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Literature summary extracted from

  • Morina, K.; Schulte, M.; Hubrich, F.; Doerner, K.; Steimle, S.; Stolpe, S.; Friedrich, T.
    Engineering the respiratory complex I to energy-converting NADPH:ubiquinone oxidoreductase (2011), J. Biol. Chem., 286, 34627-34634.
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

EC Number Cloned (Comment) Organism
7.1.1.2 expression of wild-type and mutant enzymes in Escherichia coli Escherichia coli

Protein Variants

EC Number Protein Variants Comment Organism
7.1.1.2 E183D site-directed mutagenesis, the mutant shows a 2fold increase in NADH/ferricyanide oxidoreductase activity and a 5fold increase in NADPHoxidase activity compared to the wild-type enzyme Escherichia coli
7.1.1.2 E183H site-directed mutagenesis, the mutant shows a 2fold increase in NADH/ferricyanide oxidoreductase activity and a 2fold increase in NADPHoxidase activitycompared to the wild-type enzyme Escherichia coli
7.1.1.2 E183N site-directed mutagenesis, the mutant shows a 2fold increase in NADH/ferricyanide oxidoreductase activity and a 2fold increase in NADPHoxidase activitycompared to the wild-type enzyme Escherichia coli
7.1.1.2 E183Q site-directed mutagenesis, the mutant shows a similar NADH/ferricyanide oxidoreductase activity and a 2fold increase NADPHoxidase activity compared to the wild-type enzyme Escherichia coli
7.1.1.2 additional information construction of several variants with mutations at position 183 exhibiting up to 200fold enhanced catalytic efficiency with NADPH Escherichia coli

Inhibitors

EC Number Inhibitors Comment Organism Structure
7.1.1.2 Piericidin A
-
 Escherichia coli

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
7.1.1.2 0.0057
-
 NADH mutant E183H, pH 6.0, 30°C Escherichia coli
7.1.1.2 0.0058
-
 NADH mutant E183D, pH 6.0, 30°C Escherichia coli
7.1.1.2 0.012
-
 NADH mutant E183Q, pH 6.0, 30°C Escherichia coli
7.1.1.2 0.013
-
 NADH wild-type enzyme, pH 6.0, 30°C Escherichia coli
7.1.1.2 0.014
-
 NADH mutant E183N, pH 6.0, 30°C Escherichia coli
7.1.1.2 0.025
-
 NADPH mutant E183H, pH 6.0, 30°C Escherichia coli
7.1.1.2 0.045
-
 NADPH mutant E183Q, pH 6.0, 30°C Escherichia coli
7.1.1.2 0.39
-
 NADPH mutant E183D, pH 6.0, 30°C Escherichia coli
7.1.1.2 0.48
-
 NADPH mutant E183N, pH 6.0, 30°C Escherichia coli
7.1.1.2 1.87
-
 NADPH wild-type enzyme, pH 6.0, 30°C Escherichia coli

Localization

EC Number Localization Comment Organism GeneOntology No. Textmining
7.1.1.2 membrane
-
 Escherichia coli 16020
-

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
7.1.1.2 NADH + ubiquinone + 6 H+[side 1] Escherichia coli
-
 NAD+ + ubiquinol + 7 H+[side 2]
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
7.1.1.2 Escherichia coli
-
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
7.1.1.2 recombinant wild-type and mutant enzymes from Escherichia coli Escherichia coli

Specific Activity [micromol/min/mg]

EC Number Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
7.1.1.2 additional information
-
 NADH/ferricyanide oxidoreductase activity (measured as ferricyanide reduction) and NAD(P)H oxidase activity (measured as oxygen consumption) of cytoplasmic membranes from Escherichia coli recombinantly expressing the enzyme Escherichia coli

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
7.1.1.2 NADH + ubiquinone + 6 H+[side 1]
-
 Escherichia coli NAD+ + ubiquinol + 7 H+[side 2]
-
 ?
7.1.1.2 NADPH + ubiquinone + 6 H+[side 1] NADPH is a poor substrate of the complex Escherichia coli NADP+ + ubiquinol + 7 H+[side 2]
-
 ?

Subunits

EC Number Subunits Comment Organism
7.1.1.2 More the enzyme nucleotide-binding site is made up of a unique Rossmann fold to accommodate the binding of the substrate NADH and of the primary electron acceptor flavin mononucleotide Escherichia coli

Synonyms

EC Number Synonyms Comment Organism
7.1.1.2 complex I
-
 Escherichia coli
7.1.1.2 energy-converting NADPH:ubiquinone oxidoreductase
-
 Escherichia coli

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
7.1.1.2 30
-
 assay at Escherichia coli

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
7.1.1.2 2 8 NADH mutant E183Q, pH 6.0, 30°C Escherichia coli
7.1.1.2 3
-
 NADPH wild-type enzyme, pH 6.0, 30°C Escherichia coli
7.1.1.2 10
-
 NADPH mutant E183H, pH 6.0, 30°C Escherichia coli
7.1.1.2 11
-
 NADPH mutant E183N, pH 6.0, 30°C Escherichia coli
7.1.1.2 11
-
 NADH mutant E183N, pH 6.0, 30°C Escherichia coli
7.1.1.2 14
-
 NADPH mutant E183Q, pH 6.0, 30°C Escherichia coli
7.1.1.2 26
-
 NADH wild-type enzyme, pH 6.0, 30°C Escherichia coli
7.1.1.2 34
-
 NADPH mutant E183D, pH 6.0, 30°C Escherichia coli
7.1.1.2 37
-
 NADH mutants E183D and E18H, pH 6.0, 30°C Escherichia coli

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
7.1.1.2 6 7 mutant E183H, activity with NADH Escherichia coli
7.1.1.2 6
-
 mutant E183H, activity with NADPH Escherichia coli
7.1.1.2 6
-
 wild-type enzyme, activity with NADH, first peak Escherichia coli
7.1.1.2 6.5
-
 wild-type enzyme, activity with NADPH Escherichia coli
7.1.1.2 7.5
-
 wild-type enzyme, activity with NADH, second higher peak Escherichia coli

pH Range

EC Number pH Minimum pH Maximum Comment Organism
7.1.1.2 5 8 activity range Escherichia coli

Cofactor

EC Number Cofactor Comment Organism Structure
7.1.1.2 FMN
-
 Escherichia coli
7.1.1.2 additional information NADPH is a poor substrate of the complex Escherichia coli
7.1.1.2 NADH binding of NADH includes interactions of the hydroxyl groups of the adenosine ribose with a conserved glutamic acid residue. Structural analysis revealed that due to steric hindrance and electrostatic repulsion, this residue most likely prevents the binding of NADPH, which is a poor substrate of the complex Escherichia coli

General Information

EC Number General Information Comment Organism
7.1.1.2 physiological function the respiratory complex I couples the electron transfer from NADH to ubiquinone with a translocation of protons across the membrane Escherichia coli

kcat/KM [mM/s]

EC Number kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
7.1.1.2 1.8
-
 NADPH wild-type enzyme, pH 6.0, 30°C Escherichia coli
7.1.1.2 20
-
 NADPH mutant E183N, pH 6.0, 30°C Escherichia coli
7.1.1.2 90
-
 NADPH mutant E183D, pH 6.0, 30°C Escherichia coli
7.1.1.2 320
-
 NADPH mutant E183Q, pH 6.0, 30°C Escherichia coli
7.1.1.2 400
-
 NADPH mutant E183H, pH 6.0, 30°C Escherichia coli
7.1.1.2 800
-
 NADH mutant E183N, pH 6.0, 30°C Escherichia coli
7.1.1.2 2000
-
 NADH wild-type enzyme, pH 6.0, 30°C Escherichia coli
7.1.1.2 2300
-
 NADH mutant E183Q, pH 6.0, 30°C Escherichia coli
7.1.1.2 6400
-
 NADH mutant E183D, pH 6.0, 30°C Escherichia coli
7.1.1.2 6500
-
 NADH mutant E183H, pH 6.0, 30°C Escherichia coli