Biosynthesis of plant-specific stilbene polyketides in metabolically engineered Escherichia coli.
Biosynthesis of plant-specific stilbene polyketides in metabolically engineered Escherichia coli.
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DOI：
10.1186/1472-6750-6-22
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发表时间：
2006-03-21
影响因子：
3.5
通讯作者：
Schmidt-Dannert, Claudia
中科院分区：
文献类型：
--
作者：
Watts, Kevin T;Lee, Pyung C;Schmidt-Dannert, Claudia
苯丙烷类化合物是一系列重要的植物代谢物的前体，如细胞壁成分木质素和属于类黄酮/二苯乙烯类化合物的次生代谢物。后一类植物天然产物已被证明在广泛的生物活性中发挥作用。在过去的几年里，越来越多的健康益处与这些化合物有关。特别是，它们表现出强大的抗氧化活性和选择性地抑制某些酪氨酸激酶的能力。许多医学上重要的植物次生代谢物的生物合成，包括二苯乙烯类化合物，通常不是很好的了解，并且受到严格的空间和时间控制，限制了它们从植物来源获得。作为另一种选择，我们试图开发一种通过工程重组微生物细胞生物合成不同二苯乙烯类化合物的方法。利用从拟南芥中克隆的4-香豆酰辅酶A连接酶(4CL1)和从花生中克隆的二苯乙烯合成酶(STS)，在大肠杆菌中构建了二苯乙烯生物合成途径。表达这些酶的大肠杆菌培养物将添加到生长介质中的苯丙酸前体4-香豆酸转化为二苯乙烯类白藜芦醇(100 mg/L)。以同样的方式加入咖啡酸，会产生预期的二羟基二苯乙烯--二苯乙烯酚(>；10毫克/L)。然而，阿魏酸并没有转化为预期的二苯乙烯产品--异哈桥基元。用阿魏酸优先于4-香豆酸的同源酶4CL4取代4CL1对阿魏酸的转化率没有影响。从阿魏酸中积累三酮和四酮内酯，而不考虑在大肠杆菌中表达的辅酶A连接酶，表明STS不能正确地调节和折叠相应二苯乙烯结构的中间四酮。苯丙酸，如4-香豆酸和咖啡酸，可以通过表达植物生物合成基因的重组大肠杆菌有效地转化为二苯乙烯类化合物。通过宿主代谢工程和蛋白质工程对阿魏酸前体的前体转化和环化进行优化，可能会合成结构更加多样化的二苯乙烯类化合物。
Phenylpropanoids are the precursors to a range of important plant metabolites such as the cell wall constituent lignin and the secondary metabolites belonging to the flavonoid/stilbene class of compounds. The latter class of plant natural products has been shown to function in a wide range of biological activities. During the last few years an increasing number of health benefits have been associated with these compounds. In particular, they demonstrate potent antioxidant activity and the ability to selectively inhibit certain tyrosine kinases. Biosynthesis of many medicinally important plant secondary metabolites, including stilbenes, is frequently not very well understood and under tight spatial and temporal control, limiting their availability from plant sources. As an alternative, we sought to develop an approach for the biosynthesis of diverse stilbenes by engineered recombinant microbial cells. A pathway for stilbene biosynthesis was constructed in Escherichia coli with 4-coumaroyl CoA ligase 1 4CL1) from Arabidopsis thaliana and stilbene synthase (STS) cloned from Arachis hypogaea. E. coli cultures expressing these enzymes together converted the phenylpropionic acid precursor 4-coumaric acid, added to the growth medium, to the stilbene resveratrol (>100 mg/L). Caffeic acid, added in the same way, resulted in the production of the expected dihydroxylated stilbene, piceatannol (>10 mg/L). Ferulic acid, however, was not converted to the expected stilbene product, isorhapontigenin. Substitution of 4CL1 with a homologous enzyme, 4CL4, with a preference for ferulic acid over 4-coumaric acid, had no effect on the conversion of ferulic acid. Accumulation of tri- and tetraketide lactones from ferulic acid, regardless of the CoA-ligase expressed in E. coli, suggests that STS cannot properly accommodate and fold the tetraketide intermediate to the corresponding stilbene structure. Phenylpropionic acids, such as 4-coumaric acid and caffeic acid, can be efficiently converted to stilbene compounds by recombinant E. coli cells expressing plant biosynthetic genes. Optimization of precursor conversion and cyclization of the bulky ferulic acid precursor by host metabolic engineering and protein engineering may afford the synthesis of even more structurally diverse stilbene compounds.