Cloned (Comment) | Organism |
---|---|
expression as N-terminal His6-tagged enzyme in Escherichia coli strain M15 | Rauvolfia serpentina |
Crystallization (Comment) | Organism |
---|---|
purified recombinant methylated His6-tagged enzyme wild-type and mutant A213W, hanging drop vapor diffusion method, mixing of 0.002 ml of 5.5 mg/ml protein in 10 mM Tris-HCl buffer, pH 7.0, 1 mM DTT, 10 mM EDTA, with 0002 ml reservoir solution containing 25% v/v PEG 4000, 0.1 mM sodium citrate, pH 5.6, equilibration against 1 ml of reservoir solution at 20°C for 7 days, X-ray diffraction structure determination and analysis at 2.31 A and 1.77 A resolution, respectively | Rauvolfia serpentina |
Protein Variants | Comment | Organism |
---|---|---|
A213W | site-directed mutagenesis, cofactor-bound structure analysis | Rauvolfia serpentina |
additional information | methylation of wild-type and mutant enzymes for improvement of the crystallization process | Rauvolfia serpentina |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
perakine + NADPH + H+ | Rauvolfia serpentina | - |
raucaffrinoline + NADP+ | - |
? |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Rauvolfia serpentina | Q3L181 | - |
- |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
perakine + NADPH + H+ | - |
Rauvolfia serpentina | raucaffrinoline + NADP+ | - |
? |
Subunits | Comment | Organism |
---|---|---|
More | three-dimensional structure modeling of wild-type and mutant apo methylated enzymes with (alpha/beta)8-barrels with eight parallel beta-strands and eight alpha-helices typical for AKR superfamily members, overview | Rauvolfia serpentina |
Synonyms | Comment | Organism |
---|---|---|
AKR13D1 | - |
Rauvolfia serpentina |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
30 | - |
assay at | Rauvolfia serpentina |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
7 | - |
assay at | Rauvolfia serpentina |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
NADPH | dependent on, binding mode and structure, conformational changes caused by the cofactor binding, overview | Rauvolfia serpentina |
General Information | Comment | Organism |
---|---|---|
evolution | the enzyme belongs to the AKR13D subfamily of the aldo-keto reductases, evolutionary relationship of AKR13D from Rauvolfia serpentina to annotated AKR families and subfamilies | Rauvolfia serpentina |
additional information | the active site is formed by the catalytic tetrad Asp52, Tyr57, Lys84, and His126 at the center of the (alpha/beta)8-barrel structure. Upon NADPH binding, dramatic conformational changes and movements are observed: two additional beta-strands in the C terminus become ordered to form one alpha-helix, and a movement of up to 24 A occurs. This conformational change creates a large space that allows the binding of substrates of variable size for PR and enhances the enzyme activity | Rauvolfia serpentina |
physiological function | perakine reductase catalyzes the NADPH-dependent reduction of the aldehyde perakine to yield the alcohol raucaffrinoline in the biosynthetic pathway of ajmaline in Rauvolfia, a key step in indole alkaloid biosynthesis | Rauvolfia serpentina |