Cyclization Mechanism of Amorpha-4,11-diene Synthase, a Key Enzyme in Artemisinin Biosynthesis

Cyclization of farnesyl diphosphate into amorpha-4,11-diene by amorpha-4,11-diene synthase (ADS) initiates biosynthesis of artemisinin, a clinically important antimalarial drug precursor. Three possible ring-closure mechanisms, two involving a bisabolyl carbocation intermediate followed by either a...

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Bibliographic Details
Published in:Journal of natural products (Washington, D.C.) D.C.), 2006-05, Vol.69 (5), p.758-762
Main Authors: Kim, Soon-Hee, Heo, Keon, Chang, Yung-Jin, Park, Si-Hyung, Rhee, Sang-Ki, Kim, Soo-Un
Format: Article
Language:English
Subjects:
alkyl (aryl) transferases
Alkyl and Aryl Transferases - metabolism
amorpha-4,11-diene synthase
Artemisia - enzymology
Artemisia annua
artemisinin
Artemisinins - metabolism
Bioconversions. Hemisynthesis
Biological and medical sciences
biosynthesis
Biotechnology
Cyclization
Deuterium - chemistry
Deuterium - metabolism
enzyme activity
Fundamental and applied biological sciences. Psychology
Methods. Procedures. Technologies
Molecular Structure
Nuclear Magnetic Resonance, Biomolecular
Plants, Medicinal - enzymology
Polyisoprenyl Phosphates - chemical synthesis
sesquiterpene cyclase
Sesquiterpenes - metabolism
spectral analysis
Stereoisomerism
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Summary:Cyclization of farnesyl diphosphate into amorpha-4,11-diene by amorpha-4,11-diene synthase (ADS) initiates biosynthesis of artemisinin, a clinically important antimalarial drug precursor. Three possible ring-closure mechanisms, two involving a bisabolyl carbocation intermediate followed by either a 1,3-hydride shift or two successive 1,2-shifts, and one involving a germacrenyl carbocation, were proposed and tested by analyzing the fate of farnesyl diphosphate H-1 hydrogen atoms through 1H and 2H NMR spectroscopy. Migration of one deuterium atom of [1,1-2H2]farnesyl diphosphate to H-10 of amorpha-4,11-diene singled out the bisabolyl carbocation mechanism with a 1,3-hydride shift. Further confirmation was obtained through enzyme reactions with (1R)- and (1S)-[1-2H]farnesyl diphosphate. Results showed that deuterium of the 1R compound remained at H-6, whereas that of the 1S compound migrated to H-10 of amorpha-4,11-diene. Incorporation of one deuterium into amorphadiene in the cyclization process was observed when the reaction was performed in 2H2O, as evidenced by an increase of 1 amu in the mass of the molecular ion.
ISSN:0163-3864
1520-6025
DOI:10.1021/np050356u