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Amorpha-4,11-diene synthase: Mechanism and stereochemistry of the enzymatic cyclization of farnesyl diphosphate

Picaud, S ; Mercke, P ; He, XF ; Sterner, Olov LU ; Brodelius, M ; Cane, DE and Brodelius, P E (2006) In Archives of Biochemistry and Biophysics 448(1-2). p.150-155
Abstract
Recombinant amorpha-4,11-diene synthase from Artemisia annua, expressed in Escherichia coli, was incubated with the deuterium-labeled farnesyl diphosphates, (1R)-[1-H-2]FPP, (1S)-[1-H-2]FPP, and [1,1-H-2(2)]FPP. GC-MS analysis of amorpha-4,11-diene formed from the deuterated FPPs shows that the deuterium atoms are retained in the product. Furthermore, analysis of the MS-spectra obtained with the differently labeled substrate indicates that the H-1si-proton of FPP is transferred during the cyclization reaction to carbon 10 of amorphadiene while the H-1re-proton of FPP is retained on C-6 of the product. Proton NMR and COSY experiments proved that the original H-1si-proton of FPP is located at C-10 of amorpha-4,11-diene as a result of a... (More)
Recombinant amorpha-4,11-diene synthase from Artemisia annua, expressed in Escherichia coli, was incubated with the deuterium-labeled farnesyl diphosphates, (1R)-[1-H-2]FPP, (1S)-[1-H-2]FPP, and [1,1-H-2(2)]FPP. GC-MS analysis of amorpha-4,11-diene formed from the deuterated FPPs shows that the deuterium atoms are retained in the product. Furthermore, analysis of the MS-spectra obtained with the differently labeled substrate indicates that the H-1si-proton of FPP is transferred during the cyclization reaction to carbon 10 of amorphadiene while the H-1re-proton of FPP is retained on C-6 of the product. Proton NMR and COSY experiments proved that the original H-1si-proton of FPP is located at C-10 of amorpha-4,11-diene as a result of a 1,3-hydride shift following initial 1,6-ring closure. The results obtained support the previously suggested mechanism for the cyclization of farnesyl diphosphate by amorph-4,11-diene synthase involving isomerization of FPP to (R)-nerolidyl diphosphate (NPP), ionization of NPP, and C-1,C-6-ring closure to generate a bisabolyl cation, followed by a 1,3-hydride shift, 1,10-ring closure to generate the amorphane skeleton, and deprotonation at either C-12 or C-13 to afford the final product (1S,6R,7R,10R)-amorpha-4,11-diene. (c) 2005 Elsevier Inc. All rights reserved. (Less)
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author
; ; ; ; ; and
organization
publishing date
type
Contribution to journal
publication status
published
subject
keywords
GC-MS, proton NMR, COSY, deuterated substrate, enzyme mechanism, sesquiterpenes, 11-diene synthase, amorpha-4
in
Archives of Biochemistry and Biophysics
volume
448
issue
1-2
pages
150 - 155
publisher
Academic Press
external identifiers
  • pmid:16143293
  • wos:000237493100016
  • scopus:33646148490
  • pmid:16143293
ISSN
0003-9861
DOI
10.1016/j.abb.2005.07.015
language
English
LU publication?
yes
additional info
The information about affiliations in this record was updated in December 2015. The record was previously connected to the following departments: Organic chemistry (S/LTH) (011001240)
id
f7e791e9-e933-4ddb-bbc1-8dddc382d3b3 (old id 409810)
date added to LUP
2016-04-01 12:04:01
date last changed
2021-03-24 02:08:59
@article{f7e791e9-e933-4ddb-bbc1-8dddc382d3b3,
  abstract     = {Recombinant amorpha-4,11-diene synthase from Artemisia annua, expressed in Escherichia coli, was incubated with the deuterium-labeled farnesyl diphosphates, (1R)-[1-H-2]FPP, (1S)-[1-H-2]FPP, and [1,1-H-2(2)]FPP. GC-MS analysis of amorpha-4,11-diene formed from the deuterated FPPs shows that the deuterium atoms are retained in the product. Furthermore, analysis of the MS-spectra obtained with the differently labeled substrate indicates that the H-1si-proton of FPP is transferred during the cyclization reaction to carbon 10 of amorphadiene while the H-1re-proton of FPP is retained on C-6 of the product. Proton NMR and COSY experiments proved that the original H-1si-proton of FPP is located at C-10 of amorpha-4,11-diene as a result of a 1,3-hydride shift following initial 1,6-ring closure. The results obtained support the previously suggested mechanism for the cyclization of farnesyl diphosphate by amorph-4,11-diene synthase involving isomerization of FPP to (R)-nerolidyl diphosphate (NPP), ionization of NPP, and C-1,C-6-ring closure to generate a bisabolyl cation, followed by a 1,3-hydride shift, 1,10-ring closure to generate the amorphane skeleton, and deprotonation at either C-12 or C-13 to afford the final product (1S,6R,7R,10R)-amorpha-4,11-diene. (c) 2005 Elsevier Inc. All rights reserved.},
  author       = {Picaud, S and Mercke, P and He, XF and Sterner, Olov and Brodelius, M and Cane, DE and Brodelius, P E},
  issn         = {0003-9861},
  language     = {eng},
  number       = {1-2},
  pages        = {150--155},
  publisher    = {Academic Press},
  series       = {Archives of Biochemistry and Biophysics},
  title        = {Amorpha-4,11-diene synthase: Mechanism and stereochemistry of the enzymatic cyclization of farnesyl diphosphate},
  url          = {http://dx.doi.org/10.1016/j.abb.2005.07.015},
  doi          = {10.1016/j.abb.2005.07.015},
  volume       = {448},
  year         = {2006},
}