Katsuyama Yohei, Funa Nobutaka, Horinouchi Sueharu
Department of Biotechnology, Graduate School of Agriculture and Life Sciences, The University of Tokyo, Tokyo, Japan.
Biotechnol J. 2007 Oct;2(10):1286-93. doi: 10.1002/biot.200700098.
Stilbenes are bioactive compounds that show beneficial effects for humans, such as anti-tumor activity and survival improvement. Resveratrol, a representative of stilbenes and showing various health-improving activities, is rapidly metabolized in humans, and modified resveratrols are therefore desired as anti-cancer drugs and dietary polyphenols. An Escherichia coli system, in which an artificial stilbene biosynthetic pathway, including steps of phenylalanine ammonia-lyase, 4-coumarate:CoA ligase, and stilbene synthase, was reconstructed, produced stilbenes in high yields: resveratrol from tyrosine and pinosylvin from phenylalanine. To incorporate a stilbene methyltransferase gene into this E. coli system, cDNA of Os08g06100 in Oryza sativa was expressed and its O-methylating activity toward stilbenes was confirmed. Incorporation of the pinosylvin methyltransferase (OsPMT) gene into the pathway established in E. coli led to production of mono- and di-methylated stilbenes. Furthermore, the OsPMT gene turned out to be useful in production of unnatural stilbene methyl ethers due to its rather relaxed substrate specificity; various carboxylic acids supplemented as precursors, such as p-fluorocinnamic acid, 3-(2-furyl)acrylic acid, 3-(2-thienyl)acrylic acid, and 3-(3-pyridyl)acrylic acid, to the E. coli system carrying the steps of 4-coumarate:CoA ligase, stilbene synthase, and OsPMT were converted to stilbene dimethyl ethers with the corresponding carboxylic moiety.
芪类化合物是对人体具有有益作用的生物活性化合物，如抗肿瘤活性和改善生存能力。白藜芦醇作为芪类化合物的代表，具有多种改善健康的活性，但其在人体内会迅速代谢，因此，修饰后的白藜芦醇有望作为抗癌药物和膳食多酚。在大肠杆菌系统中，构建了一条人工芪生物合成途径，该途径包括苯丙氨酸解氨酶、4-香豆酸：辅酶A连接酶和芪合酶等步骤，能够高产芪类化合物：由酪氨酸生成白藜芦醇，由苯丙氨酸生成松脂醇。为了将芪甲基转移酶基因整合到该大肠杆菌系统中，表达了水稻中Os08g06100的cDNA，并证实了其对芪类化合物的O-甲基化活性。将松脂醇甲基转移酶（OsPMT）基因整合到大肠杆菌中建立的途径中，可导致单甲基化和二甲基化芪类化合物的产生。此外，由于OsPMT基因的底物特异性相当宽松，它在生产非天然芪甲基醚方面很有用；向携带4-香豆酸：辅酶A连接酶、芪合酶和OsPMT步骤的大肠杆菌系统中添加各种作为前体的羧酸，如对氟肉桂酸、3-(2-呋喃基)丙烯酸、3-(2-噻吩基)丙烯酸和3-(3-吡啶基)丙烯酸，均可转化为带有相应羧基部分的芪二甲醚。
