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Information on EC 1.6.5.9 - NADH:quinone reductase (non-electrogenic) and Organism(s) Saccharomyces cerevisiae and UniProt Accession P32340
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1.6.5.9 NADH:quinone reductase (non-electrogenic)IUBMB Comments
A flavoprotein (FAD or FMN). Occurs in mitochondria of yeast and plants, and in aerobic bacteria. Has low activity with NADPH. Unlike EC 7.1.1.2, NADH:ubiquinone reductase (H+-translocating), this enzyme does not pump proteons of sodium ions across the membrane. It is also not sensitive to rotenone.
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Word Map
- 1.6.5.9
- ndh-2
- hydrogenases
-
proton-translocating
- ubiquinone-1
- hqno
- 2-heptyl-4-hydroxyquinoline-n-oxide
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single-subunit
- deamino-nadh
-
na+-translocating
-
rotenone-sensitive
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:quinone oxidoreductase, alternative nadh dehydrogenase, rotenone-insensitive nadh dehydrogenase, nadh dehydrogenase-2, ndh2e, nadh:q6 oxidoreductase, internal alternative nadh dehydrogenase, ndh2i, alternative nadh-quinone oxidoreductase, non-proton-pumping nadh dehydrogenase, 
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topSYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
Ndi1
NADH:ubiquinone reductase (non-electrogenic)
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-
-
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PATHWAY SOURCE
PATHWAYS
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-, -, -, -, -, -
SYSTEMATIC NAME
IUBMB Comments
NADH:quinone oxidoreductase
A flavoprotein (FAD or FMN). Occurs in mitochondria of yeast and plants, and in aerobic bacteria. Has low activity with NADPH. Unlike EC 7.1.1.2, NADH:ubiquinone reductase (H+-translocating), this enzyme does not pump proteons of sodium ions across the membrane. It is also not sensitive to rotenone.
CAS REGISTRY NUMBER
COMMENTARY
9028-04-0
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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
NADH + H+ + 2-azido-5-geranyl-3-methoxy-6-methyl-1,4-benzoquinone
NAD+ + 2-azido-5-geranyl-3-methoxy-6-methyl-1,4-benzoquinol
-
-
-
?
NADH + H+ + N-(6-[[(4E,8E)-10-(4-azido-5-methoxy-2-methyl-3,6-dioxocyclohexa-1,4-dien-1-yl)-4,8-dimethyldeca-4,8-dien-1-yl]amino]-6-oxohexyl)-6-([5-[(3aS,4S,6aR)-2-oxohexahydro-1H-thieno[3,4-d]imidazol-4-yl]pentanoyl]amino)hexanamide
NAD+ + N-(6-[[(4E,8E)-10-(4-azido-3,6-dihydroxy-5-methoxy-2-methylcyclohexa-1,4-dien-1-yl)-4,8-dimethyldeca-4,8-dien-1-yl]amino]-6-oxohexyl)-6-([5-[(3aS,4S,6aR)-2-oxohexahydro-1H-thieno[3,4-d]imidazol-4-yl]pentanoyl]amino)hexanamide
-
-
-
?
NADH + H+ + N-[(4E,8E)-10-(4-azido-5-methoxy-2-methyl-3,6-dioxocyclohexa-1,4-dien-1-yl)-4,8-dimethyldeca-4,8-dien-1-yl]-6-([5-[(3aS,6aR)-2-oxohexahydro-1H-thieno[3,4-d]imidazol-4-yl]pentanoyl]amino)hexanamide
NAD+ + N-[(4E,8E)-10-(4-azido-3,6-dihydroxy-5-methoxy-2-methylcyclohexa-1,4-dien-1-yl)-4,8-dimethyldeca-4,8-dien-1-yl]-6-([5-[(3aS,4S,6aR)-2-oxohexahydro-1H-thieno[3,4-d]imidazol-4-yl]pentanoyl]amino)hexanamide
-
-
-
?
NADH + H+ + N-[2-[2-(2-[[(4E,8E)-10-(4-azido-5-methoxy-2-methyl-3,6-dioxocyclohexa-1,4-dien-1-yl)-4,8-dimethyldeca-4,8-dien-1-yl]oxy]ethoxy)ethoxy]ethyl]-5-[(3aS,4S,6aR)-2-oxohexahydro-1H-thieno[3,4-d]imidazol-4-yl]pentanamide
NAD+ + N-[2-[2-(2-[[(4E,8E)-10-(4-azido-3,6-dihydroxy-5-methoxy-2-methylcyclohexa-1,4-dien-1-yl)-4,8-dimethyldeca-4,8-dien-1-yl]oxy]ethoxy)ethoxy]ethyl]-5-[(3aS,4S,6aR)-2-oxohexahydro-1H-thieno[3,4-d]imidazol-4-yl]pentanamide
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-
-
?
NADH + H+ + 2,6-dichloroindophenol
NAD+ + reduced 2,6-dichloroindophenol
-
-
-
-
?
NADH + H+ + 2,6-dichlorophenolindophenol
NAD+ + reduced 2,6-dichlorophenolindophenol
-
-
-
-
?
NADH + H+ + ubiquinone-2
NAD+ + ubiquinol-2
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ubiquinone is the best electron acceptor
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-
?
additional information
?
-
-
the yeast enzyme is a two-electron reaction enzyme
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-
?
additional information
?
-
-
NAD+ binds in the second Rossmann domain in a predominantly positively charged cleft, with the nicotinamide ring approaching the re-face of the FAD
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-
?
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
COFACTOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
FAD
-
binding site structure, overview. The isoalloxazine ring of the FAD is positioned between the two Rossmann domains, which define two channels along the interdomain plane that lead to the active site
NADH
-
the enzyme is specific for NADH. NADPH or deamino-NADH show rates that are at least 250times lower than obtained with NADH at any pH between 5.5 and 8.5
additional information
Ndi1 contains no iron-sulfur cluster
-
additional information
-
the enzyme contains no iron-sulfur clusters
-
additional information
-
the mature enzyme contains no iron-sulfur clusters
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INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
AMP
-
dead end reversible inhibitor, competitive inhibitor of NADH and uncompetitive inhibitor of 2,6-dichlorophenolindophenol
additional information
-
NAD+, Mg-ADP, Mg-ATP, Ca2+ (all at 2 mM) or piericidin, rotenone (both at 0.01 mM) or adenosyl-3'-O-[3-[N-(4-azido-2-nitrophenyl)-amino]propionyl]-NAD+ (0.05 mM) are without inhibitory or stimulatory effects
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additional information
-
not inhibited by rotenone, NAD+ does not inhibit the activity of the enzyme, even at concentrations as high as 10 mM
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.0181
N-(6-[[(4E,8E)-10-(4-azido-5-methoxy-2-methyl-3,6-dioxocyclohexa-1,4-dien-1-yl)-4,8-dimethyldeca-4,8-dien-1-yl]amino]-6-oxohexyl)-6-([5-[(3aS,4S,6aR)-2-oxohexahydro-1H-thieno[3,4-d]imidazol-4-yl]pentanoyl]amino)hexanamide
in 50 mM sodium phosphate buffer (pH 6.0) and 1 mM EDTA, temperature not specified in the publication
0.022
N-[(4E,8E)-10-(4-azido-5-methoxy-2-methyl-3,6-dioxocyclohexa-1,4-dien-1-yl)-4,8-dimethyldeca-4,8-dien-1-yl]-6-([5-[(3aS,6aR)-2-oxohexahydro-1H-thieno[3,4-d]imidazol-4-yl]pentanoyl]amino)hexanamide
in 50 mM sodium phosphate buffer (pH 6.0) and 1 mM EDTA, temperature not specified in the publication
0.033
N-[2-[2-(2-[[(4E,8E)-10-(4-azido-5-methoxy-2-methyl-3,6-dioxocyclohexa-1,4-dien-1-yl)-4,8-dimethyldeca-4,8-dien-1-yl]oxy]ethoxy)ethoxy]ethyl]-5-[(3aS,4S,6aR)-2-oxohexahydro-1H-thieno[3,4-d]imidazol-4-yl]pentanamide
in 50 mM sodium phosphate buffer (pH 6.0) and 1 mM EDTA, temperature not specified in the publication
0.0152
ubiquinone-1
in 50 mM sodium phosphate buffer (pH 6.0) and 1 mM EDTA, temperature not specified in the publication
0.0079
ubiquinone-2
in 50 mM sodium phosphate buffer (pH 6.0) and 1 mM EDTA, temperature not specified in the publication
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.031
NADH
-
using ubiquinone-2 as substrate, in 20 mM MOPS/KOH, 1 mM EDTA, 0.03% (v/v) Triton X-100, 200 mM KCl, pH 6.2, temperature not specified in the publication
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
11.5
AMP
-
linear uncompetitive inhibition of 2,6-dichlorophenolindophenol, pH 7.0, 25°C
0.0054
flavone
-
hyperbolic uncompetitive inhibition of NADH, pH 7.0, 25°C
0.0071
flavone
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hyperbolic noncompetitive inhibition of 2,6-dichlorophenolindophenol, pH 7.0, 25°C
IC50 VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.095
flavone
Saccharomyces cerevisiae
-
using ubiquinone-2 as substrate, in 20 mM MOPS/KOH, 1 mM EDTA, 0.03% (v/v) Triton X-100, 200 mM KCl, pH 6.2, temperature not specified in the publication
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
0.78
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crude extract, using ubiquinone-2 as substrate, at pH 6.2, temperature not specified in the publication
1671
-
after 2134fold purification, using ubiquinone-2 as substrate, at pH 6.2, temperature not specified in the publication
61.9
-
after 2134fold purification, using ubiquinone-6 as substrate, at pH 6.2, temperature not specified in the publication
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
6.2
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with ubiquinone-2 as acceptor a sharp maximum at pH 6.2 is obtained
pH RANGE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
4.5 - 9.5
-
with ubiquinone-6 as acceptor the activity is almost constant between pH 4.5 and 9.5, at pH values lower than 4.5 and higher than 9.5, enzymic activity is irreversibly destroyed
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY
GeneOntology No.
LITERATURE
SOURCE
internal surface of the inner mitochondrial membrane
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Ndi1 is a peripheral membrane protein forming an intimate dimer, in which packing of the monomeric units within the dimer creates an amphiphilic membrane-anchor domain structure, membrane-anchor domain, overview
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Ndi1 protein from Saccharomyces cerevisiae is a monotopic membrane protein, directed to the matrix
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
physiological function
inner mitochondrial isoform Ndi1 associates with complexes III and IV. Ndi1 is found in high molecular weight supercomplexes. Complexes containing the NADH dehydrogenase activity are also found in mutants lacking the external NADH dehydrogenases NDE1 and NDE2
additional information
additional information
-
Ndi1 protein from Saccharomyces cerevisiae is a monotopic membrane protein, directed to the mitochondrial matrix. It is a peripheral membrane protein forming an intimate dimer, in which packing of the monomeric units within the dimer creates an amphiphilic membrane-anchor domain structure. Structures of the Ndi1NAD+ and Ndi1UQ2 complexes show overlapping binding sites for the NAD+ and quinone substrates
additional information
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targeted expression of Ndi1 in fly neurons significantly increases lifespan without compromising fertility or physical activity. Expression of the yeast Ndi1 gene in Drosophila mitochondria leads to an increase in respiratory chain activity, overview
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
53000
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
Ndi1 in its substrate-free, NAD+- and ubiquinone-complexed states, X-ray diffraction structure determination and analysis
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pH STABILITY
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
5.5 - 8
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the free enzyme and the binary complex E-NADH are highly stable in the pH range from 5.5 to 8.0, maintaining the initial activity after 10 min incubation. Maximal stability is observed at pH 5.0 and 6.5 with no loss in activity. The enzyme is highly unstable above and below this pH range
392700
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
the enzyme is stable at low protein concentrations and in the presence of both substrates
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PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
DEAE Bio-gel column chromatography and Blue Sepharose CL-6B column chromatography, column chromatography
-
DEAE-cellulose column chromatography and blue Sepharose CL-6B column chromatography
-
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
gene Ndi1, inducible NDI1 enzyme expression in Drosophila melanogaster mitochondria of different cells and tissues. Expression of Ndi1 in fly mitochondria leads to an increase in NADH-ubiquinone oxidoreductase activity, oxygen consumption and ATP levels. In addition, exogenous Ndi1 expression results in increased CO2 production in living flies. Targeted expression of Ndi1 in fly neurons significantly increases lifespan without compromising fertility or physical activity
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EXPRESSION
ORGANISM
UNIPROT
LITERATURE
the amount of enzyme in the cell is subject to glucose repression, it increases slightly when cells, grown on glucose or lactate, enter the stationary phase
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REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
De Vries, S.; Grivell, L.A.
Purification and characterization of a rotenone-insensitive NADH:Q6 oxidoreductase from mitochondria of Saccharomyces cerevisiae
Eur. J. Biochem.
176
377-384
1988
Saccharomyces cerevisiae
Velazquez, I.; Pardo, J.P.
Kinetic characterization of the rotenone-insensitive internal NADH:ubiquinone oxidoreductase of mitochondria from Saccharomyces cerevisiae
Arch. Biochem. Biophys.
389
7-14
2001
Saccharomyces cerevisiae
Murai, M.; Yamashita, T.; Senoh, M.; Mashimo, Y.; Kataoka, M.; Kosaka, H.; Matsuno-Yagi, A.; Yagi, T.; Miyoshi, H.
Characterization of the ubiquinone binding site in the alternative NADH-quinone oxidoreductase of Saccharomyces cerevisiae by photoaffinity labeling
Biochemistry
49
2973-2980
2010
Saccharomyces cerevisiae (P32340), Saccharomyces cerevisiae
Bahadorani, S.; Cho, J.; Lo, T.; Contreras, H.; Lawal, H.O.; Krantz, D.E.; Bradley, T.J.; Walker, D.W.
Neuronal expression of a single-subunit yeast NADH-ubiquinone oxidoreductase (Ndi1) extends Drosophila lifespan
Aging Cell
9
191-202
2010
Saccharomyces cerevisiae
Iwata, M.; Lee, Y.; Yamashita, T.; Yagi, T.; Iwata, S.; Cameron, A.D.; Maher, M.J.
The structure of the yeast NADH dehydrogenase (Ndi1) reveals overlapping binding sites for water- and lipid-soluble substrates
Proc. Natl. Acad. Sci. USA
109
15247-15252
2012
Saccharomyces cerevisiae
Matus-Ortega, M.G.; Cardenas-Monroy, C.A.; Flores-Herrera, O.; Mendoza-Hernandez, G.; Miranda, M.; Gonzalez-Pedrajo, B.; Vazquez-Meza, H.; Pardo, J.P.
New complexes containing the internal alternative NADH dehydrogenase (Ndi1) in mitochondria of Saccharomyces cerevisiae
Yeast
32
629-641
2015
Saccharomyces cerevisiae (P32340), Saccharomyces cerevisiae
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