Engineering tropane biosynthetic pathway in Hyoscyamus niger hairy root cultures

被引:175
|
作者
Zhang, L
Ding, RX
Chai, YR
Bonfill, M
Moyano, E
Oksman-Caldentey, KM
Xu, TF
Pi, Y
Wang, ZN
Zhang, HM
Kai, GY
Liao, ZH
Sun, XF
Tang, KX [1 ]
机构
[1] Fudan Univ, Sch Life Sci, Morgan Tan Int Ctr Life Sci, State Key Lab Genet Engn, Shanghai 200433, Peoples R China
[2] Second Mil Med Univ, Sch Pharm, Shanghai 200433, Peoples R China
[3] Univ Barcelona, Fac Farm, Unidad Fisiol Vegetal, E-08028 Barcelona, Spain
[4] Univ Pompeu Fabra, Dept Ciencias Expt Salut, E-08003 Barcelona, Spain
[5] VTT Biotechnol, FIN-02044 Espoo, Finland
[6] SW Agr Univ, Sch Agron & Life Sci, Chongqing 400716, Peoples R China
[7] Shanghai Jiao Tong Univ, Sch Agr & Biol, Plant Biotechnol Res Ctr, Shanghai 200030, Peoples R China
关键词
Agrobacterium; hyoscyamine; 6; beta-hydroxylase; putrescine N-methyltransferase; scopolamine; transformation;
D O I
10.1073/pnas.0401391101
中图分类号
O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];
学科分类号
07 ; 0710 ; 09 ;
摘要
Scopolamine is a pharmaceutically important tropane alkaloid extensively used as an anticholinergic agent. Here, we report the simultaneous introduction and overexpression of genes encoding the rate-limiting upstream enzyme putrescine N-methyltransferase (PMT) and the downstream enzyme hyoscyamine 6 beta-hydroxylase (H6H) of scopolamine biosynthesis in transgenic henbane (Hyoscyamus niger) hairy root cultures. Transgenic hairy root lines expressing both pmt and h6h produced significantly higher (P < 0.05) levels of scopolamine compared with the wild-type and transgenic lines harboring a single gene (pmt or h6h). The best line (T-3) produced 411 mg/liter scopolamine, which was over nine times more than that in the wild type (43 mg/liter) and more than twice the amount in the highest scopolamine-producing h6h single-gene transgenic line H-11 (184 mg/liter). To our knowledge, this is the highest scopolamine content achieved through genetic engineering of a plant. We conclude that transgenic plants harboring both pmt and h6h possessed an increased flux in the tropane alkaloid biosynthetic pathway that enhanced scopolamine yield, which was more efficient than plants harboring only one of the two genes. It seems that the pulling force of the downstream enzyme (the faucet enzyme) H6H plays a more important role in stimulating scopolamine accumulation in H. niger whereas the functioning of the upstream enzyme PMT is increased proportionally. This study provides an effective approach for large-scale commercial production of scopolamine by using hairy root culture systems as bioreactors.
引用
收藏
页码:6786 / 6791
页数:6
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