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Literature summary for 1.1.1.206 extracted from

  • Kushwaha, A.K.; Sangwan, N.S.; Tripathi, S.; Sangwan, R.S.
    Molecular cloning and catalytic characterization of a recombinant tropine biosynthetic tropinone reductase from Withania coagulans leaf (2013), Gene, 516, 238-247.
    View publication on PubMed
Search for more literature for 1.1.1.206

Cloned(Commentary)

Cloned (Comment) Organism
gene TRI, isolated from aerial tissue, DNA and amino acid sequence determination and analysis, sequence comparisons and phylogenetic analysis, overexpression of the His-tagged enzyme in Escherichia coli strain BL21 (DE3) Withania coagulans
gene WcTR-I, from leaves, DNA and amino acid sequence determination and analysis, sequence comparisons, phylogenetic analysis and tree, recombinant expression of His6-tagged enzyme in Escherichia coli strain BL21(DE3) Withania coagulans

Inhibitors

Inhibitors Comment Organism Structure
K+ slight inhibitory effect at 5 mM Withania coagulans
Li+ slight inhibitory effect at 5 mM Withania coagulans
Na+ slight inhibitory effect at 5 mM Withania coagulans
NH4+ slight inhibitory effect at 5 mM Withania coagulans

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
 additional information the in vitro reaction kinetics predominantly favour the reaction direction of tropinone reduction, Michaelis-Menten kinetics Withania coagulans
0.126
-
 NADPH pH 6.6, 30°C, recombinant enzyme Withania coagulans
0.127
-
 NADPH pH 6.6, 30°C, recombinant His-tagged enzyme Withania coagulans
0.208
-
 NADP+ pH 6.6, 30°C, recombinant enzyme Withania coagulans
0.208
-
 NADP+ pH 6.6, 30°C, recombinant His-tagged enzyme Withania coagulans
1.451
-
 tropinone pH 6.6, 30°C, recombinant enzyme Withania coagulans
1.451
-
 tropinone pH 6.6, 30°C, recombinant His-tagged enzyme Withania coagulans
26.31
-
 Tropine pH 6.6, 30°C, recombinant enzyme Withania coagulans
26.31
-
 Tropine pH 6.6, 30°C, recombinant His-tagged enzyme Withania coagulans

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
29340
-
 x * 29340, sequence calculation Withania coagulans

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
additional information Withania coagulans the enzyme TR-I has a wide substrate specificity but does not cover the substrates of other well-known plant SDR related to menthol metabolism ?
-
 ?
tropinone + NADPH + H+ Withania coagulans preferred substrate tropine + NADP+
-
 r
tropinone + NADPH + H+ Withania coagulans the reaction direction of tropinone reduction is predominantly operational in planta tropine + NADP+
-
 r

Organism

Organism UniProt Comment Textmining
Withania coagulans L7QI79
-
-

Purification (Commentary)

Purification (Comment) Organism
recombinant His-tagged enzyme from Escherichia coli strain BL21 (DE3) by nickel affinity chromatography Withania coagulans
recombinant His6-tagged enzyme from Escherichia coli strain BL21(DE3) by nickel affinity chromatography Withania coagulans

Source Tissue

Source Tissue Comment Organism Textmining
leaf
-
 Withania coagulans
-

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
1-methylpyrrolidine + NADPH + H+
-
 Withania coagulans ? + NADP+
-
 ?
1-propyl-4-piperidone + NADPH + H+
-
 Withania coagulans ? + NADP+
-
 ?
2-pyrrolidone + NADPH + H+
-
 Withania coagulans ? + NADP+
-
 ?
3-methylcyclohexanone + NADPH + H+
-
 Withania coagulans ? + NADP+
-
 ?
3-methylcyclohexanone + NADPH + H+
-
 Withania coagulans 3-methylcyclohexanol + NADP+
-
 ?
4-chlor-1-methylpiperidine + NADPH + H+
-
 Withania coagulans ? + NADP+
-
 ?
4-ethylcyclohexanone + NADPH + H+
-
 Withania coagulans 4-ethylcyclohexanol + NADP+
-
 ?
4-methylcyclohexanone + NADPH + H+
-
 Withania coagulans 4-methylcyclohexanol + NADP+
-
 ?
4-piperidone + NADPH + H+
-
 Withania coagulans ? + NADP+
-
 ?
exo-6-hydroxytropinone + NADPH + H+
-
 Withania coagulans ? + NADP+
-
 ?
additional information the enzyme TR-I has a wide substrate specificity but does not cover the substrates of other well-known plant SDR related to menthol metabolism Withania coagulans ?
-
 ?
additional information no activity by TR-I with 1-methyl-2-pyrrolidone Withania coagulans ?
-
 ?
additional information wide substrate specificity, overview. No activity with 1-methyl-2-pyrrolidone. The reverse reaction of tropine oxidation is highly specific and none of the tested alcohols can be oxidized with either NADP+ or NAD+ as cofactor Withania coagulans ?
-
 ?
N-methyl-4-piperidone + NADPH + H+
-
 Withania coagulans ? + NADP+
-
 ?
N-methylpiperidine + NADPH + H+
-
 Withania coagulans ? + NADP+
-
 ?
tetrahydro-4H-thiopyran-4-one + NADPH + H+
-
 Withania coagulans ? + NADP+
-
 ?
tropine + NADP+ in vitro reaction kinetics show that the enzyme predominantly favours the reverse reaction Withania coagulans tropinone + NADPH + H+
-
 r
tropinone + NADPH + H+ best substrate Withania coagulans tropine + NADP+
-
 r
tropinone + NADPH + H+ preferred substrate Withania coagulans tropine + NADP+
-
 r
tropinone + NADPH + H+ the reaction direction of tropinone reduction is predominantly operational in planta Withania coagulans tropine + NADP+
-
 r
tropinone + NADPH + H+ the in vitro reaction kinetics predominantly favour the reaction direction of tropinone reduction Withania coagulans tropine + NADP+
-
 r

Subunits

Subunits Comment Organism
? x * 29340, sequence calculation Withania coagulans
More superimposition of three-dimensional models of tropinone reductases,overview Withania coagulans
More a three dimensional model is prepared by taking Datura stramonium TR-II, PDB ID 1ipf, as template, structure comparison, overview Withania coagulans

Synonyms

Synonyms Comment Organism
TR-I
-
 Withania coagulans
tropine biosynthetic tropinone reductase
-
 Withania coagulans
tropinone reductase I
-
 Withania coagulans
WcTR-I
-
 Withania coagulans

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
30
-
 assay at Withania coagulans

Temperature Stability [°C]

Temperature Stability Minimum [°C] Temperature Stability Maximum [°C] Comment Organism
50
-
 purified recombinant enzyme, 30 min, over 92% of maximal activity remaining Withania coagulans
50 80 purified recombinant His-tagged enzyme, pH 6.6, 30 min, over 92% activity remaining at up to 50°C, 30% remaining at 60°C, 7% at 70°C, complete inactivation occurs at 80°C Withania coagulans
60
-
 purified recombinant enzyme, 30 min, 30% of maximal activity remaining Withania coagulans
70
-
 purified recombinant enzyme, 30 min, 7% of maximal activity remaining Withania coagulans
80
-
 purified recombinant enzyme, 30 min, inactivation Withania coagulans

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
1.87
-
 Tropine pH 6.6, 30°C, recombinant enzyme Withania coagulans
1.87
-
 Tropine pH 6.6, 30°C, recombinant His-tagged enzyme Withania coagulans
2.94
-
 NADP+ pH 6.6, 30°C, recombinant enzyme Withania coagulans
2.94
-
 NADP+ pH 6.6, 30°C, recombinant His-tagged enzyme Withania coagulans
6.76
-
 NADPH pH 6.6, 30°C, recombinant enzyme Withania coagulans
6.76
-
 NADPH pH 6.6, 30°C, recombinant His-tagged enzyme Withania coagulans
16.74
-
 tropinone pH 6.6, 30°C, recombinant enzyme Withania coagulans
16.74
-
 tropinone pH 6.6, 30°C, recombinant His-tagged enzyme Withania coagulans

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
6.6
-
-
 Withania coagulans
6.6
-
 tropinone reduction Withania coagulans

pH Range

pH Minimum pH Maximum Comment Organism
additional information
-
 narrow pH range in tropinone reduction Withania coagulans
6.1 7 narrow pH range Withania coagulans

Cofactor

Cofactor Comment Organism Structure
additional information the enzyme has absolute specificity for NADPH/NADP+. The affinity of the enzyme for NADPH is much higher than for NADP+ compared to NADPH Withania coagulans
NADP+
-
 Withania coagulans
NADPH
-
 Withania coagulans

pI Value

Organism Comment pI Value Maximum pI Value
Withania coagulans sequence calculation
-
 6.12

General Information

General Information Comment Organism
evolution tropinone reductase TR-I belongs to the short chain dehydrogenase/reductase family, SDR, of enzymes and has a YSASK as the signature YXXXK motif of SDRs, catalytic tetrade comprises N, S, Y, and K residues, overview Withania coagulans
evolution tropinone reductases (TRs) are small proteins belonging to the SDR (short chain dehydrogenase/reductase) family of enzymes. The enzyme's sequence contains the signature YXXXK motif of SDRs. Residues His112, Ala160, Val168, Ile223 and Phe226 are conserved in the TRs, and conserved at corresponding positions in WcTR-I, and involved in stereospecificity of the respective TR-Is. Tropane alkaloids might have evolved independently in plants, at least in Solanaceae and Erythroxylaceae Withania coagulans
metabolism the enzyme catalyzes a step intermediary enroute to biosynthesis of tropane esters of medicinal importance, i.e. hyoscyamine/scopolamine Withania coagulans
metabolism enzymes TR-I and TR-II catalyze the conversion of tropinone into tropane alcohols (tropine and pseudotropine, respectively). The steps are intermediary enroute to biosynthesis of tropane esters of medicinal importance, hyoscyamine/scopolamine, and calystegins, respectively. The enzyme has a wide substrate specificity but does not cover the substrates of other plant SDR enzymes related to menthol metabolism. The leaf tissue expression of the enzyme with these catalytic features suggests physiologically much prodigal rate of operation of the tropane alkaloid pathway in the leaf tissue itself Withania coagulans
additional information superimposition of three-dimensional models of tropinone reductases,overview Withania coagulans
additional information a three dimensional model is prepared by taking Datura stramonium TR-II, PDB ID 1ipf, as template, structure comparison, overview Withania coagulans

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
0.071
-
 Tropine pH 6.6, 30°C, recombinant enzyme Withania coagulans
0.071
-
 Tropine pH 6.6, 30°C, recombinant His-tagged enzyme Withania coagulans
11.53
-
 tropinone pH 6.6, 30°C, recombinant enzyme Withania coagulans
11.53
-
 tropinone pH 6.6, 30°C, recombinant His-tagged enzyme Withania coagulans
14
-
 NADP+ pH 6.6, 30°C, recombinant enzyme Withania coagulans
14
-
 NADP+ pH 6.6, 30°C, recombinant His-tagged enzyme Withania coagulans
53
-
 NADPH pH 6.6, 30°C, recombinant enzyme Withania coagulans
53
-
 NADPH pH 6.6, 30°C, recombinant His-tagged enzyme Withania coagulans