Document Detail

Biochemical conservation and evolution of germacrene A oxidase in asteraceae.
MedLine Citation:
PMID:  20351109     Owner:  NLM     Status:  MEDLINE    
Sesquiterpene lactones are characteristic natural products in Asteraceae, which constitutes approximately 8% of all plant species. Despite their physiological and pharmaceutical importance, the biochemistry and evolution of sesquiterpene lactones remain unexplored. Here we show that germacrene A oxidase (GAO), evolutionarily conserved in all major subfamilies of Asteraceae, catalyzes three consecutive oxidations of germacrene A to yield germacrene A acid. Furthermore, it is also capable of oxidizing non-natural substrate amorphadiene. Co-expression of lettuce GAO with germacrene synthase in engineered yeast synthesized aberrant products, costic acids and ilicic acid, in an acidic condition. However, cultivation in a neutral condition allowed the de novo synthesis of a single novel compound that was identified as germacrene A acid by gas and liquid chromatography and NMR analyses. To trace the evolutionary lineage of GAO in Asteraceae, homologous genes were further isolated from the representative species of three major subfamilies of Asteraceae (sunflower, chicory, and costus from Asteroideae, Cichorioideae, and Carduoideae, respectively) and also from the phylogenetically basal species, Barnadesia spinosa, from Barnadesioideae. The recombinant GAOs from these genes clearly showed germacrene A oxidase activities, suggesting that GAO activity is widely conserved in Asteraceae including the basal lineage. All GAOs could catalyze the three-step oxidation of non-natural substrate amorphadiene to artemisinic acid, whereas amorphadiene oxidase diverged from GAO displayed negligible activity for germacrene A oxidation. The observed amorphadiene oxidase activity in GAOs suggests that the catalytic plasticity is embedded in ancestral GAO enzymes that may contribute to the chemical and catalytic diversity in nature.
Don Trinh Nguyen; Jens Christian Göpfert; Nobuhiro Ikezawa; Gillian Macnevin; Meena Kathiresan; Jürgen Conrad; Otmar Spring; Dae-Kyun Ro
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't     Date:  2010-03-29
Journal Detail:
Title:  The Journal of biological chemistry     Volume:  285     ISSN:  1083-351X     ISO Abbreviation:  J. Biol. Chem.     Publication Date:  2010 May 
Date Detail:
Created Date:  2010-05-24     Completed Date:  2010-06-17     Revised Date:  2011-07-28    
Medline Journal Info:
Nlm Unique ID:  2985121R     Medline TA:  J Biol Chem     Country:  United States    
Other Details:
Languages:  eng     Pagination:  16588-98     Citation Subset:  IM    
Department of Biological Sciences, University of Calgary, Calgary T2N 1N4, Canada.
Data Bank Information
Bank Name/Acc. No.:
GENBANK/GU198171;  GU256644;  GU256645;  GU256646;  GU256647
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MeSH Terms
Asteraceae / enzymology*
Chromatography, Thin Layer / methods
Cytochrome P-450 Enzyme System / chemistry
Evolution, Molecular*
Gene Expression Regulation
Genetic Variation
Magnetic Resonance Spectroscopy
Mass Spectrometry / methods
Microsomes / metabolism
Molecular Sequence Data
Oxidoreductases / chemistry*,  genetics
Plant Proteins / metabolism
Recombinant Proteins / chemistry
Sesquiterpenes, Germacrane / chemistry*,  genetics
Reg. No./Substance:
0/Plant Proteins; 0/Recombinant Proteins; 0/Sesquiterpenes, Germacrane; 28387-44-2/germacrene A; 9035-51-2/Cytochrome P-450 Enzyme System; EC 1.-/Oxidoreductases

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine

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