Information on EC 3.5.1.42 - nicotinamide-nucleotide amidase

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

EC NUMBER
COMMENTARY hide
3.5.1.42
-
RECOMMENDED NAME
GeneOntology No.
nicotinamide-nucleotide amidase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
beta-nicotinamide D-ribonucleotide + H2O = beta-nicotinate D-ribonucleotide + NH3
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
hydrolysis of carboxylic acid amide
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-
-
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PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
NAD metabolism
-
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NAD salvage pathway II
-
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Nicotinate and nicotinamide metabolism
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SYSTEMATIC NAME
IUBMB Comments
nicotinamide-D-ribonucleotide amidohydrolase
Also acts more slowly on beta-nicotinamide D-ribonucleoside.
CAS REGISTRY NUMBER
COMMENTARY hide
37355-58-1
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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
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-
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Manually annotated by BRENDA team
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-
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Manually annotated by BRENDA team
DSM 826
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-
Manually annotated by BRENDA team
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-
-
Manually annotated by BRENDA team
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-
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Manually annotated by BRENDA team
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-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
no activity in Oryctolagus cuniculus
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-
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Manually annotated by BRENDA team
no activity in Saccharomyces cerevisiae
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-
-
Manually annotated by BRENDA team
gene pncC
UniProt
Manually annotated by BRENDA team
gene pncC
UniProt
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
gene cinA
UniProt
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
evolution
metabolism
additional information
-
enzyme surface charge analysis and substrate docking performed on the enzyme structure from Agrobacterium tumefaciens, PDB code 2A9S, overview
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
beta-nicotinamide D-ribonucleotide + H2O
beta-nicotinate D-ribonucleotide + NH3
show the reaction diagram
nicotinamide + H2O
nicotinate + NH3
show the reaction diagram
-
-
-
-
?
nicotinamide mononucleotide + H2O
nicotinate mononucleotide + NH3
show the reaction diagram
nicotinamide ribonucleoside + H2O
nicotinic acid ribonucleoside + NH3
show the reaction diagram
nicotinamide riboside + H2O
nicotinate riboside + NH3
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
beta-nicotinamide D-ribonucleotide + H2O
beta-nicotinate D-ribonucleotide + NH3
show the reaction diagram
nicotinamide mononucleotide + H2O
nicotinate mononucleotide + NH3
show the reaction diagram
nicotinamide ribonucleoside + H2O
nicotinic acid ribonucleoside + NH3
show the reaction diagram
additional information
?
-
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the enzyme has no detectable deamidase activity toward NAD, NADP+, nicotinamide, and nicotinamide riboside
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-
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METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Mg2+
-
significant stimulation
additional information
-
the enzyme does not require a divalent cation for catalytic activity
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
alpha-nicotinic acid adenine dinucleotide
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non-competitive inhibitor, Ki: 8 mM
alpha-nicotinic acid mononucleotide
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non-competitive inhibitor, Ki: 21 mM
beta-nicotinic acid adenine dinucleotide
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non-competitive inhibitor, Ki: 7 mM
Cu2+
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strong inhibitor
deamido-NAD+
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product inhibition
Hg2+
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52% inhibition at 1 mM
Isonicotinic acid
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non-competitive inhibitor, Ki: 8.3 mM
nicotinic acid
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non-competitive inhibitor, Ki: 4.4 mM
p-chloromercuribenzoate
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-
p-hydroxymercuribenzoate
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26% inhibition at 1 mM
Picolinic acid
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non-competitive inhibitor, Ki: 9.8 mM
quinolinic acid
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non-competitive inhibitor, Ki: 3.9 mM
additional information
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
ATP
-
weak activator
diphosphate
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weak activator
phosphate
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weak activator
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
3.6
beta-nicotinamide D-ribonucleoside
-
-
0.007 - 1.05
beta-nicotinamide D-ribonucleotide
0.006
nicotinamide mononucleotide
additional information
additional information
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.38 - 4.1
beta-nicotinamide D-ribonucleotide
3.3
nicotinamide mononucleotide
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
2.1 - 590
beta-nicotinamide D-ribonucleotide
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
8
alpha-nicotinic acid adenine dinucleotide
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-
21
alpha-nicotinic acid mononucleotide
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7
beta-nicotinic acid adenine dinucleotide
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8.3
Isonicotinic acid
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4.4
nicotinic acid
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9.8
Picolinic acid
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3.9
quinolinic acid
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SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.003
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crude extract, at pH 8.0 and 37C
0.07
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cell-free extract
1.8
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after 600fold purification, at pH 8.0 and 37C
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5 - 5.5
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maximal activity within
5
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no activity below
5 - 10
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the enzyme shows a broad optimum pH, ranging from 5.5 to 9.0, 72 and 60% residual activity is observed at pH 10.0 and 5.0, respectively
5.5 - 10
the enzyme is active over a broad pH range, with an optimum at pH 7.4, whilst maintaining 90% activity at pH 10.0
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
25700 - 26300
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gel filtration
33000
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gel filtration
80000
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gel filtration
96100
recombinant His6-tagged enzyme, gel filtration
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
homodimer
oligomer
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heat-stable and heat-sensitive subunits
additional information
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enzyme structure comparisons
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
purified recombinant detagged enzyme free or in complex with nicotinate mononucleotide, or AMP/Mg2+, or ATP/Mg2+, or ribose-ADP/Mg2+, sitting drop vapor diffusion, mixing of 200 nl of protein with 200 nl of well solution containing 0.2 M Na2SO4, 0.1 M Bis-Tris propane, pH 6.5, and 20% w/v, PEG 3350 at 20C, rodlike crystals, 5-7 days, X-ray diffraction structure determination and analysis at 1.98-2.46 A resolution, molecular replacement and modeling
pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
3.8
-
21% retaining activity
209139
5 - 6
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stable within for 30 min
209134
7.4
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48% retaining activity
209139
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
50
purified recombinant enzyme, pH 7.3, stable up to
64
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preincubation without substrate: complete denaturation within 32 min
70
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purified proteins, wild-type enzyme loses 40% activity after 20 min, 60% after 40 min, and 80% after 60 min, the mutant E28A lose s 50%, 80% and over 90% activity after 30, 40 qnd 60 min, the mutant T31A is inactivated within 20 min
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
DTT required during purification
-
unstable during purification procedure
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STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-15C, 0.5 mg protein/ml, 12% glycerol, 1 month, more stable at 4C in the presence of 24% glycerol
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-70C, 20 mM Tris-HCl, pH 7.5, 15% glycerol, fully active within 1 month
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4C, Tris-HCl, pH 7.5, 1 week
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
ammonium sulfate precipitation, phenyl-Sepharose column chromatography, Reactive Red 120-agarose column chromatography, Resource Q column chromatography, and Superose 12 gel filtration
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near homogeneity, chromatography techniques
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partial
recombinant C-terminally His6-tagged enzyme from Escherichia coli strain Rosetta2 (DE3) by nickel affinity chromatography and gel filtration
recombinant His6-tagged enzyme from Escherichia coli strain T7 by nickel affinity chromatgraphy, cleavage of the tag by thrombin, anion exchange chromatography, and gel filtration
recombinant wild-type and mutant enzymes from Escherichia coli strain BL21(DE3) by nicke affinity chromatography
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to homogeneity, chromatography techniques
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
gene cinA, recombinant expression of His6-tagged enzyme with a thrombin cleavage site at the N terminus in Escherichia coli strain T7
gene pncC, DNA and amino acid sequence determination and analysis, sequence comparisons, phylogenetic analysis, recombinant expression of C-terminally His6-tagged enzyme in Escherichia coli strain Rosetta2 (DE3)
gene pncC, sequence comparisons, recombinant expression of wild-type and mutant enzymes in Escherichia coli strain BL21(DE3)
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
E28A
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site-directed mutagenesis, the mutant shows reduced activity compared to the wild-type enzyme
K61Q
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site-directed mutagenesis, inactive mutant
R142A
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site-directed mutagenesis, inactive mutant
S103A
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site-directed mutagenesis, the mutant shows reduced activity compared to the wild-type enzyme
S29A
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site-directed mutagenesis, inactive mutant
T31A
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site-directed mutagenesis, the mutant shows reduced activity compared to the wild-type enzyme
Y56A
-
site-directed mutagenesis, inactive mutant
Show AA Sequence (145 entries)
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