Reconstitution of monoterpene indole alkaloid biosynthesis in genome engineered Nicotiana benthamiana.
Reconstitution of monoterpene indole alkaloid biosynthesis in genome engineered Nicotiana benthamiana.
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DOI：
10.1038/s42003-022-03904-w
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发表时间：
2022-09-10
影响因子：
5.9
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中科院分区：
文献类型：
--
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单萜烯吲哚生物碱(MIA)是一类种类繁多的植物天然产物，其中包括许多具有重要药用价值的化合物。我们着手从本氏烟草的中枢代谢重建所有MIA的关键中间体--斯特列托西丁的途径。这种宿主的一个缺点是其丰富的背景代谢导致了一些异源产生的分子的衍生化。在这里，我们使用转录分析来鉴定糖基转移酶，这些糖基转移酶对生物合成中间体的反应是上调的，并产生编码它们的基因具有靶向突变的植物系。早期MIA途径在这些品系中的表达产生了更有利的产物图谱。成功地重组了马钱子苷的生物合成，14种酶的共表达获得了最好的产量，其中来自香菜的主要乳胶蛋白样酶(MLPL)是通过环烯醚类化合物途径提高通量的关键。内源性糖基转移酶的去除不会影响Strectosidine的产量，这突显了途径酶到稳定的生物合成中间体的代谢流最大限度地减少了对调节宿主内源代谢的需要。在Planta中生产Strectosidine扩大了MIA产品的范围，使其能够进行生物合成。本烟草是单萜吲哚生物碱的关键中间体，其生物合成在烟草中成功重组，展示了本烟草作为小分子生物生产底盘的潜力。
Monoterpene indole alkaloids (MIAs) are a diverse class of plant natural products that include a number of medicinally important compounds. We set out to reconstitute the pathway for strictosidine, a key intermediate of all MIAs, from central metabolism in Nicotiana benthamiana. A disadvantage of this host is that its rich background metabolism results in the derivatization of some heterologously produced molecules. Here we use transcriptomic analysis to identify glycosyltransferases that are upregulated in response to biosynthetic intermediates and produce plant lines with targeted mutations in the genes encoding them. Expression of the early MIA pathway in these lines produces a more favorable product profile. Strictosidine biosynthesis was successfully reconstituted, with the best yields obtained by the co-expression of 14 enzymes, of which a major latex protein-like enzyme (MLPL) from Nepeta (catmint) is critical for improving flux through the iridoid pathway. The removal of endogenous glycosyltransferases does not impact the yields of strictosidine, highlighting that the metabolic flux of the pathway enzymes to a stable biosynthetic intermediate minimizes the need to engineer the endogenous metabolism of the host. The production of strictosidine in planta expands the range of MIA products amenable to biological synthesis. The biosynthesis of strictosidine, a key intermediate of monoterpene indole alkaloids, was successfully reconstructed in Nicotiana benthamiana, demonstrating the potential of Nicotiana benthamiana as a bioproduction chassis for small molecules.
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