Information on EC 4.2.3.129 - (+)-sativene synthase

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The expected taxonomic range for this enzyme is: Eukaryota

EC NUMBER
COMMENTARY hide
4.2.3.129
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RECOMMENDED NAME
GeneOntology No.
(+)-sativene synthase
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REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
(2E,6E)-farnesyl diphosphate = (+)-sativene + diphosphate
show the reaction diagram
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SYSTEMATIC NAME
IUBMB Comments
(2E,6E)-farnesyl-diphosphate diphosphate-lyase (cyclizing, (+)-sativene-forming)
Isolated from the fungus Coprinus cinereus. The enzyme also forms (+)-delta-cadinene, beta-copaene, beta-cubebene, and traces of several other sequiterpenoids. See EC 4.2.3.13, (+)-delta-cadinene synthase, EC 4.2.3.127, beta-copaene synthase, and EC 4.2.3.128, beta-cubebene synthase.
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
gene cop4
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Manually annotated by BRENDA team
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UniProt
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
evolution
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the enzyme belongs to the sesquiterpene synthases
malfunction
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identification of constitutive photomorphogenic mutants, cop2, cop3, and cop4,in which dark-grown seedlings have open and enlarged cotyledons resembling those of light-grown wild-type seedlings. Mutations in each of the three loci alleviate the normal inhibition of cell-type differentiation, cell enlargement, and lateral cell division observed in cotyledons of dark-grown wild-type seedlings, but do not affect plastid differentiation. Cop4 mutation also leads to high-leve1 dark expression of nuclear, but not plastid-encoded, light-inducible genes
metabolism
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epistatic relationships of these three mutations to previously characterized phytochrome-deficient mutations suggest that COP2, COP3, and COP4 may act downstream of phytochrome in the light regulatory pathway
physiological function
additional information
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
(2E,6E)-farnesyl diphosphate
(+)-sativene + diphosphate
show the reaction diagram
(2Z,6E)-farnesyl diphosphate
(+)-sativene + diphosphate
show the reaction diagram
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conversion of (E,E)-farnesyl diphosphate proceeds via a (6S)-beta-bisabolene carbocation in the case of Cop4
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?
additional information
?
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Cop4 cultures produce several sesquiterpene compounds, e.g. beta-cubebene, sativene, beta-copaene, and cubebol
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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
(2E,6E)-farnesyl diphosphate
(+)-sativene + diphosphate
show the reaction diagram
additional information
?
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A8NU13
Cop4 cultures produce several sesquiterpene compounds, e.g. beta-cubebene, sativene, beta-copaene, and cubebol
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METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Mn2+
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substitution of Mg2+ with Mn2+ as the divalent metal ion shifts the product profile of Cop4 to germacrene D, disfavoring subsequent ring closures that produce the cadinyl cation and its tricyclic descendents
NaCl
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does not affect the product specificity of Cop4 significantly at 1 M
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
additional information
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increasing the reaction temperature to 37°C decreases the fidelity of Cop4. At this temperature Cop4 generates a relative larger fraction of products beta-cubebene, sativene, delta-cadinene and beta-copaene, that are derived from a cadinyl cation intermediate
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.011 - 0.0707
(2E,6E)-farnesyl diphosphate
additional information
additional information
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kinetic analysis, overview
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pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
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changing the pH of the reaction drastically alters the fidelity of Cop4 and makes it a highly selective enzyme. Increasing the reaction temperature to 37°C decreases the fidelity of Cop4. At this temperature Cop4 generates a relative larger fraction of products beta-cubebene, sativene, delta-cadinene and beta-copaene, that are derived from a cadinyl cation intermediate. The histidine side chain in the Cop4 loop, in particular, has a strong impact on the net charge of the loop at different pH values
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
25 - 30
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assay at
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expression of wild-type and mutant enzymes in Escherichia coli strain JM109
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gene cop4, DNA and amino acid sequence determination and analysis, phylogenetic analysis, recombinant expression in Escherichia coli strain JM109
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
H235P
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site-directed mutagenesis, no production of sativene, the mutation converts Cop4 into a much more selective enzyme that produces (-)-germacrene D as the major cyclization product with 50% of total sesquiterpenes products. The mutant makes beta-ylangene, which is a diastereomer of beta-copaene and not synthesized by wild-type Cop4
K233I
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site-directed mutagenesis, mutation of K233, interacting with the second Asp92 in the DDXXD motif of Cop4, does not significantly change the overall product promiscuity of Cop4, though beta-cubebene 4 does become the major product and püroduction of sativene is reduced by 50% compared to the wild-type enzyme
N238L
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site-directed mutagenesis, the mutant shows a altered product profile compared to the wild-type enzyme with only slight reduction in beta-cubebene synthesis, sativene levels are similar to the wild-type. The mutant does no longer show production of cubebol and has reduced (-)-germacrene D synthesis activity compared to the wild-type enzyme, synthesis of beta-cubebene, beta-copaene, delta-cadinene, and alpha-cubebene
N239L
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site-directed mutagenesis, no production of sativene, the mutation converts Cop4 into a much more selective enzyme that produces (-)-germacrene D as the major cyclization product with 50% of total sesquiterpenes products. The mutant makes beta-ylangene, which is a diastereomer of beta-copaene and not synthesized by wild-type Cop4
T236L
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site-directed mutagenesis, the mutant shows a altered product profile compared to the wild-type enzyme with an increase in sativene synthesis. The mutant does no longer show production of cubebol and (-)-germacrene D compared to the wild-type enzyme
additional information