Information on EC 2.7.1.86 - NADH kinase

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

EC NUMBER
COMMENTARY hide
2.7.1.86
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RECOMMENDED NAME
GeneOntology No.
NADH kinase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
ATP + NADH = ADP + NADPH
show the reaction diagram
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-
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REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
phospho group transfer
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-
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PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
NAD/NADH phosphorylation and dephosphorylation
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NAD/NADP-NADH/NADPH cytosolic interconversion (yeast)
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NAD/NADP-NADH/NADPH mitochondrial interconversion (yeast)
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SYSTEMATIC NAME
IUBMB Comments
ATP:NADH 2'-phosphotransferase
CTP, ITP, UTP and GTP can also act as phosphate donors (in decreasing order of activity). The enzyme is specific for NADH. Activated by acetate.
CAS REGISTRY NUMBER
COMMENTARY hide
62213-39-2
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
strain CBS 513.88 /FGSC A1513
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-
Manually annotated by BRENDA team
strain CBS 513.88 /FGSC A1513
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-
Manually annotated by BRENDA team
strain F3
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-
Manually annotated by BRENDA team
strain F3
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-
Manually annotated by BRENDA team
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-
-
Manually annotated by BRENDA team
strain H37Rv
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-
Manually annotated by BRENDA team
strain H37Rv
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-
Manually annotated by BRENDA team
strain YPH925
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-
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
malfunction
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
ATP + NAD+
ADP + NADH
show the reaction diagram
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-
-
?
ATP + NAD+
ADP + NADP+
show the reaction diagram
ATP + NADH
ADP + NADPH
show the reaction diagram
CTP + NADH
CDP + NADPH
show the reaction diagram
GTP + NADH
GDP + NADPH
show the reaction diagram
ITP + NADH
IDP + NADPH
show the reaction diagram
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61% of activity with ATP
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-
?
tetrapolyphosphate + NADH
tripolyphosphate + NADPH
show the reaction diagram
UTP + NADH
UDP + NADPH
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
ATP + NAD+
ADP + NADP+
show the reaction diagram
ATP + NADH
ADP + NADPH
show the reaction diagram
additional information
?
-
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Co2+
-
32% activity in the presence of 1mM Co2+
K+
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K+ is the only monovalent metal ion resulting higher NADH kinase activity by almost 1.5fold
Zn2+
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4% of the activity with Mg2+; far less active than Mg2+ and Mn2+
additional information
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
3-acetyl-NAD+
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1 mM, 15% inhibition
Ag2+
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1 mM Ag+ lowers the activity to 3%
beta-mercaptoethanol
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slightly inhibits NADH kinase
Ca2+
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no activity in the presence of 10 mM
Cu2+
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1 mM Cu2+ lowers the activity to 33%
diphosphate
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mixed inhibitor both to NADH and ATP
eosin-5-maleimide
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Fe2+
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no activity in the presence of 10 mM
HgCl2
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13% residual activity at 0.1 mM, 6% residual activity at 0.25 mM
iodoacetic acid
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5 mM, 87% inhibition, NADH protects
Mg2+
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at 10 mM or above
Mn2+
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at 1 mM or above
NADH
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slight inhibition
NADP+
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slightly inhibits NADH kinase
NADPH
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slight inhibition
NEM
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5 mM, 77% inhibition, NADH protects, ATP not
p-chloromercuribenzoate
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0.01 mM, 42% residual activity
phosphoenolpyruvate
Zn2+
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1 mM Zn2+ lowers the activity to 35%
additional information
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NAD+ causes no inhibition of the enzyme
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ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
acetate
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.062 - 2.59
ATP
0.759
CTP
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pH 7.9, 25C, cosubstrate NADH
0.461
GTP
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pH 7.9, 25C, cosubstrate NADH
1.3 - 4.5
NAD+
0.027 - 2
NADH
0.58 - 0.68
Tetrapolyphosphate
0.156
UTP
-
pH 7.9, 25C, cosubstrate NADH
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.84
ATP
Fusarium oxysporum
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6.2 - 7.4
NAD+
0.95 - 16.1
NADH
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
1.6 - 4.8
NAD+
7
85 - 241
NADH
8
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.078
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cell extract, at 30C
0.1
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with GTP as phosphate donor
0.22
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in the presence of 5 mM tetrapolyphosphate
0.5
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with CTP as phosphate donor
0.65
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in the presence of 5 mM ATP
1.2
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with ATP as phosphate donor
3.04
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in the presence of 5 mM tetrapolyphosphate
3.4
in the presence of 0.5 mM EGTA, using NAD+ as a substrate
5.75
in the presence of 0.5 mM EGTA, using NADH as a substrate
8.66
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in the presence of 5 mM ATP
9.475
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after 122fold purification, at 30C
15.1
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NAD+ as substrate, pH 7.9, 25C
15.6
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pH 7.9, 25C, substrate NAD+
40.1
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NADH as susbtrate, pH 7.9, 25C; pH 7.9, 25C, substrate NADH
additional information
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
9.5
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optimum pH for NADH kinase activity of Pos5
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6.2 - 9.9
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pH 6.2: about 35% of maximal activity, pH 9.9: about 50% of maximal activity
6.5 - 8.5
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less than 30% of the peak activity is displayed in pH 6.5 and 8.5
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5.46
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calculated from amino acid sequence
5.6
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isoelectric focusing
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
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restricted to lateral root primordium
Manually annotated by BRENDA team
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restricted primarily to leaf vasculature
Manually annotated by BRENDA team
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
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NADK3 localizes to the peroxisomal matrix via a novel type 1 peroxisomal targeting signal
Manually annotated by BRENDA team
additional information
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not: mitochondrial
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Manually annotated by BRENDA team
PDB
SCOP
CATH
ORGANISM
UNIPROT
Saccharomyces cerevisiae (strain ATCC 204508 / S288c)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
49000
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predicted
55900
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calculated from amino acid sequence
58250
calculated from amino acid sequence
78000
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gel filtration
156000
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gel filtration
160000
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gel filtration
480000
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gel filtration
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
dimer
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2 * 35000, SDS-PAGE
homodimer
homooctamer
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8 * 60000, gel filtration
additional information
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
Pos5 complexed with NADH, sitting drop vapor diffusion method, using 15% (v/v) 2-methyl 2,4-pentanediol, 5% (w/v) polyethylene glycol 4000 and 100 mM imidazole-HCl, pH 8.0, at 20C
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pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5.5
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the enzyme is very unstable at pH values higher than 5.5
706568
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
30
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10 min, stable
40
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10 min, 40% loss of activity
50
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the enzyme retains 8% of the activity after 1 h at 50C
54
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half of its activity is lost on treatment at 54C for 10 min
55
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5 min, 95% loss of activity
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
ammonium sulfate, 0.2 M protects against denaturation
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STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-30C, 20 mM Tris-HCl buffer, pH 7.8, 0.2 M ammonium sulfate, 20 mM MgCl2, 2 mM EDTA, stable for 1 month or more
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0-5C, 20 mM Tris-HCl buffer, pH 7.8, 0.2 M ammonium sulfate, 20 mM MgCl2, 2 mM EDTA, stable for 7 days
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4C or -20C, 95% loss of activity after overnight storage
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
; cobalt chelate affinity column chromatography
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Butyl-Toyopearl 650M column chromatography
DEAE-Toyopearl column chromatography, Butyl-Toyopearl column chromatography, Ni-chelate AF Toyopearl column chromatography
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HiTrap Chelating HP column chromatography
Ni-NTA agarose column chromatography and Superdex 200 gel filtration
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Ni-NTA column chromatography
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Q-Sepharose column chromatography, Sephacryl S300 gel filtration, SP-Sepharose column chromatography, and t-butyl hydrophobic interaction column chromatography
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
a NADK3-GFP reporter fusion construct is expressed in Arabidopsis suspension-cultured cells
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expressed in Escherichia coli
expressed in Escherichia coli BL21(DE3)pLysS cells
expressed in Escherichia coli BL21DE3 cells
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expressed in Escherichia coli BL21star(DE3) cells
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expressed in Escherichia coli RosettaBlue(DE3) cells
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expressed in Escherichia coli strain MK746 as a fusion recombinant protein with a V5 epitope and His6 tag at the C terminus
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expressed in Escherichia coli; expression in Escherichia coli as N-terminally His-tagged fusion
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expression in Escherichia coli
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overexpressed in Aspergillus AR1 derived from the natural isolate A4, pTr04 expression vector
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overexpression in Escherichia coli
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overproduction of the POS5 gene product in Escherichia coli
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EXPRESSION
ORGANISM
UNIPROT
LITERATURE
NADK3 activity is first observed in stage 1.0-1.02, most notably in the procambium and vasculature of cotyledons
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transcription of isoform NADK1 is upregulated by 3 and 8fold after treatment of plant cells with 5 mM H2O2 and irradiation, respectively
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transcription of POS5 is upregulated 3.4fold after treatment for 10-12 min with 2.5 mM Cu2+
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
G183R
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mutant with partially retained NADH kinase activity
H231D
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the mutant exhibits no activity towards NAD+ and low activity for NADH
R293H
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the mutation reduces the ratio of NADH kinase activity to NAD kinase activity from 8.6 to 2.1
R293H/H231D
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inactive
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
analysis
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the enzyme is useful for amplification of NADH in presence of excess NAD+ and is applicable to sensitive measurement of NAD+ dependent dehydrogenase or ist substrate