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Information on EC 1.14.14.151 - premnaspirodiene oxygenase
for references in articles please use BRENDA:EC1.14.14.151
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EC Tree 1 Oxidoreductases
1.14 Acting on paired donors, with incorporation or reduction of molecular oxygen
1.14.14 With reduced flavin or flavoprotein as one donor, and incorporation of one atom of oxygen into the other donor
1.14.14.151 premnaspirodiene oxygenase
IUBMB Comments
A cytochrome P-450 (heme-thiolate) protein. The enzyme from the plant Hyoscymus muticus also hydroxylates valencene at C-2 to give the α-hydroxy compound, nootkatol, and this is converted into nootkatone. 5-Epiaristolochene and epieremophilene are hydroxylated at C-2 to give a 2β-hydroxy derivatives that are not oxidized further.
The enzyme appears in viruses and cellular organisms
- 1.14.14.151
-
hyoscyamus
- muticus
-
+-valencene
- aroma
-
environmentally
-
regio
-
sesquiterpenoid
-
pichia
- grapefruit
- nootkatensis
-
+-nootkatone
- pastoris
Reaction Schemes
showSynonyms
premnaspirodiene oxygenase, cyp71d55, more
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SYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
CYP71D55
EC 1.14.13.121
-
-
formerly
-
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REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT
LITERATURE
(-)-vetispiradiene + 2 [reduced NADPH-hemoprotein reductase] + 2 O2 = solavetivone + 2 [oxidized NADPH-hemoprotein reductase] + 3 H2O
overall reaction
-
-
-
(-)-vetispiradiene + [reduced NADPH-hemoprotein reductase] + O2 = solavetivol + [oxidized NADPH-hemoprotein reductase] + H2O
(1a)
-
-
-
solavetivol + [reduced NADPH-hemoprotein reductase] + O2 = solavetivone + [oxidized NADPH-hemoprotein reductase] + 2 H2O
(1b)
-
-
-
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REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
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PATHWAY SOURCE
PATHWAYS
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SYSTEMATIC NAME
IUBMB Comments
()-vetispiradiene,[reduced NADPH-hemoprotein reductase]:oxygen 2alpha-oxidoreductase
A cytochrome P-450 (heme-thiolate) protein. The enzyme from the plant Hyoscymus muticus also hydroxylates valencene at C-2 to give the alpha-hydroxy compound, nootkatol, and this is converted into nootkatone. 5-Epiaristolochene and epieremophilene are hydroxylated at C-2 to give a 2beta-hydroxy derivatives that are not oxidized further.
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SUBSTRATE
PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
LITERATURE
COMMENTARY
Reversibility
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
()-vetispiradiene + 2 [reduced NADPH-hemoprotein reductase] + 2 O2
solavetivone + 2 [oxidized NADPH-hemoprotein reductase] + 3 H2O
Substrates: overall reaction
Products: -
Products: -
?
(+)-valencene + [reduced NADPH-hemoprotein reductase] + O2
alpha-nootkatol + beta-nootkatol + [oxidized NADPH-hemoprotein reductase] + H2O
(-)-vetispiradiene + 2 [reduced NADPH-hemoprotein reductase] + 2 O2
solavetivone + 2 [oxidized NADPH-hemoprotein reductase] + 3 H2O
(-)-vetispiradiene + [reduced NADPH-hemoprotein reductase] + O2
solavetivol + [oxidized NADPH-hemoprotein reductase] + H2O
Substrates: -
Products: solavetivol is also named premnaspirodien-2alpha-ol or (2S,4R,5S,7R)-spirovetiva-1(10),11(12)-dien-2-ol
Products: solavetivol is also named premnaspirodien-2alpha-ol or (2S,4R,5S,7R)-spirovetiva-1(10),11(12)-dien-2-ol
?
4-epi-eremophilene + [reduced NADPH-hemoprotein reductase] + O2
4-epieremophilen-2beta-ol + [oxidized NADPH-hemoprotein reductase] + H2O
Substrates: -
Products: -
Products: -
?
5-epiaristolochene + 3 [reduced NADPH-hemoprotein reductase] + 3 O2
2beta-hydroxy-5-epiaristolochene + 1beta-hydroxy-5-epiaristolochene + 3alpha-hydroxy-5-epiaristolochene + 3 [oxidized NADPH-hemoprotein reductase] + 3 H2O
Substrates: substrate affinity for 5-epiaristolochene is 2-4times lower than that for valencene or (-)-premnaspirodiene, respectively. HPO catalyzes the conversion of 5-epiaristolochene to four mono-hydroxylated products with 2beta-hydroxy-epiaristolochene comprising greater than 80% of the reaction products, 1beta-hydroxy-epiaristolochene about 5%, 3alpha-hydroxy-epiaristolochene less than 2%, and an unknown mono-hydroxylated product of about 20%
Products: -
Products: -
?
cedr-8-ene + [reduced NADPH-hemoprotein reductase] + O2
cedr-8-en-15-ol + [oxidized NADPH-hemoprotein reductase] + H2O
Substrates: -
Products: -
Products: -
?
resveratrol + 2 [reduced NADPH-hemoprotein reductase] + 2 O2
piceatannol + 2 [oxidized NADPH-hemoprotein reductase] + 3 H2O
-
Substrates: -
Products: -
Products: -
?
valencene + 2 [reduced NADPH-hemoprotein reductase] + 2 O2
alpha-nootkatol + beta-nootkatol + 2 [oxidized NADPH-hemoprotein reductase] + 2 H2O
Substrates: -
Products: -
Products: -
?
(+)-valencene + [reduced NADPH-hemoprotein reductase] + O2
alpha-nootkatol + beta-nootkatol + [oxidized NADPH-hemoprotein reductase] + H2O
Substrates: enzyme HPO-mediated biohydroxylation of (+)-valencene
Products: -
Products: -
?
(+)-valencene + [reduced NADPH-hemoprotein reductase] + O2
alpha-nootkatol + beta-nootkatol + [oxidized NADPH-hemoprotein reductase] + H2O
Substrates: -
Products: -
Products: -
?
(-)-vetispiradiene + 2 [reduced NADPH-hemoprotein reductase] + 2 O2
solavetivone + 2 [oxidized NADPH-hemoprotein reductase] + 3 H2O
Substrates: overall reaction
Products: -
Products: -
?
(-)-vetispiradiene + 2 [reduced NADPH-hemoprotein reductase] + 2 O2
solavetivone + 2 [oxidized NADPH-hemoprotein reductase] + 3 H2O
-
Substrates: overall reaction
Products: -
Products: -
?
additional information
?
-
Substrates: HPO also catalyzes the equivalent regio-specific (C-2) hydroxylation of several eremophilane-type (decalin ring system) sesquiterpenes, such as with 5-epi-aristolochene
Products: -
Products: -
?
additional information
?
-
-
Substrates: HPO also catalyzes the equivalent regio-specific (C-2) hydroxylation of several eremophilane-type (decalin ring system) sesquiterpenes, such as with 5-epi-aristolochene
Products: -
Products: -
?
additional information
?
-
Substrates: the enzyme fails to convert nootkatol to nootkatone
Products: -
Products: -
?
additional information
?
-
-
Substrates: the enzyme fails to convert nootkatol to nootkatone
Products: -
Products: -
?
additional information
?
-
Substrates: co-expression of enzyme HPO and cytochrome P450 reductase (CPR) leads to the hydroxylation of externally added (+)-valencene to trans-nootkatol in Pichia pastoris cells
Products: -
Products: -
?
additional information
?
-
-
Substrates: co-expression of enzyme HPO and cytochrome P450 reductase (CPR) leads to the hydroxylation of externally added (+)-valencene to trans-nootkatol in Pichia pastoris cells
Products: -
Products: -
?
additional information
?
-
Substrates: GC-MS metabolite analysis
Products: -
Products: -
?
additional information
?
-
-
Substrates: premnaspirodiene oxygenase is a cytochrome P450 enzyme functioning in heme-iron binding and oxidoreductase activity
Products: -
Products: -
?
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NATURAL SUBSTRATE
NATURAL PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
LITERATURE
COMMENTARY
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
(+)-valencene + [reduced NADPH-hemoprotein reductase] + O2
alpha-nootkatol + beta-nootkatol + [oxidized NADPH-hemoprotein reductase] + H2O
(-)-vetispiradiene + 2 [reduced NADPH-hemoprotein reductase] + 2 O2
solavetivone + 2 [oxidized NADPH-hemoprotein reductase] + 3 H2O
resveratrol + 2 [reduced NADPH-hemoprotein reductase] + 2 O2
piceatannol + 2 [oxidized NADPH-hemoprotein reductase] + 3 H2O
-
Substrates: -
Products: -
Products: -
?
(+)-valencene + [reduced NADPH-hemoprotein reductase] + O2
alpha-nootkatol + beta-nootkatol + [oxidized NADPH-hemoprotein reductase] + H2O
Substrates: enzyme HPO-mediated biohydroxylation of (+)-valencene
Products: -
Products: -
?
(+)-valencene + [reduced NADPH-hemoprotein reductase] + O2
alpha-nootkatol + beta-nootkatol + [oxidized NADPH-hemoprotein reductase] + H2O
Substrates: -
Products: -
Products: -
?
(-)-vetispiradiene + 2 [reduced NADPH-hemoprotein reductase] + 2 O2
solavetivone + 2 [oxidized NADPH-hemoprotein reductase] + 3 H2O
Substrates: overall reaction
Products: -
Products: -
?
(-)-vetispiradiene + 2 [reduced NADPH-hemoprotein reductase] + 2 O2
solavetivone + 2 [oxidized NADPH-hemoprotein reductase] + 3 H2O
-
Substrates: overall reaction
Products: -
Products: -
?
additional information
?
-
Substrates: HPO also catalyzes the equivalent regio-specific (C-2) hydroxylation of several eremophilane-type (decalin ring system) sesquiterpenes, such as with 5-epi-aristolochene
Products: -
Products: -
?
additional information
?
-
-
Substrates: HPO also catalyzes the equivalent regio-specific (C-2) hydroxylation of several eremophilane-type (decalin ring system) sesquiterpenes, such as with 5-epi-aristolochene
Products: -
Products: -
?
additional information
?
-
Substrates: co-expression of enzyme HPO and cytochrome P450 reductase (CPR) leads to the hydroxylation of externally added (+)-valencene to trans-nootkatol in Pichia pastoris cells
Products: -
Products: -
?
additional information
?
-
-
Substrates: co-expression of enzyme HPO and cytochrome P450 reductase (CPR) leads to the hydroxylation of externally added (+)-valencene to trans-nootkatol in Pichia pastoris cells
Products: -
Products: -
?
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COFACTOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
NADPH-hemoprotein reductase
A flavoprotein containing both FMN and FAD. This enzyme catalyses the transfer of electrons from NADPH, an obligatory two-electron donor, to microsomal P-450 monooxygenases, EC 1.14.14._
-
NADPH-hemoprotein reductase
KY366226
A flavoprotein containing both FMN and FAD. This enzyme catalyses the transfer of electrons from NADPH, an obligatory two-electron donor, to microsomal P-450 monooxygenases, EC 1.14.14._
-
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METALS and IONS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
Fe2+
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.0062
(-)-vetispiradiene
mutant enzyme V482I, in 100 mM Tris-HCl, pH 7.5, at 30°C
0.0084
(-)-vetispiradiene
mutant enzyme V480I/A484I, in 100 mM Tris-HCl, pH 7.5, at 30°C
0.013
(-)-vetispiradiene
mutant enzyme V482I/A484I, in 100 mM Tris-HCl, pH 7.5, at 30°C
0.014
(-)-vetispiradiene
wild type enzyme, in 100 mM Tris-HCl, pH 7.5, at 30°C
0.0025
5-epiaristolochene
mutant enzyme V480I/A484I, product is 2beta-hydroxy-epiaristolochene, in 100 mM Tris-HCl, pH 7.5, at 30°C
0.0033
5-epiaristolochene
wild type enzyme, product is 2beta-hydroxy-epiaristolochene, in 100 mM Tris-HCl, pH 7.5, at 30°C
0.0043
5-epiaristolochene
mutant enzyme V482I/A484I, product is 2beta-hydroxy-epiaristolochene, in 100 mM Tris-HCl, pH 7.5, at 30°C
0.0179
5-epiaristolochene
mutant enzyme V482I, product is 2beta-hydroxy-epiaristolochene, in 100 mM Tris-HCl, pH 7.5, at 30°C
0.0069
valencene
mutant enzyme V482I, product is beta-nootkatol, in 100 mM Tris-HCl, pH 7.5, at 30°C
0.0074
valencene
wild type enzyme, product is beta-nootkatol, in 100 mM Tris-HCl, pH 7.5, at 30°C
0.0121
valencene
mutant enzyme V482I/A484I, product is beta-nootkatol, in 100 mM Tris-HCl, pH 7.5, at 30°C
0.0192
valencene
mutant enzyme V480I/A484I, product is beta-nootkatol, in 100 mM Tris-HCl, pH 7.5, at 30°C
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TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
2.1
(-)-vetispiradiene
wild type enzyme, in 100 mM Tris-HCl, pH 7.5, at 30°C
3.8
(-)-vetispiradiene
mutant enzyme V482I, in 100 mM Tris-HCl, pH 7.5, at 30°C
12.7
(-)-vetispiradiene
mutant enzyme V480I/A484I, in 100 mM Tris-HCl, pH 7.5, at 30°C
20.7
(-)-vetispiradiene
mutant enzyme V482I/A484I, in 100 mM Tris-HCl, pH 7.5, at 30°C
0.2
5-epiaristolochene
wild type enzyme, product is 2beta-hydroxy-epiaristolochene, in 100 mM Tris-HCl, pH 7.5, at 30°C
0.7
5-epiaristolochene
mutant enzyme V480I/A484I, product is 2beta-hydroxy-epiaristolochene, in 100 mM Tris-HCl, pH 7.5, at 30°C
1.4
5-epiaristolochene
mutant enzyme V482I, product is 2beta-hydroxy-epiaristolochene, in 100 mM Tris-HCl, pH 7.5, at 30°C
2.8
5-epiaristolochene
mutant enzyme V482I/A484I, product is 2beta-hydroxy-epiaristolochene, in 100 mM Tris-HCl, pH 7.5, at 30°C
1.9
valencene
wild type enzyme, product is beta-nootkatol, in 100 mM Tris-HCl, pH 7.5, at 30°C
5.1
valencene
mutant enzyme V482I, product is beta-nootkatol, in 100 mM Tris-HCl, pH 7.5, at 30°C
5.7
valencene
mutant enzyme V480I/A484I, product is beta-nootkatol, in 100 mM Tris-HCl, pH 7.5, at 30°C
15.9
valencene
mutant enzyme V482I/A484I, product is beta-nootkatol, in 100 mM Tris-HCl, pH 7.5, at 30°C
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kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
150
(-)-vetispiradiene
wild type enzyme, in 100 mM Tris-HCl, pH 7.5, at 30°C
610
(-)-vetispiradiene
mutant enzyme V482I, in 100 mM Tris-HCl, pH 7.5, at 30°C
1510
(-)-vetispiradiene
mutant enzyme V480I/A484I, in 100 mM Tris-HCl, pH 7.5, at 30°C
1590
(-)-vetispiradiene
mutant enzyme V482I/A484I, in 100 mM Tris-HCl, pH 7.5, at 30°C
60
5-epiaristolochene
wild type enzyme, product is 2beta-hydroxy-epiaristolochene, in 100 mM Tris-HCl, pH 7.5, at 30°C
80
5-epiaristolochene
mutant enzyme V482I, product is 2beta-hydroxy-epiaristolochene, in 100 mM Tris-HCl, pH 7.5, at 30°C
280
5-epiaristolochene
mutant enzyme V480I/A484I, product is 2beta-hydroxy-epiaristolochene, in 100 mM Tris-HCl, pH 7.5, at 30°C
650
5-epiaristolochene
mutant enzyme V482I/A484I, product is 2beta-hydroxy-epiaristolochene, in 100 mM Tris-HCl, pH 7.5, at 30°C
260
valencene
wild type enzyme, product is beta-nootkatol, in 100 mM Tris-HCl, pH 7.5, at 30°C
300
valencene
mutant enzyme V480I/A484I, product is beta-nootkatol, in 100 mM Tris-HCl, pH 7.5, at 30°C
740
valencene
mutant enzyme V482I, product is beta-nootkatol, in 100 mM Tris-HCl, pH 7.5, at 30°C
1310
valencene
mutant enzyme V482I/A484I, product is beta-nootkatol, in 100 mM Tris-HCl, pH 7.5, at 30°C
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ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
Phytophthora tolerant black pepper cultivar IISR-Shakthi and susceptible cultivar IISR-Subhakara
-
-
brenda
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SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
SOURCE
-
transcriptome analysis in Phytophthora tolerant black pepper cultivar IISR-Shakthi and susceptible cultivar IISR-Subhakara
brenda
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LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY
GeneOntology No.
LITERATURE
SOURCE
-
determination of a predicted signal peptide and transmembrane topology in the sequence
brenda
Highest Expressing Human Cell Lines
Cell Line LinksPlease wait a moment until the data is sorted. This message will disappear when the data is sorted.
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
physiological function
-
involvement of signal proteins and defence enzymes premnaspirodiene oxygenase, phosphatase 2C-like domain-containing protein, nitrous oxide reductase family maturation protein, putative disease resistance protein RGA3, aspartyl protease, beta-glucosidase enzyme, cP450 signal protein, serine/threonine protein kinase WAG1 and nucleoredoxin 1-1 enzyme in field tolerance of black pepper to foot rot oomycete Phytophthora capsici, molecular mechanisms, overview. Foot rot or quick wilt caused by oomycete Phytophthora capsici is a serious disease in black pepper with blight on runner shoots, foliage, spikes and branches, spike shedding, defoliation, die back, collar and foot rot and death of plants. Premnaspirodiene oxygenase is involved in the hydroxylation of resveratrol to piceatannol, a phytoalexin, by which it promotes the intolerance against the pathogen
additional information
KY366226
overexpression of rice premnaspirodiene oxygenase reduces the infection rate of Xanthomonas oryzae pv. oryzae. Gene OsCYP71 does not contain a signal peptide
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UNIPROT
ENTRY NAME
ORGANISM
NO. OF AA
NO. OF TRANSM. HELICES
MOLECULAR WEIGHT[Da]
SOURCE
SEQUENCE
LOCALIZATION PREDICTION
The reliability class of the localization prediction ranges from 1 (very reliable) to 5 (not reliable).
The reliability class of the localization prediction ranges from 1 (very reliable) to 5 (not reliable). Please wait a moment until the data is sorted. This message will disappear when the data is sorted.
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
V480S
the mutation improves the kcat for the conversion of (-)-vetispiradiene to solavetivol about 2fold
V482I
V482I/A484I
additional information
V482I
the mutation improves the kcat for the conversion of (-)-vetispiradiene to solavetivol about 2fold
V482I/A484I
the mutant possesses a 5fold improvement in its catalytic efficiency for nootkatol biosynthesis and a 10fold improvement for 2beta-hydroxy-epiaristolochene biosynthesis
V482I/A484I
site-directed mutagenesis, the mutant shows a fivefold improved catalytic efficiency in the oxidation of (+)-valencene compared to wild-type
additional information
production of the sesquiterpenoid (+)-nootkatone by metabolic engineering of Pichia pastoris by generation of a strain co-expressing the premnaspirodiene oxygenase of Hyoscyamus muticus (HPO) and the Arabidopsis thaliana cytochrome P450 reductase (CPR) that hydroxylates extracellularly added (+)-valencene. Intracellular production of (+)-valencene by co-expression of valencene synthase from Callitropsis nootkatensis resolves the phase-transfer issues of (+)-valencene. Bi-phasic cultivations of Pichia pastoris result in the production of trans-nootkatol, which is oxidized to (+)-nootkatone by an intrinsic Pichia pastoris activity. Additional overexpression of a Pichia pastoris alcohol dehydrogenase and truncated hydroxy-methylglutaryl-CoA reductase (tHmg1p) significantly enhances the (+)-nootkatone yield to 208 mg/l cell culture in bioreactor cultivations. After 12 h of biotransformation about 50% of added (+)-valencene is converted to (+)-nootkatone without residual trans-nootkatol or ot herby-products, but with a moderate overall yield of 48% due to high substrate loss overtime. HPO,CPR and ADH-C3 protein levels are only marginally decreased by co-overexpression of tHMG1
additional information
-
production of the sesquiterpenoid (+)-nootkatone by metabolic engineering of Pichia pastoris by generation of a strain co-expressing the premnaspirodiene oxygenase of Hyoscyamus muticus (HPO) and the Arabidopsis thaliana cytochrome P450 reductase (CPR) that hydroxylates extracellularly added (+)-valencene. Intracellular production of (+)-valencene by co-expression of valencene synthase from Callitropsis nootkatensis resolves the phase-transfer issues of (+)-valencene. Bi-phasic cultivations of Pichia pastoris result in the production of trans-nootkatol, which is oxidized to (+)-nootkatone by an intrinsic Pichia pastoris activity. Additional overexpression of a Pichia pastoris alcohol dehydrogenase and truncated hydroxy-methylglutaryl-CoA reductase (tHmg1p) significantly enhances the (+)-nootkatone yield to 208 mg/l cell culture in bioreactor cultivations. After 12 h of biotransformation about 50% of added (+)-valencene is converted to (+)-nootkatone without residual trans-nootkatol or ot herby-products, but with a moderate overall yield of 48% due to high substrate loss overtime. HPO,CPR and ADH-C3 protein levels are only marginally decreased by co-overexpression of tHMG1
additional information
metabolic engineering Saccharomyces cerevisiae for de novo production of the sesquiterpenoid (+)-nootkatone. As the direct precursor of (+)-nootkatone biosynthesis, (+)-valencene is first produced in large quantities in Saccharomyces cerevisiae by overexpressing (+)-valencene synthase CnVS of Callitropsis nootkatensis in combination with various mevalonate pathway (MVA) engineering strategies, including the expression of CnVS and farnesyl diphosphate synthase (ERG20) as a fused protein, overexpression of a truncated form of the rate-limiting enzyme 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) reductase (tHMG1), and downregulating the squalene synthase enzyme (ERG9). These approaches altogether bring the production of (+)-valencene to 217.95 mg/l. Secondly, the (+)-valencene oxidation is adressed by overexpressing the Hyoscyamus muticus premnaspirodiene oxygenase (HPO) variant V482I/A484I and cytochrome P450 reductase (ATR1) from Arabidopsis thaliana. (+)-Valencene is predominantly oxidized to beta-nootkatol, and only minor amounts of (+)-nootkatone (9.66 mg/l) are produced. The short-chain dehydrogenase/reductase (SDR) superfamily dehydrogenases ZSD1 of Zingiber zerumbet and ABA2 of Citrus sinensis are capable of effectively catalyzing beta-nootkatol oxidation to (+)-nootkatone. The yield of (+)-nootkatone increases to 59.78 mg/l and 53.48 mg/l by combined overexpression of ZSD1 and ABA2, respectively. Metabolites GC analysis. Pathway reconstruction mechanism, overview
additional information
development of a viable bioprocess using premnaspirodiene oxygenase from Hyoscyamus muticus (HPO), cytochrome P450 reductase from Arabidopsis thaliana (AtCPR) and alcohol dehydrogenase (ADH1) from Saccharomyces cerevisiae overexpressed in CEN-PK2-1Ca, yielding beta-nootkatol and (+)-nootkatone with 170.5 and 45.6 mg/l ethyl acetate, respectively. A combinational engineering strategy including promoter change, regulator ROX1 knockout, squalene pathway inhibition, and tHMGR overexpression is performed to achieve de novo (+)-valencene production. Subsequent culture investigations show that galactose as the induced carbon source and a lower temperature of 25°C are beneficial to target accumulation. Considering the toxic effects of (+)-valencene to Saccharomyces cerevisiae, the (+)-valencene synthase gene under a weaker endogenous promoter of TEF1 (PTEF1) is cloned, yielding the recombinant yeast of PK2-14. Also, replacing the inducible promoters (GAL1) of HPO and AtCPR with constitutive promoters, HXT7 and CYC1, increases the beta-nootkatol accumulation from 108.2 to 327.8 mg/l ethyl acetate in resting-cell experiments using (+)-valencene as a substrate. The total terpenoid titer of the engineered strain of PK2-25 using glucose as a carbon source is improved 56fold to 157.8 mg/l cell culture. Bi-phasic batch cultivation in shake flasks. Method development and evaluation, overview
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CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
co-expression of C-terminally FLAG-tagged premnaspirodiene oxygenase of Hyoscyamus muticus (HPO) and the Arabidopsis thaliana cytochrome P450 reductase (CPR) in Pichia pastoris strains ADH-C1 and -C3
CYP71D55, recombinant expression in Saccharomyces cerevisiae strain PK2-25 under control of a constitutive promoter, coexpression with cytochrome P450 reductase from Arabidopsis thaliana (AtCPR) and alcohol dehydrogenase (ADH1) from Saccharomyces cerevisiae
DNA and amino acid sequence determination and analysis, sequence comparisons, quantitative DDRT-PCR enzyme expression analysis
-
gene OsCYP71, DNA and amino acid seuence determination and analysis, sequence comparisons and phylogenetic analysis. Overexpression of full-length cDNA of premnaspirodiene oxygenase from Oryza sativa subsp. japonica in the wild-type, Dongjin, which is susceptible to bacterial blight Korean strain K2, transformed via Agrobacterium tumefaciens strain EHA105 reduces the infection rate of Xanthomonas oryzae pv. oryzae
KY366226
overexpression of premnaspirodiene oxygenase mutant V482I/A484I in Saccharomyces cerevisiae strain W303
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EXPRESSION
ORGANISM
UNIPROT
LITERATURE
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REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Takahashi, S.; Yeo, Y.S.; Zhao, Y.; O'Maille, P.E.; Greenhagen, B.T.; Noel, J.P.; Coates, R.M.; Chappell, J.
Functional characterization of premnaspirodiene oxygenase, a cytochrome P450 catalyzing regio- and stereo-specific hydroxylations of diverse sesquiterpene substrates
J. Biol. Chem.
282
31744-31754
2007
Hyoscyamus muticus (A6YIH8), Hyoscyamus muticus
brenda
Nino, M.; Song, J.; Nogoy, F.; Kim, M.; Jung, Y.; Kang, K.; Nou, I.; Cho, Y.
Overexpression of rice premnaspirodiene oxygenase reduces the infection rate of Xanthomonas oryzae pv. oryzae
J. Plant Biotechnol.
43
422-431
2016
Oryza sativa Japonica Group (KY366226)
-
brenda
Wriessnegger, T.; Augustin, P.; Engleder, M.; Leitner, E.; Mueller, M.; Kaluzna, I.; Schuermann, M.; Mink, D.; Zellnig, G.; Schwab, H.; Pichler, H.
Production of the sesquiterpenoid (+)-nootkatone by metabolic engineering of Pichia pastoris
Metab. Eng.
24
18-29
2014
Hyoscyamus muticus (A6YIH8), Hyoscyamus muticus
brenda
Meng, X.; Liu, H.; Xu, W.; Zhang, W.; Wang, Z.; Liu, W.
Metabolic engineering Saccharomyces cerevisiae for de novo production of the sesquiterpenoid (+)-nootkatone
Microb. Cell Fact.
19
21
2020
Hyoscyamus muticus (A6YIH8)
brenda
Babu Paul, B.; Mathew, D.; Beena, S.; Shylaja, M.
Comparative transcriptome analysis reveals the signal proteins and defence genes conferring foot rot (Phytophthora capsici sp. nov.) resistance in black pepper (Piper nigrum L.)
Physiol. Mol. Plant Pathol.
108
101436
2019
Piper nigrum
-
brenda
Ouyang, X.; Cha, Y.; Li, W.; Zhu, C.; Zhu, M.; Li, S.; Zhuo, M.; Huang, S.; Li, J.
Stepwise engineering of Saccharomyces cerevisiae to produce (+)-valencene and its related sesquiterpenes
RSC Adv.
9
30171-30181
2019
Hyoscyamus muticus (A6YIH8), no activity in Saccharomyces cerevisiae strain PK2-C
brenda
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