Literature summary for 4.2.3.89 extracted from

Salvagnin, U.; Carlin, S.; Angeli, S.; Vrhovsek, U.; Anfora, G.; Malnoy, M.; Martens, S.
Homologous and heterologous expression of grapevine E-(beta)-caryophyllene synthase (VvGwECar2) (2016), Phytochemistry, 131, 76-83 .

Cloned(Commentary)

Cloned (Comment)	Organism
gene VvGwECar2, recombinant overexpression in Escherichia coli strain BL21-CodonPlus(DE3)-RIPL, and transgenic recombinant overexpression under the control of cauliflower mosaic virus (CaMV) 35S promoter n the cytoplasm of Arabidopsis thaliana ecotype Col-0 and Vitis vinifera. In Arabidopsis thaliana, the stable enzyme expression changes the plant head space release, showing a higher selectivity for E-(beta)-caryophyllene, but also the production of thujopsene instead of germacrene D. Overall, plants increase the E-(beta)-caryophyllene emission in the headspace collection by 8fold compared to Col-0 control plants. In grapevine VvGwECar2 overexpression result in higher E-(beta)-caryophyllene emissions. Quantitative RT-PCR expression analysis	Vitis vinifera

Cloned (Comment)

Organism

gene VvGwECar2, recombinant overexpression in Escherichia coli strain BL21-CodonPlus(DE3)-RIPL, and transgenic recombinant overexpression under the control of cauliflower mosaic virus (CaMV) 35S promoter n the cytoplasm of Arabidopsis thaliana ecotype Col-0 and Vitis vinifera. In Arabidopsis thaliana, the stable enzyme expression changes the plant head space release, showing a higher selectivity for E-(beta)-caryophyllene, but also the production of thujopsene instead of germacrene D. Overall, plants increase the E-(beta)-caryophyllene emission in the headspace collection by 8fold compared to Col-0 control plants. In grapevine VvGwECar2 overexpression result in higher E-(beta)-caryophyllene emissions. Quantitative RT-PCR expression analysis

Vitis vinifera

Protein Variants

Protein Variants	Comment	Organism
additional information	the enzyme from transgenic Arabidopsis thalina plants shows, in contrast with the behavior of the enzyme in vitro, no trace of germacrene D production. A low thujopsene production, also a higher selectivity for E-(beta)-caryophyllene production: the headspace of the modified plants is indeed dominated by alpha-humulene, thujopsene and E-(beta)-caryophyllene, in a 1:0.75:5 mass ratio (average). But a great variation in sesquiterpenes quantity among the different lines is observed, probably because of the difference in T-DNA copy number and expression. Overall, plants increase the E-(beta)-caryophyllene emission in the headspace collection by 8fold compared to Col-0 control plants. In grapevine VvGwECar2 overexpression result in higher E-(beta)-caryophyllene emissions. No differences in plant phenotypes or growth speed are observed	Vitis vinifera

KM Value [mM]

KM Value [mM]	KM Value Maximum [mM]	Substrate	Comment	Organism	Structure
0.0314	-	(2E,6E)-farnesyl diphosphate	recombinant enzyme, pH 7.0, 30°C	Vitis vinifera

Localization

Localization	Comment	Organism	GeneOntology No.	Textmining
cytoplasm	-	Vitis vinifera	5737	-

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
(2E,6E)-farnesyl diphosphate	Vitis vinifera	-	(+)-beta-caryophyllene + diphosphate	-	?

Organism

Organism	UniProt	Comment	Textmining
Vitis vinifera	-	-	-

Source Tissue

Source Tissue	Comment	Organism	Textmining
berry	-	Vitis vinifera	-

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.	Reac.
(2E,6E)-farnesyl diphosphate	-	Vitis vinifera	(+)-beta-caryophyllene + diphosphate	-	?
additional information	enzyme VGwECar2 catalyzes the cyclization of farnesyl diphosphate to both beta-caryophyllene and alpha-humulene, plus a small amount of germacrene D, GC-MS analysis	Vitis vinifera	?	-	?

Synonyms

Synonyms	Comment	Organism
E-(beta)-caryophyllene synthase	-	Vitis vinifera
VvGwECar2	-	Vitis vinifera

Temperature Optimum [°C]

Temperature Optimum [°C]	Temperature Optimum Maximum [°C]	Comment	Organism
30	-	assay at	Vitis vinifera

Turnover Number [1/s]

Turnover Number Minimum [1/s]	Turnover Number Maximum [1/s]	Substrate	Comment	Organism	Structure
0.19	-	(2E,6E)-farnesyl diphosphate	recombinant enzyme, pH 7.0, 30°C	Vitis vinifera

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
7.8	-	recombinant enzyme	Vitis vinifera

General Information

General Information	Comment	Organism
evolution	in grapevine five genes are known to code for beta-caryophyllene synthases, but only one of them, VvGwECar2, is actually expressed in all plant tissues and therefore accounts for most of the volatile production in vegetative parts and berries, although in flowers two other genes, VvGwECar1 and VvPNECar2, seem to play a big role	Vitis vinifera
physiological function	the TPS enzyme (VvGwECar2) is responsible for most of the production of E-(beta)-caryophyllene in grapevine (VvGwECar2). It might play a key role in the plant regulation machinery	Vitis vinifera

kcat/KM [mM/s]

kcat/KM Value [1/mMs^-1]	kcat/KM Value Maximum [1/mMs^-1]	Substrate	Comment	Organism	Structure
6.1	-	(2E,6E)-farnesyl diphosphate	recombinant enzyme, pH 7.0, 30°C	Vitis vinifera