Department of Biotechnology, Yeungnam University, Gyeongsan, Gyeongbuk, 38541, Republic of Korea.
Department of Life Science and Biochemical Engineering, SunMoon University, 70 Sunmoon-ro 221, Tangjeong-myeon, Asan-si, Chungnam, 31460, Republic of Korea.
Appl Biochem Biotechnol. 2020 Jan;190(1):325-340. doi: 10.1007/s12010-019-03206-8. Epub 2019 Dec 18.
Curcuminoids are natural phenylpropanoids that are biosynthesized via an L-phenylalanine metabolism pathway in turmeric (Curcuma longa L.). Curcuminoids have various chemopreventive activities and pharmaceutical applications in human life. In this study, we synthesized dicinnamoylmethane, one principal component of curcuminoids, from cinnamic acid by means of co-expression of Oryza sativa curcuminoid synthase and Petroselinum crispum 4-coumarate-CoA ligase in Escherichia coli BL21 (DE3). Moreover, we used CRISPRi systems to knock down the genes in a tricarboxylic acid cycle and fatty acid biosynthesis pathway. The repression of target genes led to an increase of up to 0.236 μmol g DCW of malonyl-CoA in cytosol-engineered E. coli and subsequently increased the biosynthesis of dicinnamoylmethane. We found that the S10 strain containing a CRISPRi repression for three genes, fabF, fabD, and mdh, showed the highest amount of dicinnamoylmethane of 7.54 μM, which is 5.76-fold higher than that of the wild-type strain. Finally, 41.94 μM (~ 11.6 mg) of dicinnamoylmethane was obtained in a 3-L fermenter.
姜黄色素是天然苯丙烷类化合物，通过姜黄（Curcuma longa L.）中的 L-苯丙氨酸代谢途径生物合成。姜黄色素在人类生活中具有多种化学预防和药物应用。在这项研究中，我们通过在大肠杆菌 BL21（DE3）中共同表达水稻姜黄素合成酶和芹菜 4-香豆酰辅酶 A 连接酶，由肉桂酸合成姜黄色素的主要成分之一二肉桂酰甲烷。此外，我们使用 CRISPRi 系统敲低三羧酸循环和脂肪酸生物合成途径中的基因。靶基因的抑制导致细胞溶质工程大肠杆菌中丙二酰辅酶 A 的产量增加高达 0.236 μmol g DCW，随后增加了二肉桂酰甲烷的生物合成。我们发现，含有三个基因 fabF、fabD 和 mdh 的 CRISPRi 抑制的 S10 菌株表现出最高量的二肉桂酰甲烷，为 7.54 μM，比野生型菌株高 5.76 倍。最后，在 3-L 发酵罐中获得了 41.94 μM（约 11.6 mg）的二肉桂酰甲烷。
