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Rational metabolic-flow switching for the production of exogenous secondary metabolites in bamboo suspension cells

机译:在竹悬浮细胞中产生外源次生代谢产物的合理代谢流转换

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摘要

The synthetic biology-driven production of high-value plant secondary metabolites in microbial hosts has attracted extensive attention despite various challenges, including correct protein expression and limited supplies of starting materials. In contrast, plant cell cultures are rarely used for this purpose owing to their slow proliferation rates and laborious transformation processes. Here, we propose a “rational metabolic-flow switching” strategy to efficiently produce exogenous secondary metabolites using suspension-cultured bamboo (Phyllostachys nigra; Pn) cells as model production hosts. The Pn cells biosynthesise hydroxycinnamic acid amides (HCAAs) of putrescine as major secondary metabolites, which indicates that the phenylpropanoid and polyamine biosynthetic pathways are highly active and that the Pn cells may produce alternative secondary metabolites derived from those pathways. Stable transformants of Pn cells expressing agmatine coumaroyltransferase of barley (Hordeum vulgare) were generated with the expectation of metabolic-flow switching from HCAAs of putrescine to those of agmatine. In the recombinant Pn cells, the levels of HCAAs of putrescine decreased and the HCAAs of agmatine were produced instead. The production titre of the major product, p-coumaroylagmatine, reached approximately 360 mg/L, providing a proof-of-concept for the usefulness of “rational metabolic-flow switching” in synthetic biology using plant cell hosts.
机译:尽管存在各种挑战,包括正确的蛋白质表达和有限的起始原料供应,但在微生物宿主中合成生物学驱动的高价值植物次生代谢产物的生产引起了广泛关注。相反,植物细胞培养物由于其缓慢的增殖速率和费力的转化过程而很少用于该目的。在这里,我们提出了一种“合理的代谢流转换”策略,以使用悬浮培养的竹(Phyllostachys nigra; Pn)细胞作为模型生产宿主来有效地生产外源性次生代谢产物。 Pn细胞可生物合成腐胺的羟基肉桂酸酰胺(HCAAs)作为主要的次要代谢产物,这表明苯丙烷和多胺的生物合成途径具有很高的活性,并且Pn细胞可产生源自这些途径的替代性次要代谢产物。期望从腐胺的HCAA转化为胍丁胺的HCAA的代谢流转换,产生了表达大麦的胍丁胺香豆酰香豆酰转移酶的Pn细胞的稳定转化体。在重组Pn细胞中,腐胺的HCAA水平降低,而产生了胍丁胺的HCAA。主要产品对-香豆素嘧啶的生产效价达到约360 mg / L,为使用植物细胞宿主的合成生物学中“合理代谢流转换”的有效性提供了概念证明。

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