Cloned (Comment) | Organism |
---|---|
gene TR-I from roots, DNA and amino acid sequence determination and analysis, sequence comparisons and phylogenetic analysis and tree, recombinant functional overexpression of His6-tagged enzyme in Escherichia coli strain BL21(DE3) | Withania somnifera |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
additional information | - |
additional information | Michaelis-Menten kinetics | Withania somnifera | |
0.00043 | - |
NADPH | recombinant enzyme, pH 6.7, 30°C | Withania somnifera | |
0.0962 | - |
NADP+ | recombinant enzyme, pH 6.7, 30°C | Withania somnifera | |
1.27 | - |
tropinone | recombinant enzyme, pH 6.7, 30°C | Withania somnifera | |
1.74 | - |
N-methylpiperidine | recombinant enzyme, pH 6.7, 30°C | Withania somnifera | |
2.11 | - |
3-methylcyclohexanone | recombinant enzyme, pH 6.7, 30°C | Withania somnifera | |
114.9 | - |
Tropine | recombinant enzyme, pH 6.7, 30°C | Withania somnifera |
Metals/Ions | Comment | Organism | Structure |
---|---|---|---|
Mg2+ | slightly activating | Withania somnifera | |
additional information | except slight enhancement of enzyme activity by Mg2+ and Na+, none of the following cations, Mn2+, Li+, NH4+, Mg2+, Na+, and Co2+, 5 mM each, affects the activity significantly | Withania somnifera | |
Na+ | slightly activating | Withania somnifera |
Molecular Weight [Da] | Molecular Weight Maximum [Da] | Comment | Organism |
---|---|---|---|
60000 | - |
2 * 60000, SDS-PAGE | Withania somnifera |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
tropinone + NADPH + H+ | Withania somnifera | - |
tropine + NADP+ | - |
r |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Withania somnifera | U5PUZ1 | cv. NMITLI-118 | - |
Purification (Comment) | Organism |
---|---|
recombinant His6-tagged enzyme from Escherichia coli strain BL21(DE3) by affinity chromatography and gel filtration | Withania somnifera |
Source Tissue | Comment | Organism | Textmining |
---|---|---|---|
berry | low enzyme level, all developmental stages | Withania somnifera | - |
leaf | low enzyme level, all developmental stages | Withania somnifera | - |
additional information | enzyme activity and tropine/tropinone contents during different developmental stages, overview | Withania somnifera | - |
root | high enzyme level in bark, moderate in medulla | Withania somnifera | - |
shoot | high enzyme level | Withania somnifera | - |
stem | high enzyme level in bark, moderate in medulla | Withania somnifera | - |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
1-propyl-4-piperidone + NADPH + H+ | low activity | Withania somnifera | 1-propylpiperidin-4-ol + NADP+ | - |
? | |
2-pyrrolidone + NADPH + H+ | very low activity | Withania somnifera | 2-hydroxypyrrolidine + NADP+ | - |
? | |
3-methylcyclohexanone + NADPH + H+ | - |
Withania somnifera | 3-methylcyclohexanol + NADP+ | - |
? | |
4-chloro-1-methylpiperidine + NADPH + H+ | moderate activity | Withania somnifera | ? + NADP+ | - |
? | |
4-ethylcyclohexanone + NADPH + H+ | low activity | Withania somnifera | 4-ethylcyclohexanol + NADP+ | - |
? | |
4-methylcyclohexanone + NADPH + H+ | low activity | Withania somnifera | 4-methylcyclohexanol + NADP+ | - |
? | |
4-piperidone + NADPH + H+ | very low activity | Withania somnifera | 4-hydroxypiperidin + NADP+ | - |
? | |
exo-6-hydroxytropinone + NADPH + H+ | low activity | Withania somnifera | ? + NADP+ | - |
? | |
additional information | TLC and GC-MS analysis of substrates and products. The enzyme also shows comparatively efficient catalysis of NADPH-dependent reduction of cyclic ketones 3-methylcyclohexanone and N-methylpiperidine, substrate specificity, overview. No activity with 1-methyl-2-pyrrolidone | Withania somnifera | ? | - |
? | |
N-methyl-4-piperidone + NADPH + H+ | low activity | Withania somnifera | 1-methylpiperidin-4-ol + NADP+ | - |
? | |
N-methylpiperidine + NADPH + H+ | - |
Withania somnifera | ? + NADP+ | - |
? | |
tetrahydro-4H-thiopyran-4-one + NADPH + H+ | moderate activity | Withania somnifera | ? + NADP+ | - |
? | |
tropine + NADP+ | catalytic kinetics of the enzyme favour the forward reaction, tropine formation | Withania somnifera | tropinone + NADPH + H+ | - |
r | |
tropinone + NADPH + H+ | - |
Withania somnifera | tropine + NADP+ | - |
r |
Subunits | Comment | Organism |
---|---|---|
homodimer | 2 * 60000, SDS-PAGE | Withania somnifera |
More | the homodimer is the catalytically active form of the recombinant WsTR-I | Withania somnifera |
Synonyms | Comment | Organism |
---|---|---|
TR-I | - |
Withania somnifera |
tropinone reductase | - |
Withania somnifera |
tropinone reductase-I | - |
Withania somnifera |
WsTR-I | - |
Withania somnifera |
WsTR-I protein | - |
Withania somnifera |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
30 | - |
assay at | Withania somnifera |
Temperature Stability Minimum [°C] | Temperature Stability Maximum [°C] | Comment | Organism |
---|---|---|---|
50 | 60 | purified recombinant enzyme, retains about 75% of maximal catalytic activity at up to 50°C, 4.27% activity remaining on incubation at 60°C | Withania somnifera |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.54 | - |
NADP+ | recombinant enzyme, pH 6.7, 30°C | Withania somnifera | |
1.67 | - |
Tropine | recombinant enzyme, pH 6.7, 30°C | Withania somnifera | |
8.97 | - |
3-methylcyclohexanone | recombinant enzyme, pH 6.7, 30°C | Withania somnifera | |
9.23 | - |
N-methylpiperidine | recombinant enzyme, pH 6.7, 30°C | Withania somnifera | |
9.51 | - |
tropinone | recombinant enzyme, pH 6.7, 30°C | Withania somnifera | |
33.34 | - |
NADPH | recombinant enzyme, pH 6.7, 30°C | Withania somnifera |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
6.7 | - |
- |
Withania somnifera |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
NADP+ | - |
Withania somnifera | |
NADPH | - |
Withania somnifera |
Organism | Comment | Expression |
---|---|---|
Withania somnifera | the enzyme-encoding TR-I gene expression is upregulated on exposure to signal molecules, i.e. methyl jasmonate and salicylic acid, and on mechanical injury/wounding | up |
General Information | Comment | Organism |
---|---|---|
evolution | molecular evolution of WsTR-I, overview | Withania somnifera |
metabolism | the enzyme catalyzes the NADPH-dependent tropinone to tropine conversion step in tropane metabolism, metabolic pathway of tropane alkaloid biosynthesis, overview | Withania somnifera |
additional information | enzyme three-dimensional structure modelling | Withania somnifera |
physiological function | physiological role of tropinone reductase enzyme in tropane alkaloid biosynthesis in aerial tissues of the plant. The metabolic step of tropine formation may be regulated by, besides through the transcript and protein levels, the tissue concentration of tropinone | Withania somnifera |
kcat/KM Value [1/mMs-1] | kcat/KM Value Maximum [1/mMs-1] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.014 | - |
Tropine | recombinant enzyme, pH 6.7, 30°C | Withania somnifera | |
0.559 | - |
NADP+ | recombinant enzyme, pH 6.7, 30°C | Withania somnifera | |
4.38 | - |
N-methylpiperidine | recombinant enzyme, pH 6.7, 30°C | Withania somnifera | |
5.17 | - |
3-methylcyclohexanone | recombinant enzyme, pH 6.7, 30°C | Withania somnifera | |
7.49 | - |
tropinone | recombinant enzyme, pH 6.7, 30°C | Withania somnifera | |
77540 | - |
NADPH | recombinant enzyme, pH 6.7, 30°C | Withania somnifera |