The T296V Mutant of Amorpha-4,11-diene Synthase Is Defective in Allylic Diphosphate Isomerization but Retains the Ability To Cyclize the Intermediate (3R)-Nerolidyl Diphosphate to Amorpha-4,11-diene.
Biochemistry
; 55(48): 6599-6604, 2016 Dec 06.
Article
em En
| MEDLINE
| ID: mdl-27933789
ABSTRACT
The T296V mutant of amorpha-4,11-diene synthase catalyzes the abortive conversion of the natural substrate (E,E)-farnesyl diphosphate mainly into the acyclic product (E)-ß-farnesene (88%) instead of the natural bicyclic sesquiterpene amorphadiene (7%). Incubation of the T296V mutant with (3R,6E)-nerolidyl diphosphate resulted in cyclization to amorphadiene. Analysis of additional mutants of amino acid residue 296 and in vitro assays with the intermediate analogue (2Z,6E)-farnesyl diphosphate as well as (3S,6E)-nerolidyl diphosphate demonstrated that the T296V mutant can no longer catalyze the allylic rearrangement of farnesyl diphosphate to the normal intermediate (3R,6E)-nerolidyl diphosphate, while retaining the ability to cyclize (3R,6E)-nerolidyl diphosphate to amorphadiene. The T296A mutant predominantly retained amorphadiene synthase activity, indicating that neither the hydroxyl nor the methyl group of the Thr296 side chain is required for cyclase activity.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Proteínas de Plantas
/
Fosfatos de Poli-Isoprenil
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Sesquiterpenos
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Difosfatos
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Alquil e Aril Transferases
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Mutação de Sentido Incorreto
Idioma:
En
Revista:
Biochemistry
Ano de publicação:
2016
Tipo de documento:
Article