BRENDA - Enzyme Database
show all sequences of 1.1.1.317

Purification, cloning, functional expression and characterization of perakine reductase: the first example from the AKR enzyme family, extending the alkaloidal network of the plant Rauvolfia

Sun, L.; Ruppert, M.; Sheludko, Y.; Warzecha, H.; Zhao, Y.; Stöckigt, J.; Plant Mol. Biol. 67, 455-467 (2008)

Data extracted from this reference:

Cloned(Commentary)
Cloned (Commentary)
Organism
functionally expressed in Escherichia coli as the N-terminal His6-tagged protein
Rauvolfia serpentina
Engineering
Protein Variants
Commentary
Organism
D52A
inactive mutant enzyme
Rauvolfia serpentina
H126A
inactive mutant enzyme
Rauvolfia serpentina
K84A
inactive mutant enzyme
Rauvolfia serpentina
additional information
site-directed mutagenesis of each of the functional residues of catalytic tetrad Asp52, Tyr57, Lys84, His126 to an alanine residue results in more than 97.8% loss of enzyme activity
Rauvolfia serpentina
Y57A
inactive mutant enzyme
Rauvolfia serpentina
KM Value [mM]
KM Value [mM]
KM Value Maximum [mM]
Substrate
Commentary
Organism
Structure
0.28
-
4-nitrobenzaldehyde
pH 8.5, 37°C
Rauvolfia serpentina
0.83
-
perakine
pH 8.5, 37°C
Rauvolfia serpentina
Molecular Weight [Da]
Molecular Weight [Da]
Molecular Weight Maximum [Da]
Commentary
Organism
37159
-
x * 37159, calculated from sequence
Rauvolfia serpentina
38000
-
x * 38000, SDS-PAGE
Rauvolfia serpentina
Natural Substrates/ Products (Substrates)
Natural Substrates
Organism
Commentary (Nat. Sub.)
Natural Products
Commentary (Nat. Pro.)
Organism (Nat. Pro.)
Reversibility
ID
perakine + NADPH + H+
Rauvolfia serpentina
the biosynthesis of raucaffrinoline from perakine is a side route of the ajmaline biosynthesis pathway (monoterpenoid indole alkaloid)
raucaffrinoline + NADP+
-
-
?
Organism
Organism
UniProt
Commentary
Textmining
Rauvolfia serpentina
Q3L181
-
-
Purification (Commentary)
Purification (Commentary)
Organism
-
Rauvolfia serpentina
Substrates and Products (Substrate)
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
Substrate Product ID
4-nitrobenzaldehyde + NADPH + H+
-
716558
Rauvolfia serpentina
4-nitrobenzoate + NADP+
-
-
-
?
additional information
the enzyme also catalyses the reduction of coniferyl aldehyde, sinapyl aldehyde, 4-coumaryl aldehyde, 3-(3,4,5-trimethoxyphenyl)propenal
716558
Rauvolfia serpentina
?
-
-
-
?
perakine + NADPH + H+
the biosynthesis of raucaffrinoline from perakine is a side route of the ajmaline biosynthesis pathway (monoterpenoid indole alkaloid)
716558
Rauvolfia serpentina
raucaffrinoline + NADP+
-
-
-
?
perakine + NADPH + H+
NADPH can not be replaced by NADH. In presence of the oxidized cofactor NADP+ the reverse reaction of the reductase is low, with less than 0.5% relative activity of the forward reaction
716558
Rauvolfia serpentina
raucaffrinoline + NADP+
-
-
-
?
Subunits
Subunits
Commentary
Organism
?
x * 38000, SDS-PAGE
Rauvolfia serpentina
?
x * 37159, calculated from sequence
Rauvolfia serpentina
Temperature Optimum [°C]
Temperature Optimum [°C]
Temperature Optimum Maximum [°C]
Commentary
Organism
37
-
assay at
Rauvolfia serpentina
50
-
-
Rauvolfia serpentina
Turnover Number [1/s]
Turnover Number Minimum [1/s]
Turnover Number Maximum [1/s]
Substrate
Commentary
Organism
Structure
0.014
-
perakine
pH 8.5, 37°C
Rauvolfia serpentina
8.26
-
4-nitrobenzaldehyde
pH 8.5, 37°C
Rauvolfia serpentina
pH Optimum
pH Optimum Minimum
pH Optimum Maximum
Commentary
Organism
7
-
-
Rauvolfia serpentina
8.5
-
assay at
Rauvolfia serpentina
pH Range
pH Minimum
pH Maximum
Commentary
Organism
6.5
8.5
half-maximal activity
Rauvolfia serpentina
Cofactor
Cofactor
Commentary
Organism
Structure
NADPH
can not be replaced by NADH
Rauvolfia serpentina
pI Value
Organism
Commentary
pI Value Maximum
pI Value
Rauvolfia serpentina
calculated from sequence
-
5.9
Cloned(Commentary) (protein specific)
Commentary
Organism
functionally expressed in Escherichia coli as the N-terminal His6-tagged protein
Rauvolfia serpentina
Cofactor (protein specific)
Cofactor
Commentary
Organism
Structure
NADPH
can not be replaced by NADH
Rauvolfia serpentina
Engineering (protein specific)
Protein Variants
Commentary
Organism
D52A
inactive mutant enzyme
Rauvolfia serpentina
H126A
inactive mutant enzyme
Rauvolfia serpentina
K84A
inactive mutant enzyme
Rauvolfia serpentina
additional information
site-directed mutagenesis of each of the functional residues of catalytic tetrad Asp52, Tyr57, Lys84, His126 to an alanine residue results in more than 97.8% loss of enzyme activity
Rauvolfia serpentina
Y57A
inactive mutant enzyme
Rauvolfia serpentina
KM Value [mM] (protein specific)
KM Value [mM]
KM Value Maximum [mM]
Substrate
Commentary
Organism
Structure
0.28
-
4-nitrobenzaldehyde
pH 8.5, 37°C
Rauvolfia serpentina
0.83
-
perakine
pH 8.5, 37°C
Rauvolfia serpentina
Molecular Weight [Da] (protein specific)
Molecular Weight [Da]
Molecular Weight Maximum [Da]
Commentary
Organism
37159
-
x * 37159, calculated from sequence
Rauvolfia serpentina
38000
-
x * 38000, SDS-PAGE
Rauvolfia serpentina
Natural Substrates/ Products (Substrates) (protein specific)
Natural Substrates
Organism
Commentary (Nat. Sub.)
Natural Products
Commentary (Nat. Pro.)
Organism (Nat. Pro.)
Reversibility
ID
perakine + NADPH + H+
Rauvolfia serpentina
the biosynthesis of raucaffrinoline from perakine is a side route of the ajmaline biosynthesis pathway (monoterpenoid indole alkaloid)
raucaffrinoline + NADP+
-
-
?
Purification (Commentary) (protein specific)
Commentary
Organism
-
Rauvolfia serpentina
Substrates and Products (Substrate) (protein specific)
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
ID
4-nitrobenzaldehyde + NADPH + H+
-
716558
Rauvolfia serpentina
4-nitrobenzoate + NADP+
-
-
-
?
additional information
the enzyme also catalyses the reduction of coniferyl aldehyde, sinapyl aldehyde, 4-coumaryl aldehyde, 3-(3,4,5-trimethoxyphenyl)propenal
716558
Rauvolfia serpentina
?
-
-
-
?
perakine + NADPH + H+
the biosynthesis of raucaffrinoline from perakine is a side route of the ajmaline biosynthesis pathway (monoterpenoid indole alkaloid)
716558
Rauvolfia serpentina
raucaffrinoline + NADP+
-
-
-
?
perakine + NADPH + H+
NADPH can not be replaced by NADH. In presence of the oxidized cofactor NADP+ the reverse reaction of the reductase is low, with less than 0.5% relative activity of the forward reaction
716558
Rauvolfia serpentina
raucaffrinoline + NADP+
-
-
-
?
Subunits (protein specific)
Subunits
Commentary
Organism
?
x * 38000, SDS-PAGE
Rauvolfia serpentina
?
x * 37159, calculated from sequence
Rauvolfia serpentina
Temperature Optimum [°C] (protein specific)
Temperature Optimum [°C]
Temperature Optimum Maximum [°C]
Commentary
Organism
37
-
assay at
Rauvolfia serpentina
50
-
-
Rauvolfia serpentina
Turnover Number [1/s] (protein specific)
Turnover Number Minimum [1/s]
Turnover Number Maximum [1/s]
Substrate
Commentary
Organism
Structure
0.014
-
perakine
pH 8.5, 37°C
Rauvolfia serpentina
8.26
-
4-nitrobenzaldehyde
pH 8.5, 37°C
Rauvolfia serpentina
pH Optimum (protein specific)
pH Optimum Minimum
pH Optimum Maximum
Commentary
Organism
7
-
-
Rauvolfia serpentina
8.5
-
assay at
Rauvolfia serpentina
pH Range (protein specific)
pH Minimum
pH Maximum
Commentary
Organism
6.5
8.5
half-maximal activity
Rauvolfia serpentina
pI Value (protein specific)
Organism
Commentary
pI Value Maximum
pI Value
Rauvolfia serpentina
calculated from sequence
-
5.9
KCat/KM [mM/s]
kcat/KM Value [1/mMs-1]
kcat/KM Value Maximum [1/mMs-1]
Substrate
Commentary
Organism
Structure
0.016
-
perakine
pH 8.5, 37°C
Rauvolfia serpentina
29.5
-
4-nitrobenzaldehyde
pH 8.5, 37°C
Rauvolfia serpentina
KCat/KM [mM/s] (protein specific)
KCat/KM Value [1/mMs-1]
KCat/KM Value Maximum [1/mMs-1]
Substrate
Commentary
Organism
Structure
0.016
-
perakine
pH 8.5, 37°C
Rauvolfia serpentina
29.5
-
4-nitrobenzaldehyde
pH 8.5, 37°C
Rauvolfia serpentina
Other publictions for EC 1.1.1.317
No.
1st author
Pub Med
title
organims
journal
volume
pages
year
Activating Compound
Application
Cloned(Commentary)
Crystallization (Commentary)
Engineering
General Stability
Inhibitors
KM Value [mM]
Localization
Metals/Ions
Molecular Weight [Da]
Natural Substrates/ Products (Substrates)
Organic Solvent Stability
Organism
Oxidation Stability
Posttranslational Modification
Purification (Commentary)
Reaction
Renatured (Commentary)
Source Tissue
Specific Activity [micromol/min/mg]
Storage Stability
Substrates and Products (Substrate)
Subunits
Synonyms
Temperature Optimum [°C]
Temperature Range [°C]
Temperature Stability [°C]
Turnover Number [1/s]
pH Optimum
pH Range
pH Stability
Cofactor
Ki Value [mM]
pI Value
IC50 Value
Activating Compound (protein specific)
Application (protein specific)
Cloned(Commentary) (protein specific)
Cofactor (protein specific)
Crystallization (Commentary) (protein specific)
Engineering (protein specific)
General Stability (protein specific)
IC50 Value (protein specific)
Inhibitors (protein specific)
Ki Value [mM] (protein specific)
KM Value [mM] (protein specific)
Localization (protein specific)
Metals/Ions (protein specific)
Molecular Weight [Da] (protein specific)
Natural Substrates/ Products (Substrates) (protein specific)
Organic Solvent Stability (protein specific)
Oxidation Stability (protein specific)
Posttranslational Modification (protein specific)
Purification (Commentary) (protein specific)
Renatured (Commentary) (protein specific)
Source Tissue (protein specific)
Specific Activity [micromol/min/mg] (protein specific)
Storage Stability (protein specific)
Substrates and Products (Substrate) (protein specific)
Subunits (protein specific)
Temperature Optimum [°C] (protein specific)
Temperature Range [°C] (protein specific)
Temperature Stability [°C] (protein specific)
Turnover Number [1/s] (protein specific)
pH Optimum (protein specific)
pH Range (protein specific)
pH Stability (protein specific)
pI Value (protein specific)
Expression
General Information
General Information (protein specific)
Expression (protein specific)
KCat/KM [mM/s]
KCat/KM [mM/s] (protein specific)
725466
Sun
Crystal structure of perakine ...
Rauvolfia serpentina
J. Biol. Chem.
287
11213-11221
2012
-
-
1
1
2
-
-
-
-
-
-
1
-
3
-
-
-
-
-
-
-
-
1
1
1
1
-
-
-
1
-
-
1
-
-
-
-
-
1
1
1
2
-
-
-
-
-
-
-
-
1
-
-
-
-
-
-
-
-
1
1
1
-
-
-
1
-
-
-
-
3
3
-
-
-
716558
Sun
Purification, cloning, functio ...
Rauvolfia serpentina
Plant Mol. Biol.
67
455-467
2008
-
-
1
-
5
-
-
2
-
-
2
1
-
7
-
-
1
-
-
-
-
-
4
2
-
2
-
-
2
2
1
-
1
-
1
-
-
-
1
1
-
5
-
-
-
-
2
-
-
2
1
-
-
-
1
-
-
-
-
4
2
2
-
-
2
2
1
-
1
-
-
-
-
2
2
713623
Rosenthal
Expression, purification, crys ...
Rauvolfia serpentina
Acta Crystallogr. Sect. F
62
1286-1289
2006
-
-
1
1
-
-
-
-
-
-
-
-
-
1
-
-
1
-
-
-
-
-
1
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
1
-
1
-
-
-
-
-
-
-
-
-
-
-
-
-
1
-
-
-
-
1
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-