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4.2.3.24: amorpha-4,11-diene synthase

This is an abbreviated version!
For detailed information about amorpha-4,11-diene synthase, go to the full flat file.

Word Map on EC 4.2.3.24

Reaction

(2E,6E)-farnesyl diphosphate
=
amorpha-4,11-diene
 +
diphosphate

Synonyms

ADS, ADS1, ADS3963, amorpha-4,11-diene synthase, amorphadiene synthase, AMS

ECTree

     4 Lyases
         4.2 Carbon-oxygen lyases
             4.2.3 Acting on phosphates
                4.2.3.24 amorpha-4,11-diene synthase

Cloned

Cloned on EC 4.2.3.24 - amorpha-4,11-diene synthase

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CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
Escherichia coli DH10B is used for cloning and strain DH1 is used for expression, developing and optimizing of a simple cloning system for expression of the amorphadiene biosynthetic pathway. The pathway genes are originally constructed by assembly of the operons onto three different plasmids, each with a different copy number and Escherichia coli promoter. Escherichia coli DH1 harboring pAM45MevT and pADS exhibits a significant increase in amorphadiene production (21 mg/l at 75 h), which is 3fold higher than the original base-strain DH1 pMevTpMBISpADS. To increase ADS expression, transforming of DH1pAM92 with pADS is done, resulting in a combined ADS gene dosage of 30-45 copies/cell. The strain exhibits an increase in amorphadiene production over DH1pAM92, but still does not approach the peak titers obtained with DH1pAM45pADS
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expressed in Escherichia coli BL21(DE3) cells
expressed in Escherichia coli BL21-CodonPlus (DE3)-RIL cells
expressed in Escherichia coli Rosetta(DE3) cells
expression in Escherichia coli
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expression in Escherichia coli with His-tag
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expression of a synthetic Artemesia annua amorphadiene synthase gene under control of a strong constitutive promoter ((p)gpdA) in Aspergillus nidulans yields altered product distribution. The transformants produces only small amounts of amorphadiene, but much larger amounts of similar sesquiterpenes normally produced as minor by-products in plants. Expression of ADS in Escherichia coli produces almost exclusively amorpha-4,11-diene. The host environment can greatly impact the terpenes produced from terpene synthases.
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into the pET28 vector for expression in Escherichia coli BL21DE3, BL21DE3 Tuner, BL21DE3 pLysS and BL21DE3 pLysS Tuner cells using different inducing agents
into the pET30a+ vector for expression in Escherichia coli
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into the pYeDP60 vector for expression in Saccharomyces cerevisiae, and, in addition, introduced into the yeast genome by homologous recombination
The ADS gene is mutated to the yeast-conform variant ADSm (based on the original DNA sequence of the wild-type gene, the plant gene codons are modified to Saccharomyces cerevisiae preferred with the intact protein primary structure, production in a transformed yeast.) The 8 low-usage codons, AGG (Arg), CGA (Arg), CGG (Arg), CGC (Arg), CCG (Pro), CTC (Leu), GCG (Ala) and ACG (Thr) in ADSm gene are changed with bias codons in Saccharomyces cerevisiae. The synonymous codons of 5 amino acids are changed into one preferred codon in ADSm gene, such as Gln (CAA), Glu (GAA), Cys (TGT), Arg (AGA) and Gly (GGT), which can improve the abundance of cognate tRNA when translated. Three genes, encoding the FPP synthase, HMG-CoA reductase and ADS, are plasmid-transformed together.
the open reading frame of amorpha-4,11-diene synthase is fused to the green fluorescent proten reporter gene for introducing into the Arabidopsis protoplasts
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