Molecular genetics of isoquinoline alkaloid biosynthesis in the opium poppy Papaver somniferum

罂粟“罂粟”今天仍然是我们最重要的药用植物之一。在这种植物产生的80种生物碱中，有三种是重要的药用成分。它们是麻醉性镇痛药吗啡，镇痛和止咳药可待因和止咳药诺斯卡品。可待因和吗啡的生物合成几乎完全在酶水平上阐明。然而，关于诺斯卡品的生物合成了解相对较少。 我们对生物碱的生物合成是如何调节的，以及这些化合物在植物中的生物学作用也知之甚少。我们正在系统地分离编码罂粟生物碱生物合成的独特酶的cDNA。这些cDNA在细菌和昆虫细胞培养物中功能异源表达并表征。我们在此特别建议纯化和部分测序吗啡生物合成蒂巴因合酶，并分离相应的cDNA克隆。我们还将分离编码甲基转移酶特异性生物碱生物合成在这种植物的cDNA。我们已经从“罂粟”中分离到的7个cDNA将用于原位杂交，并将编码的异源蛋白用于免疫定位研究，以确定由该植物产生的一些不同类别的异喹啉生物碱（吗啡喃，苯并[c]菲啶，邻苯二甲酰异喹啉）的生物合成的细胞位点。这些结果应提供第一个洞察如何生物合成和积累的各种类别的这些生物碱在这种植物的调节。

The opium poppy "Papaver somniferum" is still today one of our most important medicinal plants. Among the 80 alkaloids produced by this plant, three are medicinally important. These are the narcotic analgesic morphine, the analgesic and the antitussive codeine and the antitussive noscapine. The biosynthesis of codeine and morphine is almost completely elucidated at the enzyme level. Relatively little is understood, however, concerning the biosynthesis of noscapine. We also understand very little of how the alkaloid biosynthesis is regulated and of the biological role of these compounds in the plant. We are systematically isolating cDNAs that encode the unique enzymes of alkaloid biosynthesis in opium poppy. These cDNAs are functionally heterologously expressed in bacterial and insect cell cultures and characterized. We propose specifically herein to purify and partially sequence thebaine synthase of morphine biosynthesis, and to isolate the corresponding cDNA clone. We will also isolate cDNAs encoding methyltransferases specific to alkaloid biosynthesis in this plant. The seven cDNAs that we have isolated to date from "P. somniferum" will be used in situ hybridization and the encoded heterologous proteins in immunolocalization studies in order to identify the cellular sites of biosynthesis of some of the various classes of isoquinoline alkaloids (morphinan, benzo[c]phenanthridine, phtalideisoquinoline) produced by this plant. These results should provide the first insight as to how biosynthesis and accumulation of the various classes of these alkaloids is regulated in this plant.