Information on EC 3.5.2.10 - creatininase

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

EC NUMBER
COMMENTARY hide
3.5.2.10
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RECOMMENDED NAME
GeneOntology No.
creatininase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
creatinine + H2O = creatine
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
hydrolysis of peptide bond
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-
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PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
Arginine and proline metabolism
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creatinine degradation I
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creatinine degradation
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SYSTEMATIC NAME
IUBMB Comments
creatinine amidohydrolase
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CAS REGISTRY NUMBER
COMMENTARY hide
9025-13-2
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
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-
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Manually annotated by BRENDA team
Sigma, Cat. No.C-7399, lot 20 Ho349
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Manually annotated by BRENDA team
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-
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Manually annotated by BRENDA team
strain PS-7
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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
creatinine + H2O
?
show the reaction diagram
creatinine + H2O
creatine
show the reaction diagram
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
creatinine + H2O
?
show the reaction diagram
creatinine + H2O
creatine
show the reaction diagram
-
-
-
-
?
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Co2+
-
activates
Fe2+
-
activates
Mg2+
-
activates
Ni2+
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activates
Zinc
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removal of the zinc ions by dialyzing against buffers containing o-phenanthroline results in a partial decrease of 10-30% in activity and zinc content. Wild-type enzyme, purified in the presence of EDTA, shows only half the activity of the wild-type enzyme, the activity is recovered by the addition of ZnCl2 or MnCl2
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
azide
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complete inhibition at the ionic strength of 2 mM
cyanide
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complete inhibition by 1 mM, much less inhibition by both azide and cyanide
Ethoxyformic anhydride
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formaldehyde
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Heavy metal ions
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-
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Hg2+
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complete inhibition at the ionic strength of 0.1 mM HgCl, 20-30% activity restored at 20 mM 2-mercaptoethanol
methanol
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N-bromosuccinimide
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o-phenanthroline
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Photooxidation
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additional information
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ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.17 - 350
creatinine
additional information
additional information
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steady-state Michaelis-Menten kinetics
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TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
1.5 - 1340
creatinine
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.008 - 17.4
creatinine
2015
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.13
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mutant E122Q, presence of Zn2+, pH not specified in the publication, temperature not specified in the publication
4.9
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mutant W154F, presence of Mn2+, pH not specified in the publication, temperature not specified in the publication
13.9
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mutant E122Q, presence of Mn2+, pH not specified in the publication, temperature not specified in the publication
140
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mutant Y121A, presence of Mn2+, pH not specified in the publication, temperature not specified in the publication
178
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wild-type, presence of EDTA, semi-apo-enzyme, pH not specified in the publication, temperature not specified in the publication
370
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wild-type, presence of Zn2+, pH not specified in the publication, temperature not specified in the publication
740
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mutant W174F, presence of Mn2+, pH not specified in the publication, temperature not specified in the publication
1560
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wild-type, presence of Mn2+, pH not specified in the publication, temperature not specified in the publication
additional information
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by Escherichia coli transformants pCRN741 without and with IPTG 0.678 U per ml
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7.4
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assay at
7.7
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immobilized coupled enzyme system
8
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immobilized
8.5
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immobilized coupled enzyme system
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7.5 - 8.5
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immobilized
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
20
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assay at
25
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immobilized coupled enzyme system
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
23000
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8 * 23000, SDS-PAGE
28399
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6 * 28399, calculated from amino acid sequence
28400
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6 * 28400, MALDI-TOF mass spectrometry
28440
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recombinant polypeptide deduced from the nucleotide sequence
160000
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170400
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SDS-PAGE
173000
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dynamic light scattering
175000
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ultracentrifugal analysis
224000
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gel filtration
240000
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gel filtration
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
dimer
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dimer with a molecular weight of 175000
hexamer
homohexamer
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6 * 28399, calculated from amino acid sequence; 6 * 28400, MALDI-TOF mass spectrometry
octamer
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
glycoprotein
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3.5% glucose
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
; crystal structures of the native and the Mn2+-activated enzyme determined by a difference Fourier method, crystal structure of Mn2+-activated enzyme determined by the single isomorphous replacement method
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crystal structure of the enzyme complexed with a 1-methylguanidine. All subunits in the complex exist as the closed form. X-ray crystallographic study and the atomic absorption spectrometry analysis of the E122Q mutant-substrate complex reveal that the drastic decrease of the activity of the E122Q is caused by the loss of one Zn ion at the Metal1 site and a critical function of Glu122
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hanging drop vapour diffusion method with 1.5 M lithium sulfate in 0.1 M Na-HEPES pH 7.5
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sitting drop vapour diffusion method with 22% ethanol, 20% PEG 8000, and 0.1 M MOPS pH 7.5
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pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5 - 10
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172080
6 - 12
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for 3 h at 30°C
172079
6 - 9
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the enzyme tends to precipitate at pH values of under 6.0 and over 9.0 corresponding to the stability range of the enzyme
677306
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
55
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30 min at pH 7.1, apoenzyme
70
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30 min at pH 7.1, native enzyme
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
enzyme immobilized have a long-term stability, unused 6 month
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OXIDATION STABILITY
ORGANISM
UNIPROT
LITERATURE
unstable to photooxidation
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172079
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-20°C, 6 months
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4°C, 0.1 M phosphate buffer, pH 7.7, enzymes controlled-pore glass immobilized
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4°C, 5 days
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lyophilized
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
ammonium sulfate fractionation, DEAE Sepharose CL-6B and Sepharose CL-6B column chromatographies, 50% total activity
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heating, precipitation of nucleic acids with streptomycin, ammonium sulfate precipitation, Sephadex gel filtration, and DEAE-cellulose column chromatography
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native and Mn2+-activated enzymes purified, Toyopearl HW65C column and DEAE-Toyopearl column
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purified by column chromarographies on sarcosine-HM-Sepharose and DEAE-cellulose, and gel filtration on Sephadex G-200
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
cloned, sequenced and expressed in Escherichia coli DH5?, vector pCRN741, creatininase gene consists of 771 bp, encodes 257 amino acids, similarity of RS65 to PS-7 72.8% and Arthrobacter sp. TE1826 43.2%
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expressed in Escherichia coli
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expression in Escherichia coli
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in Escherichia coli C600 harboring pCNH5-13, Insert contains 780 bp, 259 amino acids
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
E122Q
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almost complete loss of activity
E183Q
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complete loss of activity
H178A
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complete loss of activity
W154A
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complete loss of activity
W154F
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almost complete loss of activity
W174A
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almost complete loss of activity
W174F
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about 50% of wild-type activity
Y121A
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about 10% of wild-type catalytic efficiency
additional information
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construction of an electrochemical biosensor for creatinine based on the immobilization of creatininase, creatinase and sarcosine oxidase onto a ferrocene/horseradish peroxidase/gold nanoparticles/multi-walled carbon nanotubes/Teflon composite electrode,method development and evaluation, overview
Renatured/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
wild-type enzyme, purified in the presence of EDTA, shows only half the activity of the wild-type enzyme, the activity is recovered by the addition of ZnCl2 or MnCl2
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APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
analysis
diagnostics
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the immobilized enzyme can be used in a biosensor for determining creatinine in human serum
medicine
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