Yamada Yasuyuki, Tamagaki Emi, Shitan Nobukazu, Sato Fumihiko
Laboratory of Medicinal Cell Biology, Kobe Pharmaceutical University, Kobe, Hyogo 658-8558, Japan.
Graduate School of Biostudies, Kyoto University, Kyoto, Kyoto 606-8502, Japan.
Plant Biotechnol (Tokyo). 2024 Sep 25;41(3):267-276. doi: 10.5511/plantbiotechnology.24.0205a.
Structurally diverse benzylisoquinoline alkaloids (BIAs) are found in specific plant families, some of which are desirable for their efficient production because of their strong biological activities. plants (e.g., ) of the family Papaveraceae also produce various BIAs; thus, they have been used in traditional Chinese medicine. Because metabolic engineering and synthetic biology using microorganisms are promising technologies for the effective production of useful metabolites, elucidation of the biosynthetic pathway of each BIA is indispensable. Although several enzyme genes involved in the biosynthesis of BIAs have recently been isolated, many remain unknown, such as the protoberberine alkaloid -methyltransferase involved in the biosynthesis of corydaline, one of the main BIAs found in the tubers of plants. In this study, we performed transcriptome analysis combined with metabolite profiling of different tissues of . Based on the high accumulation of several BIAs, including protopine, allocryptopine, and corydaline, genes encoding putative biosynthetic enzymes, including cytochrome P450, methyltransferase, and oxidase proteins, that were highly expressed in the tubers were screened. Two OMT genes, and , were highly expressed in the tuber, and further characterization using crude enzyme preparations demonstrated that CsOMT1 showed 7--methylation activity against reticuline, whereas CsOMT2 catalyzed 9--methylation of scoulerine, followed by 2--methylation of tetrahydrocolumbamine. Our findings provide valuable information for the isolation of novel biosynthetic enzyme genes in species.
结构多样的苄基异喹啉生物碱（BIAs）存在于特定的植物科中，其中一些因其强大的生物活性而有望实现高效生产。罂粟科植物（如……）也能产生多种BIAs；因此，它们已被用于传统中药。由于利用微生物的代谢工程和合成生物学是有效生产有用代谢产物的有前景的技术，阐明每种BIA的生物合成途径是必不可少的。尽管最近已经分离出了一些参与BIAs生物合成的酶基因，但仍有许多未知基因，例如参与紫堇碱生物合成的原小檗碱生物碱 -甲基转移酶，紫堇碱是……植物块茎中发现的主要BIAs之一。在本研究中，我们对……不同组织进行了转录组分析并结合代谢物谱分析。基于原阿片碱、别隐品碱和紫堇碱等几种BIAs的高积累，筛选出了在块茎中高表达的编码假定生物合成酶的基因，包括细胞色素P450、甲基转移酶和氧化酶蛋白。两个OMT基因，……和……，在块茎中高表达，使用粗酶制剂的进一步表征表明，CsOMT1对网状番荔枝碱具有7--甲基化活性，而CsOMT2催化番荔枝宁的9--甲基化，随后是四氢小檗胺的2--甲基化。我们的研究结果为分离……物种中的新型生物合成酶基因提供了有价值的信息。
