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Literature summary for 1.14.14.149 extracted from

  • Takahashi, S.; Zhao, Y.; O'Maille, P.E.; Greenhagen, B.T.; Noel, J.P.; Coates, R.M.; Chappell, J.
    Kinetic and molecular analysis of 5-epiaristolochene 1,3-dihydroxylase, a cytochrome P450 enzyme catalyzing successive hydroxylations of sesquiterpenes (2005), J. Biol. Chem., 280, 3686-3696.
    View publication on PubMedView publication on EuropePMC
Search for more literature for 1.14.14.149

Activating Compound

Activating Compound Comment Organism Structure
dimethyl sulfoxide maximum conversion of 5-epiaristolochene to capsidiol activity is observed at final concentrations of 2-5% (v/v) dimethyl sulfoxide Nicotiana tabacum

Cloned(Commentary)

Cloned (Comment) Organism
expressed in WAT11 yeast strain Nicotiana tabacum

Protein Variants

Protein Variants Comment Organism
I468A the mutant produces significant amounts 1beta-hydroxy-5-epiaristolochene, but negligible amounts of capsidiol from 5-epiaristolochene Nicotiana tabacum
S368A the mutant retains its ability to fully convert 5-epiaristolochene to capsidiol, but the turnover rates for capsidiol formation is 3-13times lower compared with that of the wild type enzyme and is also able to convert 1beta-hydroxy-5-epiaristolochene to capsidiol with kcat values comparable with that of wild type enzyme, but with Km values for 1beta-hydroxy-5-epiaristolochene 4-6times greater compared with that of wild type Nicotiana tabacum
S368C the mutant exhibits wild type catalytic efficiency for 1beta-hydroxy-5-epiaristolochene biosynthesis, but is devoid of the successive hydroxylation activity for capsidiol biosynthesis Nicotiana tabacum
S368F devoid of any hydroxylase activity for 5-epiaristolochene or 1beta-hydroxy-5-epiaristolochene Nicotiana tabacum
S368I devoid of any hydroxylase activity for 5-epiaristolochene or 1beta-hydroxy-5-epiaristolochene Nicotiana tabacum
S368T the mutant retains its ability to fully convert 5-epiaristolochene to capsidiol, but the turnover rates for capsidiol formation is 3-13times lower compared with that of the wild type enzyme and is also able to convert 1beta-hydroxy-5-epiaristolochene to capsidiol with kcat values comparable with that of wild type enzyme, but with Km values for 1beta-hydroxy-5-epiaristolochene 4-6times greater compared with that of wild type Nicotiana tabacum
S368V the mutant exhibits wild type catalytic efficiency for 1beta-hydroxy-5-epiaristolochene biosynthesis, but is devoid of the successive hydroxylation activity for capsidiol biosynthesis, the mutant catalyzes the relative equal biosynthesis of 1beta-hydroxy-5-epiaristolochene, 2beta-hydroxy-5-epiaristolochene, and 3beta-hydroxy-5-epiaristolochene from 5-epiaristolochene with wild type efficiency and converts about 1.5% of these monohydroxylated products to their respective ketone forms Nicotiana tabacum

Inhibitors

Inhibitors Comment Organism Structure
dimethyl sulfoxide conversion of 5-epiaristolochene to capsidiol activity is inhibited at concentrations above 10% (v/v) dimethyl sulfoxide Nicotiana tabacum

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.00174
-
 1beta-hydroxy-5-epiaristolochene wild type enzyme, the reaction product is capsidiol, in 100 mM Tris-HCl (pH 7.5), at 30°C Nicotiana tabacum
0.0062
-
 1beta-hydroxy-5-epiaristolochene mutant enzyme S368A, the reaction product is capsidiol, in 100 mM Tris-HCl (pH 7.5), at 30°C Nicotiana tabacum
0.00997
-
 1beta-hydroxy-5-epiaristolochene mutant enzyme S368T, the reaction product is capsidiol, in 100 mM Tris-HCl (pH 7.5), at 30°C Nicotiana tabacum
0.01518
-
 5-epiaristolochene mutant enzyme S368T, the reaction product is capsidiol, in 100 mM Tris-HCl (pH 7.5), at 30°C Nicotiana tabacum
0.01672
-
 5-epiaristolochene mutant enzyme S386V, the reaction product is 1beta-hydroxy-5-epiaristolochene, in 100 mM Tris-HCl (pH 7.5), at 30°C Nicotiana tabacum
0.01918
-
 5-epiaristolochene wild type enzyme, the reaction product is capsidiol, in 100 mM Tris-HCl (pH 7.5), at 30°C Nicotiana tabacum
0.02124
-
 1beta-hydroxy-5-epiaristolochene mutant enzyme I486A, the reaction product is capsidiol, in 100 mM Tris-HCl (pH 7.5), at 30°C Nicotiana tabacum
0.02267
-
 5-epiaristolochene mutant enzyme S368A, the reaction product is capsidiol, in 100 mM Tris-HCl (pH 7.5), at 30°C Nicotiana tabacum
0.03863
-
 5-epiaristolochene mutant enzyme S386C, the reaction product is 1beta-hydroxy-5-epiaristolochene, in 100 mM Tris-HCl (pH 7.5), at 30°C Nicotiana tabacum
0.0688
-
 5-epiaristolochene mutant enzyme I486A, the reaction product is 1beta-hydroxy-5-epiaristolochene, in 100 mM Tris-HCl (pH 7.5), at 30°C Nicotiana tabacum

Organism

Organism UniProt Comment Textmining
Nicotiana tabacum Q94FM7
-
-

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
1beta-hydroxy-5-epiaristolochene + [reduced NADPH-hemoprotein reductase] + O2 the catalytic efficiency for 1beta-hydroxy-5-epiaristolochene is about 10times greater than that for 3alpha-hydroxy-5-epiaristolochene Nicotiana tabacum capsidiol + [oxidized NADPH-hemoprotein reductase] + H2O
-
 ?
3alpha-hydroxy-5-epiaristolochene + [reduced NADPH-hemoprotein reductase] + O2 the catalytic efficiency for 3alpha-hydroxy-5-epiaristolochene is about 10times lower than that for 1beta-hydroxy-5-epiaristolochene Nicotiana tabacum capsidiol + [oxidized NADPH-hemoprotein reductase] + H2O
-
 ?
5-epiaristolochene + 2 [reduced NADPH-hemoprotein reductase] + 2 O2
-
 Nicotiana tabacum capsidiol + 2 [oxidized NADPH-hemoprotein reductase] + 2 H2O
-
 ?
5-epiaristolochene + [reduced NADPH-hemoprotein reductase] + O2 the release of an 1beta-hydroxy-5-epiaristolochene intermediate at high 5-epiaristolochene concentrations and a 10fold catalytic preference for 1beta-hydroxy-5-epiaristolochene versus 3alpha-hydroxy-5-epiaristolochene is indicative of a preferred reaction order of hydroxylation at C-1, followed by that at C-3 Nicotiana tabacum 1beta-hydroxy-5-epiaristolochene + [oxidized NADPH-hemoprotein reductase] + H2O
-
 ?

Synonyms

Synonyms Comment Organism
EAH
-
 Nicotiana tabacum

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.037
-
 5-epiaristolochene mutant enzyme S368T, the reaction product is capsidiol, in 100 mM Tris-HCl (pH 7.5), at 30°C Nicotiana tabacum
0.132
-
 1beta-hydroxy-5-epiaristolochene mutant enzyme I486A, the reaction product is capsidiol, in 100 mM Tris-HCl (pH 7.5), at 30°C Nicotiana tabacum
0.154
-
 5-epiaristolochene mutant enzyme S368A, the reaction product is capsidiol, in 100 mM Tris-HCl (pH 7.5), at 30°C Nicotiana tabacum
0.278
-
 5-epiaristolochene mutant enzyme S386C, the reaction product is 1beta-hydroxy-5-epiaristolochene, in 100 mM Tris-HCl (pH 7.5), at 30°C Nicotiana tabacum
0.293
-
 5-epiaristolochene mutant enzyme I486A, the reaction product is 1beta-hydroxy-5-epiaristolochene, in 100 mM Tris-HCl (pH 7.5), at 30°C Nicotiana tabacum
0.482
-
 1beta-hydroxy-5-epiaristolochene mutant enzyme S368T, the reaction product is capsidiol, in 100 mM Tris-HCl (pH 7.5), at 30°C Nicotiana tabacum
0.493
-
 5-epiaristolochene wild type enzyme, the reaction product is capsidiol, in 100 mM Tris-HCl (pH 7.5), at 30°C Nicotiana tabacum
0.553
-
 5-epiaristolochene mutant enzyme S386V, the reaction product is 1beta-hydroxy-5-epiaristolochene, in 100 mM Tris-HCl (pH 7.5), at 30°C Nicotiana tabacum
0.581
-
 1beta-hydroxy-5-epiaristolochene mutant enzyme S368A, the reaction product is capsidiol, in 100 mM Tris-HCl (pH 7.5), at 30°C Nicotiana tabacum
0.582
-
 1beta-hydroxy-5-epiaristolochene wild type enzyme, the reaction product is capsidiol, in 100 mM Tris-HCl (pH 7.5), at 30°C Nicotiana tabacum

Cofactor

Cofactor Comment Organism Structure
heme
-
 Nicotiana tabacum

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
2.4
-
 5-epiaristolochene mutant enzyme S368T, the reaction product is capsidiol, in 100 mM Tris-HCl (pH 7.5), at 30°C Nicotiana tabacum
4.3
-
 5-epiaristolochene mutant enzyme I486A, the reaction product is 1beta-hydroxy-5-epiaristolochene, in 100 mM Tris-HCl (pH 7.5), at 30°C Nicotiana tabacum
6.2
-
 1beta-hydroxy-5-epiaristolochene mutant enzyme I486A, the reaction product is capsidiol, in 100 mM Tris-HCl (pH 7.5), at 30°C Nicotiana tabacum
6.8
-
 5-epiaristolochene mutant enzyme S368A, the reaction product is capsidiol, in 100 mM Tris-HCl (pH 7.5), at 30°C Nicotiana tabacum
7.2
-
 5-epiaristolochene mutant enzyme S386C, the reaction product is 1beta-hydroxy-5-epiaristolochene, in 100 mM Tris-HCl (pH 7.5), at 30°C Nicotiana tabacum
25.7
-
 5-epiaristolochene wild type enzyme, the reaction product is capsidiol, in 100 mM Tris-HCl (pH 7.5), at 30°C Nicotiana tabacum
33.1
-
 5-epiaristolochene mutant enzyme S386V, the reaction product is 1beta-hydroxy-5-epiaristolochene, in 100 mM Tris-HCl (pH 7.5), at 30°C Nicotiana tabacum
48.3
-
 1beta-hydroxy-5-epiaristolochene mutant enzyme S368T, the reaction product is capsidiol, in 100 mM Tris-HCl (pH 7.5), at 30°C Nicotiana tabacum
93.7
-
 1beta-hydroxy-5-epiaristolochene mutant enzyme S368A, the reaction product is capsidiol, in 100 mM Tris-HCl (pH 7.5), at 30°C Nicotiana tabacum
334.4
-
 1beta-hydroxy-5-epiaristolochene wild type enzyme, the reaction product is capsidiol, in 100 mM Tris-HCl (pH 7.5), at 30°C Nicotiana tabacum