Information on EC 4.2.3.23 - germacrene-A synthase

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The expected taxonomic range for this enzyme is: Eukaryota, Bacteria

EC NUMBER
COMMENTARY hide
4.2.3.23
-
RECOMMENDED NAME
GeneOntology No.
germacrene-A synthase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
(2E,6E)-farnesyl diphosphate = (+)-(R)-gemacrene A + diphosphate
show the reaction diagram
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
Biosynthesis of secondary metabolites
-
-
costunolide biosynthesis
-
-
delta-guaiene biosynthesis
-
-
germacrene biosynthesis
-
-
Sesquiterpenoid and triterpenoid biosynthesis
-
-
SYSTEMATIC NAME
IUBMB Comments
(2E,6E)-farnesyl-diphosphate diphosphate-lyase [(1E,4S,5E,7R)-germacra-1(10),5-dien-11-ol-forming]
Requires Mg2+ for activity. While germacrene A is an enzyme-bound intermediate in the biosynthesis of a number of phytoalexins, e.g. EC 4.2.3.9 (aristolochene synthase) from some species and EC 4.2.3.21 (vetispiradiene synthase), it is the sole sesquiterpenoid product formed in chicory [1].
CAS REGISTRY NUMBER
COMMENTARY hide
213763-55-4
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
-
SwissProt
Manually annotated by BRENDA team
-
UniProt
Manually annotated by BRENDA team
strain PCC 7120, previously Anabaena sp.
-
-
Manually annotated by BRENDA team
-
SwissProt
Manually annotated by BRENDA team
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
(+)-(R)-germacrene A + diphosphate
show the reaction diagram
(2E,6E)-farnesyl diphosphate
germacrene A + diphosphate
show the reaction diagram
2-trans,6-trans-farnesyl diphosphate
(+)-(10R)-germacrene A + diphosphate
show the reaction diagram
additional information
?
-
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
2-trans,6-trans-farnesyl diphosphate
(+)-(10R)-germacrene A + diphosphate
show the reaction diagram
additional information
?
-
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
NADPH
-
assay at
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
farnesyl diphosphate
substrate inhibition above 0.0025 mM
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
Tween 20
-
at 0.1%, activates the enzyme by 5fold
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.00074 - 0.00106
(2E,6E)-farnesyl diphosphate
0.0025 - 14.9
2-trans,6-trans-farnesyl diphosphate
additional information
additional information
-
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.00035 - 4.4
2-trans,6-trans-farnesyl diphosphate
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6.24
-
purified enzyme
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6.7
-
about, broad optimum
7.5 - 8
-
recombinant enzyme
8
-
assay at
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5.1 - 7.3
-
half maximal activities at pH 5.1 and pH 7.3
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
37
-
recombinant enzyme
pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5.2
-
DNA sequence calculation
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
normal and etiolated; normal and etiolated
Manually annotated by BRENDA team
head core tissue
Manually annotated by BRENDA team
especially in outer tissue, highest expression level of the long isozyme; highest expression level of the short isozyme in outer and inner tissues
Manually annotated by BRENDA team
additional information
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
54000
-
gel filtration
56000
-
1 * 56000, about, SDS-PAGE
64200
x * 64200, calculated
64400
x * 64400, calculated
64700
x * 64700, about, DNA sequence calculation
67100
-
x * 67100, DNA sequence calculation
80000
x * 64500, calculated. x * 80000, SDS-PAGE of recombinant N-terminal thioredoxin fusion protein
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
monomer
-
1 * 56000, about, SDS-PAGE
additional information
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
glycerol stabilizes the enzyme
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-80°C, partially purified root enzyme, in 15 mM MOPS, pH 7.0, 10 mM MgCl2, 2 mM DTT, 30% v/v glycerol, stable for several months
-
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
201fold from roots by anion exchange and dye-ligand affinity chromatography
-
cell debris is cleared by centrifugation and the cleared protein extract is used to purify proteins by metal-affinity chromatography
-
Ni-NTA affinity chromatography; Ni-NTA affinity chromatography
partially, recombinant enzyme
-
recombinant long isozyme from Escherichia coli, native isozyme partially from chicory heads by anion exchange chromatography; recombinant short isozyme from Escherichia coli, native isozyme partially from chicory heads by anion exchange chromatography
recombinant thioredoxin fusion enzyme from Escherichia coli by nickel chelate affinity chromatography
recombinant wild-type and mutant enzymes from bacteria
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
cloning from a constructed glandular trichome cDNA library, DNA and amino acid sequence determination and analysis, phylogenetic analysis, functional expression in Escherichia coli strain BL21-DE3-RIL
DNA and amino acid sequence determination and analysis, phylogenetic analysis, expression in Escherichia coli; DNA and amino acid sequence determination and analysis, phylogenetic analysis, expression in Escherichia coli
DNA library construction, phage display, DNA and amino acid sequence determination and analysis, functional expression in Escherichia coli as N-terminal thioredoxin fusion protein
DNA sequence determination and analysis
-
expressed in Escherichia coli
-
expression in Escherichia coli
expression in Escherichia coli, N-terminal thioredoxin fusion protein
expression of wild-type and mutant enzymes in bacteria
NS1 is overexpressed at high levels in Escherichia coli. The gene encoding NS1 is flanked by an ORF encoding a putative cytochrome P450 monooxygenase
-
thioredoxin-fusion protein expressed in Escherichia coli; thioredoxin-fusion protein expressed in Escherichia coli
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
A435T
site-directed mutagenesis, altered kinetics but similar activity compared to the wild-type enzyme
S434A
site-directed mutagenesis, activity and kinetics similar to the wild-type enzyme
S434T/A435T
site-directed mutagenesis, inactive mutant
T433A
site-directed mutagenesis, altered kinetics and reduced activity compared to the wild-type enzyme
T433A/A435T
site-directed mutagenesis, altered kinetics and reduced activity compared to the wild-type enzyme
T433A/S434A
site-directed mutagenesis, inactive mutant
T433A/S434A/A435T
site-directed mutagenesis, inactive mutant
T433A/S434T
site-directed mutagenesis, inactive mutant
additional information
-
deletion of the N-terminal 44 amino acid residues leads to loss of 90% of catalytic activity
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
analysis