Prosser, I., Phillips, A., Gittings, S., Lewis, M., Hooper, A., Pickett, J.  ORCID: https://orcid.org/0000-0002-8386-3770 and Beale, M.
2002.
(+)-(10R)-germacrene A synthase from goldenrod, Solidago canadensis; cDNA isolation, bacterial expression and functional analysis.
Phytochemistry
60
(7)
, pp. 691-702.
10.1016/S0031-9422(02)00165-6
|
Abstract
Profiling of sesquiterpene hydrocarbons in extracts of goldenrod, Solidago canadensis, by GC–MS revealed the presence of both enantiomers of germacrene D and lesser amounts of germacrene A, α-humulene, and β-caryophyllene. A similarity-based cloning strategy using degenerate oligonucleotide primers, based on conserved amino acid sequences in known plant sesquiterpene synthases and RT-PCR, resulted in the isolation of a full length sesquiterpene synthase cDNA. Functional expression of the cDNA in E. coli, as an N-terminal thioredoxin fusion protein using the pET32b vector yielded an enzyme that was readily purified by nickel-chelate affinity chromatography. Chiral GC–MS analysis of products from of 3H- and 2H-labelled farnesyl diphosphate identified the enzyme as (+)-(10R)-germacrene A synthase. Sequence analysis and molecular modelling was used to compare this enzyme with the mechanistically related epi-aristolochene synthase from tobacco.
| Item Type: | Article |
|---|---|
| Date Type: | Publication |
| Status: | Published |
| Schools: | Schools > Chemistry |
| Publisher: | Elsevier |
| ISSN: | 0031-9422 |
| Last Modified: | 05 Mar 2026 15:30 |
| URI: | https://orca.cardiff.ac.uk/id/eprint/185535 |
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