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Information on EC 2.7.1.86 - NADH kinase and Organism(s) Saccharomyces cerevisiae and UniProt Accession P32622
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2.7.1.86 NADH kinaseIUBMB Comments
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.
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The taxonomic range for the selected organisms is: Saccharomyces cerevisiae
The expected taxonomic range for this enzyme is: Eukaryota, Bacteria
The expected taxonomic range for this enzyme is: Eukaryota, Bacteria
Synonyms
nadh kinase, pos5p, atp-nadh kinase, atp:nadh 2'-phosphotransferase, nicotinamide adenine dinucleotide hydride kinase, 
more
topSYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
ATP:NADH 2'-phosphotransferase
ATP:NADH2 2'-phosphotransferase
-
-
-
-
DPNH kinase
-
-
-
-
kinase, reduced nicotinamide adenine dinucleotide (phosphorylating)
-
-
-
-
NADH kinase
-
-
-
-
NADH2 kinase
-
-
-
-
Pos5
Pos5p
reduced diphosphopyridine nucleotide kinase
-
-
-
-
reduced nicotinamide adenine dinucleotide kinase (phosphorylating)
-
-
-
-
Utr1
ATP:NADH 2'-phosphotransferase
-
-
-
-
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
phospho group transfer
-
-
-
-
PATHWAY SOURCE
PATHWAYS
-
-, -, -
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
62213-39-2
-
SUBSTRATE
PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
ATP + NAD+
ADP + NADP+
-
2fold preference for NADH over NAD+
-
?
ATP + NAD+
ADP + NADP+
-
activity with NAD+ is 50fold lower than with NADH
-
-
?
ATP + NAD+
ADP + NADP+
the enzyme has much higher NADH kinase than NAD kinase activity
-
-
?
ATP + NADH
ADP + NADPH
-
2fold preference for NADH over NAD+
-
?
ATP + NADH
ADP + NADPH
-
may be involved in maintaining the level of triphosphopyridine nucleotide in the mitochondria
-
?
ATP + NADH
ADP + NADPH
-
the enzyme has primarily NADH kinase activity in mitochondria, mitochondrial NADPH is largely provided by the POS5 NADH kinase
-
?
ATP + NADH
ADP + NADPH
phosphorylates NADH with 2% of the specific activity of its NAD kinase activity
-
-
?
ATP + NADH
ADP + NADPH
possesses 33% activity with NADH and ATP compared to the activity obtained with NAD and ATP
-
-
?
ATP + NADH
ADP + NADPH
Pos5 is responsible for all mitochondrial NAD/NADH kinase. Transitory exposure of Pos5 to the cytoplasm is required for the viability of utr1 mutants
-
-
?
ATP + NADH
ADP + NADPH
isoform Pos5p shows 167% activity compared to NAD+
-
-
?
ATP + NADH
ADP + NADPH
the enzyme has much higher NADH kinase than NAD kinase activity
-
-
?
additional information
?
-
-
POS5 NADH kinase is required for mitochondrial stability with a critical role in detoxification of reactive oxygen species
-
-
?
additional information
?
-
-
NADH kinase does not increase the rate of xylose consumption
-
-
?
additional information
?
-
NADH kinase does not increase the rate of xylose consumption
-
-
?
additional information
?
-
NADH kinase does not increase the rate of xylose consumption
-
-
?
additional information
?
-
does not accept poly(P) as phosphoryl donor
-
-
?
additional information
?
-
enzyme additionally show NAD+ kinase activity, Km value for NAD+ is 0.5 mM
-
-
?
additional information
?
-
-
enzyme additionally show NAD+ kinase activity, Km value for NAD+ is 0.5 mM
-
-
?
NATURAL SUBSTRATE
NATURAL PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
ATP + NADH
ADP + NADPH
-
may be involved in maintaining the level of triphosphopyridine nucleotide in the mitochondria
-
?
ATP + NADH
ADP + NADPH
-
the enzyme has primarily NADH kinase activity in mitochondria, mitochondrial NADPH is largely provided by the POS5 NADH kinase
-
?
ATP + NADH
ADP + NADPH
Pos5 is responsible for all mitochondrial NAD/NADH kinase. Transitory exposure of Pos5 to the cytoplasm is required for the viability of utr1 mutants
-
-
?
additional information
?
-
-
POS5 NADH kinase is required for mitochondrial stability with a critical role in detoxification of reactive oxygen species
-
-
?
additional information
?
-
-
NADH kinase does not increase the rate of xylose consumption
-
-
?
additional information
?
-
NADH kinase does not increase the rate of xylose consumption
-
-
?
additional information
?
-
NADH kinase does not increase the rate of xylose consumption
-
-
?
COFACTOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
Mg2+
-
either Mn2+ or Mg2+ activate at low concentrations. Km: Km: 1.0 mM
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
phosphoenolpyruvate
-
20 mM, 63% inhibition, reconstituted enzyme. Solubilized enzyme is inhibited 55% by 30 mM
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
acetate
-
high concentrations of carboxylic acid, e.g. acetate required for maximal activity
acetate
-
maximal enhancement of activity, about 7fold is observed with 100 mM acetate, enzyme reconstituted in liposomes, maximal enhancement of solubilized enzyme with 400 mM acetate
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.17
ATP
pH and temperature not specified in the publication
2
NADH
pH and temperature not specified in the publication
1.3
NAD+
mutant enzyme R293H, in 100 mM Tris-HCl, pH 8.0, at 37°C
4.5
NAD+
wild type enzyme, in 100 mM Tris-HCl, pH 8.0, at 37°C
0.032
NADH
mutant enzyme R293H, in 100 mM Tris-HCl, pH 8.0, at 37°C
0.19
NADH
wild type enzyme, in 100 mM Tris-HCl, pH 8.0, at 37°C
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
6.2
NAD+
mutant enzyme R293H, in 100 mM Tris-HCl, pH 8.0, at 37°C
7.4
NAD+
wild type enzyme, in 100 mM Tris-HCl, pH 8.0, at 37°C
7.7
NADH
mutant enzyme R293H, in 100 mM Tris-HCl, pH 8.0, at 37°C
16.1
NADH
wild type enzyme, in 100 mM Tris-HCl, pH 8.0, at 37°C
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
1.6
NAD+
wild type enzyme, in 100 mM Tris-HCl, pH 8.0, at 37°C
4.8
NAD+
mutant enzyme R293H, in 100 mM Tris-HCl, pH 8.0, at 37°C
85
NADH
wild type enzyme, in 100 mM Tris-HCl, pH 8.0, at 37°C
241
NADH
mutant enzyme R293H, in 100 mM Tris-HCl, pH 8.0, at 37°C
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
pH RANGE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
6.2 - 9.9
-
pH 6.2: about 35% of maximal activity, pH 9.9: about 50% of maximal activity
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY
GeneOntology No.
LITERATURE
SOURCE
-
the enzyme is localized and functions at the intermembrane space side of the inner membrane
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
malfunction
physiological function
malfunction
-
yeast cells lacking the mitochondrial NADH kinase encoded by POS5 display increased sensitivity to hydrogen peroxide, a slow-growth phenotype, reduced mitochondrial function and increased levels of mitochondrial protein oxidation and mitochondrial DNA mutations
malfunction
-
deletion of the POS5 gene results in respiratory deficiency, anti-oxidative dysfunction and arginine auxotrophy of cells, and the respiratory deficiency is fatal for mitochondrial function
malfunction
-
DELTApos5 mitochondria cannot efficiently synthesize Fe-S clusters on endogenous aconitase or imported ferredoxin
physiological function
-
the enzyme contributes to the maintenance of respiratory chain complex activities
physiological function
-
the enzyme is required for efficient iron-sulfur cluster biogenesis in wild type mitochondria
physiological function
a triple mutant lacking the activities of ATP-NAD/NADH kinase Utr1, ATP-NADH kinase Pos5 and ATP-NADH kinase Yel041wp, i.e. Yef1, is viable and does not show growth defects. Presence of Utr1 protein is critical for growth in low iron medium
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
additional information
-
presence of 2 or more subunits of different size
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 20°C
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
H231D
the mutant exhibits no activity towards NAD+ and low activity for NADH
R293H
the mutation reduces the ratio of NADH kinase activity to NAD kinase activity from 8.6 to 2.1
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-30°C, 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
-
0-5°C, 20 mM Tris-HCl buffer, pH 7.8, 0.2 M ammonium sulfate, 20 mM MgCl2, 2 mM EDTA, stable for 7 days
-
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
Ni-NTA agarose column chromatography and Superdex 200 gel filtration
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
expressed in Escherichia coli
expression in Escherichia coli as N-terminally His-tagged fusion
overpressed in Pichia pastoris, either directed to the cytosol or to the mitochondria. POS5 overexpression increases the NADPH/NADPx02 ratio in all the strains and under all conditions tested
-
pos5 is co-expressed with poly-3-hydroxybutyrate (PHB) synthetic operon phbCAB in Escherichia coli. The recombinant strain carrying pos5 and phbCAB co-expression plasmid reaches 5.96 g/l cell dry weight with 64.1% PHB accumulation in 72 h shake flask cultivation, while the control strain without pos5 yields 3.93 g/l cell dry weight with 58.5% PHB content
EXPRESSION
ORGANISM
UNIPROT
LITERATURE
transcription of POS5 is upregulated 3.4fold after treatment for 10-12 min with 2.5 mM Cu2+
APPLICATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
biotechnology
synthesis
NADH kinase can be employed as an effective metabolic manipulation target to improve poly-3-hydroxybutyrate synthesis
biotechnology
NADH kinase can be employed as an effective metabolic manipulation target to improve poly-3-hydroxybutyrate synthesis
biotechnology
-
recombinant protein overproduction often results in oxidative stress, causing deviations from the optimal redox cofactor regeneration balance. This becomes one of the limiting factors in obtaining high levels of heterologous protein production. Overexpression of NADH kinase enzyme from Saccharomyces cerevisiae (Pos5) in the cytosol of Pichia pastoris results in a significant improvement in the production of the model protein
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Griffiths, M.M.; Bernofsky, C.
Purification and properties of reduced diphosphopyridine nucleotide kinase from yeast mitochondria
J. Biol. Chem.
247
1473-1478
1972
Saccharomyces cerevisiae
Iwahashi, Y.; Nakamura, T.
Localization of the NADH kinase in the inner membrane of yeast mitochondria
J. Biochem.
105
916-921
1989
Saccharomyces cerevisiae, Saccharomyces cerevisiae X2181-1A
Iwahashi, Y.; Nakamura, T.
Orientation and reactivity of NADH kinase in proteoliposomes
J. Biochem.
105
922-926
1989
Saccharomyces cerevisiae, Saccharomyces cerevisiae X2181-1A
Iwahashi, Y.; Hitoshio, A.; Tajima, N.; Nakamura, T.
Characterization of NADH kinase from Saccharomyces cerevisiae
J. Biochem.
105
588-593
1989
Saccharomyces cerevisiae, Saccharomyces cerevisiae X2181-1A
Strand, M.K.; Stuart, G.R.; Longley, M.J.; Graziewicz, M.A.; Dominick, O.C.; Copeland, W.C.
POS5 gene of Saccharomyces cerevisiae encodes a mitochondrial NADH kinase required for stability of mitochondrial DNA
Eukaryot. Cell
2
809-820
2003
Saccharomyces cerevisiae
Outten, C.E.; Culotta, V.C.
A novel NADH kinase is the mitochondrial source of NADPH in Saccharomyces cerevisiae
EMBO J.
22
2015-2024
2003
Saccharomyces cerevisiae
Shi, F.; Kawai, S.; Mori, S.; Kono, E.; Murata, K.
Identification of ATP-NADH kinase isozymes and their contribution to supply of NADP(H) in Saccharomyces cerevisiae
FEBS J.
272
3337-3349
2005
Saccharomyces cerevisiae (P21373), Saccharomyces cerevisiae
Bieganowski, P.; Seidle, H.F.; Wojcik, M.; Brenner, C.
Synthetic lethal and biochemical analyses of NAD and NADH kinases in Saccharomyces cerevisiae establish separation of cellular functions
J. Biol. Chem.
281
22439-22445
2006
Saccharomyces cerevisiae, Saccharomyces cerevisiae (Q06892)
Kawai, S.; Murata, K.
Structure and function of NAD kinase and NADP phosphatase: key enzymes that regulate the intracellular balance of NAD(H) and NADP(H)
Biosci. Biotechnol. Biochem.
72
919-930
2008
Arabidopsis thaliana, Saccharomyces cerevisiae (C7GJD6)
Hou, J.; Vemuri, G.N.; Bao, X.; Olsson, L.
Impact of overexpressing NADH kinase on glucose and xylose metabolism in recombinant xylose-utilizing Saccharomyces cerevisiae
Appl. Microbiol. Biotechnol.
82
909-919
2009
Saccharomyces cerevisiae, Saccharomyces cerevisiae (P32622), Saccharomyces cerevisiae (Q06892)
Stuart, G.R.; Humble, M.M.; Strand, M.K.; Copeland, W.C.
Transcriptional response to mitochondrial NADH kinase deficiency in Saccharomyces cerevisiae
Mitochondrion
9
211-221
2009
Saccharomyces cerevisiae, Saccharomyces cerevisiae YPH925
Shi, F.; Li, Z.; Sun, M.; Li, Y.
Role of mitochondrial NADH kinase and NADPH supply in the respiratory chain activity of Saccharomyces cerevisiae
Acta Biochim. Biophys. Sin.
43
989-995
2011
Saccharomyces cerevisiae, Saccharomyces cerevisiae BY4742
Lee, W.H.; Kim, J.W.; Park, E.H.; Han, N.S.; Kim, M.D.; Seo, J.H.
Effects of NADH kinase on NADPH-dependent biotransformation processes in Escherichia coli
Appl. Microbiol. Biotechnol.
97
1561-1569
2013
Saccharomyces cerevisiae, Saccharomyces cerevisiae CEN.PK2-1D
Lee, W.H.; Kim, M.D.; Jin, Y.S.; Seo, J.H.
Engineering of NADPH regenerators in Escherichia coli for enhanced biotransformation
Appl. Microbiol. Biotechnol.
97
2761-2772
2013
Saccharomyces cerevisiae
Pain, J.; Balamurali, M.M.; Dancis, A.; Pain, D.
Mitochondrial NADH kinase, Pos5p, is required for efficient iron-sulfur cluster biogenesis in Saccharomyces cerevisiae
J. Biol. Chem.
285
39409-39424
2010
Saccharomyces cerevisiae, Saccharomyces cerevisiae BY4741
Ando, T.; Ohashi, K.; Ochiai, A.; Mikami, B.; Kawai, S.; Murata, K.
Structural determinants of discrimination of NAD+ from NADH in yeast mitochondrial NADH kinase Pos5
J. Biol. Chem.
286
29984-29992
2011
Saccharomyces cerevisiae (Q06892), Saccharomyces cerevisiae
Tomas-Gamisans, M.; Andrade, C.C.P.; Maresca, F.; Monforte, S.; Ferrer, P.; Albiol, J.
Redox engineering by ectopic overexpression of NADH kinase in recombinant Pichia pastoris (Komagataella phaffii) impact on cell physiology and recombinant production of secreted proteins
Appl. Environ. Microbiol.
86
e02038-19
2020
Saccharomyces cerevisiae
Li, B.; Wang, X.; Tai, L.; Ma, T.; Shalmani, A.; Liu, W.; Li, W.; Chen, K.
NAD kinases Metabolic targets controlling redox co-enzymes and reducing power partitioning in plant stress and development
Front. Plant Sci.
9
379
2018
Saccharomyces cerevisiae (P32622), Saccharomyces cerevisiae (Q06892), Saccharomyces cerevisiae ATCC 204508 (P32622), Saccharomyces cerevisiae ATCC 204508 (Q06892)
Hong, P.H.; Zhang, J.; Liu, X.J.; Tan, T.W.; Li, Z.J.
Effect of NADH kinase on poly-3-hydroxybutyrate production by recombinant Escherichia coli
J. Biosci. Bioeng.
122
685-688
2016
Saccharomyces cerevisiae (Q06892), Saccharomyces cerevisiae, Saccharomyces cerevisiae ATCC 204508 (Q06892)
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