-
0.62
MgATP2-
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pH 7.5, 35°C
0.5
MgdATP2-
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pH 7.5, 35°C
0.68 - 1.17
MnADP-
additional information
additional information
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TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
319
ADP
Giardia intestinalis
A8BB85
pH 7.5, 25°C
43
Carbamate
Giardia intestinalis
A8BB85
pH 7.5, 25°C
319
Carbamoyl phosphate
Giardia intestinalis
A8BB85
pH 7.5, 25°C
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
70000
ADP
Giardia intestinalis
A8BB85
pH 7.5, 25°C
13
0.001
Carbamate
Giardia intestinalis
A8BB85
pH 7.5, 25°C
3992
85000
Carbamoyl phosphate
Giardia intestinalis
A8BB85
pH 7.5, 25°C
440
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
6.2 - 9.4
ATP
4.4
CTP
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plus Mg2+, 30°C, pH 5.0
0.15
Phosphonoacetate
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pH 7.4, 37°C
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
980
-
carbamate kinase is determined in incubations containing 1 mM ADP, 20 mM MgSO4, 0.15 mM luciferin, 1 mg firefly lantern extract, and 1 mM carbamoyl phosphate, in 50 mM potassium phosphate buffer, pH 7.6, ATP formation is determined by monitoring the luminescence using a photomultiplier tube
1100
recombinant protein
additional information
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5.4 - 8
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30°C, pH 5.0
8.1 - 8.3
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-
additional information
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activity in HEPES buffer is twice that in potassium citrate buffer at pH 6.0
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5 - 8
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pH 5.0: about 50% of maximum activity, pH 8.0: about 60% of maximum activity
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4.75
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isoeletric focusing, recombinant protein
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
PDB
SCOP
CATH
ORGANISM
UNIPROT
Aeropyrum pernix (strain ATCC 700893 / DSM 11879 / JCM 9820 / NBRC 100138 / K1)
Enterococcus faecalis (strain ATCC 700802 / V583)
Enterococcus faecalis (strain ATCC 700802 / V583)
Giardia intestinalis (strain ATCC 50803 / WB clone C6)
Giardia intestinalis (strain ATCC 50803 / WB clone C6)
Giardia intestinalis (strain ATCC 50803 / WB clone C6)
Giardia intestinalis (strain ATCC 50803 / WB clone C6)
Giardia intestinalis (strain ATCC 50803 / WB clone C6)
Pyrococcus furiosus (strain ATCC 43587 / DSM 3638 / JCM 8422 / Vc1)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
31000
-
2 * 31000, deduced from amino acid composition
33000
-
x * 33000, recombinant protein, SDS-PAGE, x * 328000, deduced from gene sequence, 14 residue C-terminal sequence is important for enzyme activity
34000
-
x * 34000, SDS-PAGE
36000
x * 36000, SDS-PAGE, recombinant protein including His-tag
37000
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2 * 37000, SDS-PAGE
40000 - 45300
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sedimentation equilibrium method
61000
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sucrose density gradient technique
66000
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sedimentation data
70000
gel filtration, recombinant protein
74000
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sucrose density gradient centrifugation
97000
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gel electrophoresis
328000
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x * 33000, recombinant protein, SDS-PAGE, x * 328000, deduced from gene sequence, 14 residue C-terminal sequence is important for enzyme activity
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
purified detagged recombinant enzyme in complex with nonhydrolyzable ATP analogue, adenosine 5'-adenylyl-beta,gamma-imidodiphosphate (AMP-PNP), and with citric acid, hanging drop vapor diffusion method, mixing of 30 mg/ml protein in 50 mM Tris-HCl, pH 8.0, 0.1 M NaCl, 5 mM MgCl2, and 1 mM DTT, with an equal volume of mother liquor containing 0.4 M ammonium citrate dibasic, pH 5.0, and 21% w/v PEG 3350, soaking of crystals in 50 mM AMP-PNP, X-ray diffraction structure determination and analysis at 2.1-2.6 A resolution
the three-dimensional structure of carbamate kinase from the human parasite Giardia lamblia is determined at 3 A resolution. The crystals belong to the monoclinic space group P21, with unit-cell parameters a = 69.77, b = 85.41, c = 102.1 A, beta = 106.8°
crystals of the pyrococcal enzyme grown in the absence or presence of MgATP (Fig. 7) diffracted with a conventional X-ray source to at least 2.6 and 2.0 Å resolution, respectively
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TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
55
-
unstable above
60
-
2 min, 85% loss of activity
additional information
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2-mercaptoethanol protects against heat denaturation
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
2-mercaptoethanol protects against heat denaturation
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80% loss of activity on dialysis against 0.04 M Tris, pH 8.5, 4°C, 18 h, Streptococcus lactis enzyme
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all the component activities associated with putrescine synthase are stabilized in dilute solutions, 0.05 mg of protein per ml, for about 3-4 h at 37°C by 0.250 mg/ml bovine serum albumin
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ammonium sulfate, 0.5 M, stabilizes against inactivation
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DTT is required to prevent the protein from aggregation
prolonged dialysis and freeze-thawing, loss of activity
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purified enzyme is highly unstable even in presence of glycerol, dithiothreitol and Mg2+
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unstable in dilute solutions, even at very low temperatures
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-20°C, 1 week, 50% loss of activity
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-20°C, 2 weeks, stable
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4°C, 48 h, complete loss of activity
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
enzyme has inherent activities of agmatine iminohydrolase, putrescine transcarbamylase, ornithine transcarbamylase and carbamate kinase
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recombinant enzyme from Escherichia coli strain DH5alpha by affinity chromatography on StrepTactin resin
recombinant protein from Saccharomyces cerevisiae
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
; expression in Escherichia coli
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comparison of carbamate kinases and carbamoyl phosphate synthases
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expressed in Escherichia coli Rosetta(DE3) and Saccharomyces cerevisiae
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gene arcC, genetic organization in the arc operon, sequence comparisons, recombinant expression in Escherichia coli strain DH5alpha
gene cbk, recombinant expression in Escherichia coli strain BL21(DE3)Star as a TEV protease-cleavable, His-tagged, maltose binding protein fusion protein
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
D208A/D210A
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0.1% of wild type activity
E136A/E138A/E141A/K140A
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not detrimental
additional information
-
enzyme can replace in vivo carbamoyl phosphate synthetase of Escherichia coli
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
degradation
drug development
additional information
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