Information on EC 4.2.3.32 - levopimaradiene synthase

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The enzyme appears in viruses and cellular organisms

EC NUMBER
COMMENTARY hide
4.2.3.32
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RECOMMENDED NAME
GeneOntology No.
levopimaradiene synthase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
(+)-copalyl diphosphate = abieta-8(14),12-diene + diphosphate
show the reaction diagram
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-
-
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REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
cyclization
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elimination of diphosphate
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PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
Biosynthesis of secondary metabolites
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Diterpenoid biosynthesis
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levopimaric acid biosynthesis
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SYSTEMATIC NAME
IUBMB Comments
(+)-copalyl-diphosphate diphosphate-lyase [abieta-8(14),12-diene-forming]
In Ginkgo, the enzyme catalyses the initial cyclization step in the biosynthesis of ginkgolides, a structurally unique family of diterpenoids that are highly specific platelet-activating-factor receptor antagonists [1]. Levopimaradiene is widely distributed in higher plants. In some species the enzyme also forms abietadiene, palustradiene, and neoabietadiene [2].
CAS REGISTRY NUMBER
COMMENTARY hide
369596-13-4
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
bifunctional levopimaradiene/abietadiene synthase
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-
Manually annotated by BRENDA team
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SwissProt
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
physiological function
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levopimaradiene synthase promoter-driven expression of beta-glucuronidase in Arabidopsis shows beta-glucuronidase accumulation mainly in the epidermis of leaves, phloem of the shoots, ovaries and stamens of flowers, and vasculature of roots. Treatment of methyl jasmonate on the transformant leads to significant upregulation of the reporter gene in the roots with little change in leaves and flowers. Findings support biosynthesis of ginkgolide in the roots of Ginkgo plant and suggest translocation occurs through the phloem
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
(+)-copalyl diphosphate
abieta-8(14),12-diene + diphosphate
show the reaction diagram
-
-
products are levopimaradiene, abieta-7,13-diene, and sandaracopimaradiene
-
?
(+)-copalyl-diphosphate
(-)-abietadiene + levopimaradiene + neoabietadiene + palustradiene + diphosphate
show the reaction diagram
(+)-copalyl-diphosphate
13-hydroxy-8(14)-abietene + diphosphate
show the reaction diagram
-
the initial products formed and released in vitro are unstable alcohols identified as epimeric thermally unstable allylic tertiary alcohols, 13-hydroxy-8(14)-abietene. The dehydration of the alcohol products, yielding the well established diterpene products levopimaradiene, abietadiene, neoabietadiene, and palustradiene, occurs due to the conditions of the GC-MS analysis typically used
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?
(+)-copalyl-diphosphate
levopimaradiene + diphosphate
show the reaction diagram
reaction intermediate is labdadienyl diphosphate
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-
?
additional information
?
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NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
additional information
?
-
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Mn2+
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required
additional information
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enzyme is not dependent on Mg2+
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
dithiothreitol
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optimal activity in the presence of 5 mM DTT and 5% glycerol
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
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resin duct epithelial cells are the main site of diterpene biosynthesis in Sitka spruce, diterpenoid biosynthesis is induced in cortical resin duct epithelial cells early upon treatment with methyljasmonate, and immature developing traumatic resin duct epithelial cells produce levopimaradiene/abietadiene synthase
Manually annotated by BRENDA team
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male strobili
Manually annotated by BRENDA team
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root of seedling
Manually annotated by BRENDA team
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
100289
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x * 100289, calculated
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
?
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x * 100289, calculated
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
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sequence includes a putative N-terminal plastid transit peptide and three aspartate-rich regions
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
homology model of the second active site of enzyme based on the structure of 5-epiaristolochene synthase from Nicotiana tabacum, Protein Data Bank ID code 5EAT
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expression in Arabidopsis thaliana
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expression in Escherichia coli
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expression of green fluorescent fusion protein in tobacco cells
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
A620T
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130% of wild-type productivity. Products are 87% levopimaradiene, 11% abieta-7,13-diene, and 2% sandaracopimaradiene
A727S
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inactive
A729G
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2% of wild-type productivity. Product is 100% sandaracopimaradiene
C618N
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20% of wild-type productivity. Products are 91% levopimaradiene, 9% abieta-7,13-diene, and traces of sandaracopimaradiene
E777A
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inactive
G854T
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140% of wild-type productivity. Products are 79% levopimaradiene, 14% abieta-7,13-diene, and 7% sandaracopimaradiene
I855L
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70% of wild-type productivity. Products are 90% levopimaradiene, 7% abieta-7,13-diene, and 3% sandaracopimaradiene
K723S
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180% of wild-type productivity. Products are 90% levopimaradiene, 7% abieta-7,13-diene, and 3% sandaracopimaradiene
L619F
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150% of wild-type productivity. Products are 94% levopimaradiene, 6% abieta-7,13-diene, and traces of sandaracopimaradiene
L696Q
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160% of wild-type productivity. Products are 86% levopimaradiene, 7% abieta-7,13-diene, and 7% sandaracopimaradiene
M593I
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370% of wild-type productivity. Products are 83% levopimaradiene, 12% abieta-7,13-diene, and 5% sandaracopimaradiene
N769A
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inactive
N838E
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220% of wild-type productivity. Products are 92% levopimaradiene, 6% abieta-7,13-diene, and 2% sandaracopimaradiene
V731L
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8% of wild-type productivity. Product is 100% levopimaradiene
Y700F
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500% of wild-type productivity. Products are 80% levopimaradiene, 15% abieta-7,13-diene, and 5% sandaracopimaradiene
Y700H
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40% of wild-type productivity. Products are 71% levopimaradiene, traces of abieta-7,13-diene, and 29% sandaracopimaradiene
Y700M
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500% of wild-type productivity. Products are 80% levopimaradiene, 13% abieta-7,13-diene, and 7% sandaracopimaradiene
Y700W
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400% of wild-type productivity. Products are 85% levopimaradiene, 7% abieta-7,13-diene, and 8% sandaracopimaradiene
A713S
products are isopimaradiene and sandaracopimaradiene
G651V
no change in product
V717L
no change in product
W679L/Y686H/A713S/V717L
products are isopimaradiene and sandaracopimaradiene
Y686H
no change in product
Y686H/A713S
main products are isopimaradiene and sandaracopimaradiene
Y686H/A713S A713S
main products are isopimaradiene and sandaracopimaradiene
Y686H/A713S/V717L
products are isopimaradiene and sandaracopimaradiene
additional information
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
synthesis
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increase of levopimaradiene synthesis in Escherichia coli by amplification of the flux toward isopentenyl diphosphate and dimethylallyl diphosphate precursors and reprogramming the rate-limiting downstream pathway by generating combinatorial mutations in geranylgeranyl diphosphate synthase and levopimaradiene synthase. The most productive pathway, combining precursor flux amplification and mutant synthases, confers approximately 2600fold increase in levopimaradiene levels. A maximum titer of approximately 700 mg/l is obtained by cultivation in a benchscale bioreactor