BRENDA - Enzyme Database

Purification and properties of soluble hydrogenase from Alcaligenes eutrophus H 16

Schneider, K.; Schlegel, H.G.; Biochim. Biophys. Acta 452, 66-80 (1976)

Data extracted from this reference:

General Stability
EC Number
General Stability
Organism
1.12.1.2
unstable in reduced active form
Cupriavidus necator
Inhibitors
EC Number
Inhibitors
Commentary
Organism
Structure
1.12.1.2
high ionic strength
200 mM phosphate buffer, 70% inhibition
Cupriavidus necator
KM Value [mM]
EC Number
KM Value [mM]
KM Value Maximum [mM]
Substrate
Commentary
Organism
Structure
1.12.1.2
0.037
-
H2
-
Cupriavidus necator
1.12.1.2
0.123
-
NAD+
-
Cupriavidus necator
1.12.1.2
0.56
-
NAD+
-
Cupriavidus necator
Localization
EC Number
Localization
Commentary
Organism
GeneOntology No.
Textmining
1.12.1.2
soluble
-
Cupriavidus necator
-
-
Metals/Ions
EC Number
Metals/Ions
Commentary
Organism
Structure
1.12.1.2
Iron
no increase of activity by addition of Co2+, Mn2+, Ni2+ or Fe2+; non-heme iron protein
Cupriavidus necator
1.12.1.2
additional information
no increase of activity by addition of Co2+, Mn2+, Ni2+ or Fe2+
Cupriavidus necator
Molecular Weight [Da]
EC Number
Molecular Weight [Da]
Molecular Weight Maximum [Da]
Commentary
Organism
1.12.1.2
204500
-
gel filtration
Cupriavidus necator
1.12.1.2
206000
-
sucrose density gradient centrifugation
Cupriavidus necator
Organism
EC Number
Organism
Primary Accession No. (UniProt)
Commentary
Textmining
1.12.1.2
Cupriavidus necator
-
; formerly Alcaligenes eutrophus
-
1.12.1.2
Cupriavidus necator H16 / ATCC 23440 / NCIB 10442 / S-10-1
-
-
-
Oxidation Stability
EC Number
Oxidation Stability
Organism
1.12.1.2
oxidized hydrogenase purified under aerobic conditions is highly stable but not reactive, reductive activation of the enzyme by H2 in the presence of catalytic amounts of NADH or by reducing agents destabilizes
Cupriavidus necator
Purification (Commentary)
EC Number
Commentary
Organism
1.12.1.2
sp. H16, cetavlon, ammonium sulfate, DEAE-cellulose, Sephadex G-200, hydroxyapatite
Cupriavidus necator
Specific Activity [micromol/min/mg]
EC Number
Specific Activity Minimum [µmol/min/mg]
Specific Activity Maximum [µmol/min/mg]
Commentary
Organism
1.12.1.2
54.46
-
-
Cupriavidus necator
Storage Stability
EC Number
Storage Stability
Organism
1.12.1.2
-20°C, O2, several months, no loss in activity
Cupriavidus necator
Substrates and Products (Substrate)
EC Number
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
1.12.1.2
2 ferricyanide + NAD(P)H
-
439699
Cupriavidus necator
2 ferrocyanide + NAD(P)+ + H+
-
439699
Cupriavidus necator
?
1.12.1.2
H2 + NAD+
electron acceptor FMN
439699
Cupriavidus necator
H+ + NADH
-
439699
Cupriavidus necator
r
1.12.1.2
H2 + NAD+
electron acceptor FAD
439699
Cupriavidus necator
H+ + NADH
-
439699
Cupriavidus necator
r
1.12.1.2
H2 + NAD+
artificial electron acceptors: Janus green, 2,6-dichlorophenolindophenol, phenazine methosulfate, menaquinone, ubiquinone, cytochrome c
439699
Cupriavidus necator
H+ + NADH
-
439699
Cupriavidus necator
r
1.12.1.2
H2 + NAD+
artificial electron acceptor benzyl viologen
439699
Cupriavidus necator
H+ + NADH
-
439699
Cupriavidus necator
r
1.12.1.2
H2 + NAD+
artificial electron acceptor methyl viologen
439699
Cupriavidus necator
H+ + NADH
-
439699
Cupriavidus necator
r
1.12.1.2
H2 + NAD+
weak electron acceptor O2
439699
Cupriavidus necator
H+ + NADH
-
439699
Cupriavidus necator
r
1.12.1.2
H2 + NAD+
enzyme also has diaphorase and NAD(P)H oxidase activity
439699
Cupriavidus necator
H+ + NADH
-
439699
Cupriavidus necator
r
1.12.1.2
H2 + NAD+
electron acceptor NAD+
439699
Cupriavidus necator
H+ + NADH
-
439699
Cupriavidus necator
r
1.12.1.2
H2 + NAD+
artificial electron acceptor ferricyanide
439699
Cupriavidus necator
H+ + NADH
-
439699
Cupriavidus necator
r
1.12.1.2
H2 + NAD+
artificial electron acceptor methylene blue
439699
Cupriavidus necator
H+ + NADH
-
439699
Cupriavidus necator
r
1.12.1.2
H2 + NAD+
electron acceptor FMN
439699
Cupriavidus necator H16 / ATCC 23440 / NCIB 10442 / S-10-1
H+ + NADH
-
439699
Cupriavidus necator H16 / ATCC 23440 / NCIB 10442 / S-10-1
r
1.12.1.2
H2 + NAD+
electron acceptor FAD
439699
Cupriavidus necator H16 / ATCC 23440 / NCIB 10442 / S-10-1
H+ + NADH
-
439699
Cupriavidus necator H16 / ATCC 23440 / NCIB 10442 / S-10-1
r
1.12.1.2
H2 + NAD+
artificial electron acceptors: Janus green, 2,6-dichlorophenolindophenol, phenazine methosulfate, menaquinone, ubiquinone, cytochrome c
439699
Cupriavidus necator H16 / ATCC 23440 / NCIB 10442 / S-10-1
H+ + NADH
-
439699
Cupriavidus necator H16 / ATCC 23440 / NCIB 10442 / S-10-1
r
1.12.1.2
H2 + oxidized benzyl viologen
-
439699
Cupriavidus necator
H+ + reduced benzyl viologen
-
439699
Cupriavidus necator
r
1.12.1.2
H2 + oxidized methyl viologen
-
439699
Cupriavidus necator
H+ + reduced methyl viologen
-
439699
Cupriavidus necator
r
1.12.1.2
H2 + oxidized methyl viologen
-
439699
Cupriavidus necator H16 / ATCC 23440 / NCIB 10442 / S-10-1
H+ + reduced methyl viologen
-
439699
Cupriavidus necator H16 / ATCC 23440 / NCIB 10442 / S-10-1
r
1.12.1.2
NAD(P)H + H+ + oxidized 2,6-dichlorophenolindophenol
-
439699
Cupriavidus necator
NAD(P)+ + reduced 2,6-dichlorophenolindophenol
-
439699
Cupriavidus necator
?
1.12.1.2
NAD(P)H + H+ + oxidized 2,6-dichlorophenolindophenol
-
439699
Cupriavidus necator H16 / ATCC 23440 / NCIB 10442 / S-10-1
NAD(P)+ + reduced 2,6-dichlorophenolindophenol
-
439699
Cupriavidus necator H16 / ATCC 23440 / NCIB 10442 / S-10-1
?
1.12.1.2
oxidized cytochrome c + NAD(P)H
-
439699
Cupriavidus necator
reduced cytchrome c + NAD(P)+
-
439699
Cupriavidus necator
?
1.12.1.2
oxidized methylene blue + NAD(P)H
-
439699
Cupriavidus necator
reduced methylene blue + NAD(P)+
-
439699
Cupriavidus necator
?
Temperature Optimum [°C]
EC Number
Temperature Optimum [°C]
Temperature Optimum Maximum [°C]
Commentary
Organism
1.12.1.2
33
-
-
Cupriavidus necator
pH Optimum
EC Number
pH Optimum Minimum
pH Optimum Maximum
Commentary
Organism
1.12.1.2
8
-
-
Cupriavidus necator
Cofactor
EC Number
Cofactor
Commentary
Organism
Structure
1.12.1.2
ATP
30% stimulation
Cupriavidus necator
1.12.1.2
flavin
flavoprotein
Cupriavidus necator
1.12.1.2
FMN
25% stimulation
Cupriavidus necator
Cofactor (protein specific)
EC Number
Cofactor
Commentary
Organism
Structure
1.12.1.2
ATP
30% stimulation
Cupriavidus necator
1.12.1.2
flavin
flavoprotein
Cupriavidus necator
1.12.1.2
FMN
25% stimulation
Cupriavidus necator
General Stability (protein specific)
EC Number
General Stability
Organism
1.12.1.2
unstable in reduced active form
Cupriavidus necator
Inhibitors (protein specific)
EC Number
Inhibitors
Commentary
Organism
Structure
1.12.1.2
high ionic strength
200 mM phosphate buffer, 70% inhibition
Cupriavidus necator
KM Value [mM] (protein specific)
EC Number
KM Value [mM]
KM Value Maximum [mM]
Substrate
Commentary
Organism
Structure
1.12.1.2
0.037
-
H2
-
Cupriavidus necator
1.12.1.2
0.123
-
NAD+
-
Cupriavidus necator
1.12.1.2
0.56
-
NAD+
-
Cupriavidus necator
Localization (protein specific)
EC Number
Localization
Commentary
Organism
GeneOntology No.
Textmining
1.12.1.2
soluble
-
Cupriavidus necator
-
-
Metals/Ions (protein specific)
EC Number
Metals/Ions
Commentary
Organism
Structure
1.12.1.2
Iron
no increase of activity by addition of Co2+, Mn2+, Ni2+ or Fe2+; non-heme iron protein
Cupriavidus necator
1.12.1.2
additional information
no increase of activity by addition of Co2+, Mn2+, Ni2+ or Fe2+
Cupriavidus necator
Molecular Weight [Da] (protein specific)
EC Number
Molecular Weight [Da]
Molecular Weight Maximum [Da]
Commentary
Organism
1.12.1.2
204500
-
gel filtration
Cupriavidus necator
1.12.1.2
206000
-
sucrose density gradient centrifugation
Cupriavidus necator
Oxidation Stability (protein specific)
EC Number
Oxidation Stability
Organism
1.12.1.2
oxidized hydrogenase purified under aerobic conditions is highly stable but not reactive, reductive activation of the enzyme by H2 in the presence of catalytic amounts of NADH or by reducing agents destabilizes
Cupriavidus necator
Purification (Commentary) (protein specific)
EC Number
Commentary
Organism
1.12.1.2
sp. H16, cetavlon, ammonium sulfate, DEAE-cellulose, Sephadex G-200, hydroxyapatite
Cupriavidus necator
Specific Activity [micromol/min/mg] (protein specific)
EC Number
Specific Activity Minimum [µmol/min/mg]
Specific Activity Maximum [µmol/min/mg]
Commentary
Organism
1.12.1.2
54.46
-
-
Cupriavidus necator
Storage Stability (protein specific)
EC Number
Storage Stability
Organism
1.12.1.2
-20°C, O2, several months, no loss in activity
Cupriavidus necator
Substrates and Products (Substrate) (protein specific)
EC Number
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
1.12.1.2
2 ferricyanide + NAD(P)H
-
439699
Cupriavidus necator
2 ferrocyanide + NAD(P)+ + H+
-
439699
Cupriavidus necator
?
1.12.1.2
H2 + NAD+
electron acceptor FMN
439699
Cupriavidus necator
H+ + NADH
-
439699
Cupriavidus necator
r
1.12.1.2
H2 + NAD+
electron acceptor FAD
439699
Cupriavidus necator
H+ + NADH
-
439699
Cupriavidus necator
r
1.12.1.2
H2 + NAD+
artificial electron acceptors: Janus green, 2,6-dichlorophenolindophenol, phenazine methosulfate, menaquinone, ubiquinone, cytochrome c
439699
Cupriavidus necator
H+ + NADH
-
439699
Cupriavidus necator
r
1.12.1.2
H2 + NAD+
artificial electron acceptor benzyl viologen
439699
Cupriavidus necator
H+ + NADH
-
439699
Cupriavidus necator
r
1.12.1.2
H2 + NAD+
artificial electron acceptor methyl viologen
439699
Cupriavidus necator
H+ + NADH
-
439699
Cupriavidus necator
r
1.12.1.2
H2 + NAD+
weak electron acceptor O2
439699
Cupriavidus necator
H+ + NADH
-
439699
Cupriavidus necator
r
1.12.1.2
H2 + NAD+
enzyme also has diaphorase and NAD(P)H oxidase activity
439699
Cupriavidus necator
H+ + NADH
-
439699
Cupriavidus necator
r
1.12.1.2
H2 + NAD+
electron acceptor NAD+
439699
Cupriavidus necator
H+ + NADH
-
439699
Cupriavidus necator
r
1.12.1.2
H2 + NAD+
artificial electron acceptor ferricyanide
439699
Cupriavidus necator
H+ + NADH
-
439699
Cupriavidus necator
r
1.12.1.2
H2 + NAD+
artificial electron acceptor methylene blue
439699
Cupriavidus necator
H+ + NADH
-
439699
Cupriavidus necator
r
1.12.1.2
H2 + NAD+
electron acceptor FMN
439699
Cupriavidus necator H16 / ATCC 23440 / NCIB 10442 / S-10-1
H+ + NADH
-
439699
Cupriavidus necator H16 / ATCC 23440 / NCIB 10442 / S-10-1
r
1.12.1.2
H2 + NAD+
electron acceptor FAD
439699
Cupriavidus necator H16 / ATCC 23440 / NCIB 10442 / S-10-1
H+ + NADH
-
439699
Cupriavidus necator H16 / ATCC 23440 / NCIB 10442 / S-10-1
r
1.12.1.2
H2 + NAD+
artificial electron acceptors: Janus green, 2,6-dichlorophenolindophenol, phenazine methosulfate, menaquinone, ubiquinone, cytochrome c
439699
Cupriavidus necator H16 / ATCC 23440 / NCIB 10442 / S-10-1
H+ + NADH
-
439699
Cupriavidus necator H16 / ATCC 23440 / NCIB 10442 / S-10-1
r
1.12.1.2
H2 + oxidized benzyl viologen
-
439699
Cupriavidus necator
H+ + reduced benzyl viologen
-
439699
Cupriavidus necator
r
1.12.1.2
H2 + oxidized methyl viologen
-
439699
Cupriavidus necator
H+ + reduced methyl viologen
-
439699
Cupriavidus necator
r
1.12.1.2
H2 + oxidized methyl viologen
-
439699
Cupriavidus necator H16 / ATCC 23440 / NCIB 10442 / S-10-1
H+ + reduced methyl viologen
-
439699
Cupriavidus necator H16 / ATCC 23440 / NCIB 10442 / S-10-1
r
1.12.1.2
NAD(P)H + H+ + oxidized 2,6-dichlorophenolindophenol
-
439699
Cupriavidus necator
NAD(P)+ + reduced 2,6-dichlorophenolindophenol
-
439699
Cupriavidus necator
?
1.12.1.2
NAD(P)H + H+ + oxidized 2,6-dichlorophenolindophenol
-
439699
Cupriavidus necator H16 / ATCC 23440 / NCIB 10442 / S-10-1
NAD(P)+ + reduced 2,6-dichlorophenolindophenol
-
439699
Cupriavidus necator H16 / ATCC 23440 / NCIB 10442 / S-10-1
?
1.12.1.2
oxidized cytochrome c + NAD(P)H
-
439699
Cupriavidus necator
reduced cytchrome c + NAD(P)+
-
439699
Cupriavidus necator
?
1.12.1.2
oxidized methylene blue + NAD(P)H
-
439699
Cupriavidus necator
reduced methylene blue + NAD(P)+
-
439699
Cupriavidus necator
?
Temperature Optimum [°C] (protein specific)
EC Number
Temperature Optimum [°C]
Temperature Optimum Maximum [°C]
Commentary
Organism
1.12.1.2
33
-
-
Cupriavidus necator
pH Optimum (protein specific)
EC Number
pH Optimum Minimum
pH Optimum Maximum
Commentary
Organism
1.12.1.2
8
-
-
Cupriavidus necator