Information on EC 4.2.3.77 - (+)-germacrene D synthase

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The enzyme appears in viruses and cellular organisms

EC NUMBER
COMMENTARY hide
4.2.3.77
-
RECOMMENDED NAME
GeneOntology No.
(+)-germacrene D synthase
-
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
(2E,6E)-farnesyl diphosphate = (+)-germacrene D + diphosphate
show the reaction diagram
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
germacrene biosynthesis
-
-
SYSTEMATIC NAME
IUBMB Comments
2E,6E)-farnesyl-diphosphate diphosphate-lyase [(+)-germacrene-D-forming]
Requires Mg2+, Mn2+, Ni2+ or Co2+. The formation of (+)-germacrene D involves a 1,2-hydride shift whereas for (-)-germacrene D there is a 1,3-hydride shift (see EC 4.2.3.75).
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
-
UniProt
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
(+)-germacrene D + diphosphate
show the reaction diagram
additional information
?
-
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Co2+
-
1 mM, 33% of the activity with Mg2+
Ni2+
-
0.5 mM, 6% of the activity with Mg2+
additional information
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0025 - 0.379
(2E,6E)-farnesyl diphosphate
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.017 - 0.04
(2E,6E)-farnesyl diphosphate
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
6.7
(2E,6E)-farnesyl diphosphate
81
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6.5 - 9
-
almost completely inactive below and above
pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5.6
-
calculated
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
expression is significantly higher in flowers than in leaf tissue. Expression increases at 08.00 h and then drops to about a half by 20.00 h
Manually annotated by BRENDA team
expression is significantly higher in flowers than in leaf tissue
Manually annotated by BRENDA team
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
60000
-
x * 63800, calculated, x * 60000, SDS-PAGE
63800
-
x * 63800, calculated, x * 60000, SDS-PAGE
64802
x * 64802, calculated
65200
x * 65200, calculated
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
recombinant protein
-
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
Agrobacterium tumefaciens-mediated expression in Nicotiana benthamiana
expression in Escherichia coli
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
N303D
mimicking of corresponding amino acid in (-)-germacrene D synthase Sc19. Kinetic data similar to wild-type
N303E
less than 0.25% of wild-type activity
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
analysis
use of NMR spectroscopy as a tool to analyze the kinetics of enzyme reactions using progress curves. The protocol presented is also a simple and direct approach for the measurement of enzyme kinetics in the presence of synthetic inhibitors. The conversion of farnesyl diphosphate into (+)-germacrene D by the enzyme germacrene D synthase involves an amphiphilic substrate forming micelles and a water insoluble product. Using proper controls, the conversion can well be analyzed by the progress curve approach using the Lambert W function
synthesis
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improved conditions for expression of Zingiber officinale (+)-germacrene synthase in Escherichia coli. Comparison of bacterial strains; BL21 (DE3), BL21 (DE3) Tuner BL21(DE3) pLysS and BL21 (DE3) pLysS Tuner using different inducing agents. The change between BL21 (DE3) cells and BL21 (DE3) Tuner, induced with IPTG, leads to a twofold increase in enzyme activity in the soluble fraction while a reduction in activity is observed when using the pLysS strains. The same doubling of activity is observed for germacrene synthase when the commonly used BL.21 (DE3) is induced with The Inducer. Addition of 2.5 mM glycine betaine and 660 mM sorbitol to the bacterial growth media results in reduction of growth rate and biomass yield but under these conditions the best overall protein production is obtained