Amorpha-4,11-diene synthase: Mechanism and stereochemistry of the enzymatic cyclization of farnesyl diphosphate
(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
- Picaud, S ; Mercke, P ; He, XF ; Sterner, Olov LU ; Brodelius, M ; Cane, DE and Brodelius, P E
- organization
- publishing date
- 2006
- 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
- 2022-03-20 23:01:05
@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}},
keywords = {{GC-MS; proton NMR; COSY; deuterated substrate; enzyme mechanism; sesquiterpenes; 11-diene synthase; amorpha-4}},
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}},
}