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Identification and functional characterization of a PDR transporter in Tripterygium wilfordii Hook.f. that mediates the efflux of triptolide
Key Message TwPDR1, a PDR transporter from Tripterygium wilfordii Hook.f., was proved to efflux triptolide and its stability could be enhanced by A 1033 T mutation. Triptolide, an abietane-type diterpene in Tripterygium wilfordii Hook.f., possesses many pharmacological activities. However, triptolid...
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Published in: | Plant molecular biology 2021-05, Vol.106 (1-2), p.145-156 |
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Main Authors: | , , , , |
Format: | Article |
Language: | English |
Subjects: | |
Citations: | Items that this one cites Items that cite this one |
Online Access: | Get full text |
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Summary: | Key Message
TwPDR1, a PDR transporter from
Tripterygium wilfordii
Hook.f., was proved to efflux triptolide and its stability could be enhanced by A
1033
T mutation.
Triptolide, an abietane-type diterpene in
Tripterygium wilfordii
Hook.f., possesses many pharmacological activities. However, triptolide is in short supply and very expensive because it is present at low amounts in natural plants and lack alternative production methods. Transporter engineering, which increases the extracellular secretion of secondary metabolites in in vitro culture systems, is an effective strategy in metabolic engineering but is rarely reported. In this study, TwPDR1, a pleiotropic drug resistance-type ATP binding cassette transporter, was identified as the best efflux pump candidate for diterpenoids through bioinformatics analysis. TwPDR1 was located in the plasma membrane, highly expressed in adventitious roots, and induced by methyl jasmonate. The triptolide efflux function of TwPDR1 was confirmed by transient expression in tobacco BY-2 cells and by downregulation via RNA interference in the native host. However, the overexpression of
TwPDR1
had a limited effect on the secretion of triptolide. As shown by previous studies, a single amino acid mutation might increase the abundance of TwPDR1 by increasing protein stability. We identified the A
1033
residue in TwPDR1 by sequence alignment and confirmed that A
1033
T mutation could increase the expression of TwPDR1 and result in the higher release ratio of triptolide (78.8%) of the mutants than that of control (60.1%). The identification and functional characterization of TwPDR1 will not only provide candidate gene material for the metabolic engineering of triptolide but also guide other transporter engineering researches in the future. |
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ISSN: | 0167-4412 1573-5028 |
DOI: | 10.1007/s11103-021-01134-y |