中国科学院昆明植物研究所黄胜雄团队报道了可卡因生物合成途径的发现与工程。相关研究成果于2022年11月14日发表于国际顶尖学术期刊《美国化学会杂志》。

可卡因，一种来自植物赤藓属的原型三羟甲基丙烷生物碱，最近被临床用作粘膜表面麻醉。尽管如此，从所鉴定的中间体4-(1-甲基-2-吡啶基)-3-氧丁酸(MPOA)中提取所需的三烷骨架（甲基环烯酮）的关键生物合成步骤直到目前为止仍然未知。

文中，研究人员通过本氏烟草中候选基因的瞬时表达，鉴定了可卡因中烷类骨架生物合成所必需的两种缺失酶（EnCYP81AN15和EnMT4）。观察到细胞色素P450 EnCYP81AN15选择性地介导S-MPOA的氧化环化，以产生不稳定的中间体芽子碱，然后通过甲基转移酶EnMT4将其甲基化，形成光学活性的甲基芽子碱。该途径的建立纠正了长期以来（但不正确的）MPOA甲基化第一和氧化环化第二的生物合成假说。值得注意的是，利用两个新发现的基因以及四个已知的基因，在本烟草中实现了可卡因的从头重建。

该项研究不仅报告了可卡因的近乎完整的生物合成途径，并为植物中的烷类生物碱（可卡因和东莨菪碱）的代谢网络提供了新的见解，而且还能够在其他（微生物）生物中异源合成烷类生物碱，这对药物生产具有重要意义。

附：英文原文

Title: Discovery and Engineering of the Cocaine Biosynthetic Pathway

Author: Yong-Jiang Wang, Jian-Ping Huang, Tian Tian, Yijun Yan, Yin Chen, Jing Yang, Jianghua Chen, Yu-Cheng Gu, Sheng-Xiong Huang

Issue&Volume: November 14, 2022

Abstract: Cocaine, the archetypal tropane alkaloid from the plant genus Erythroxylum, has recently been used clinically as a topical anesthesia of the mucous membranes. Despite this, the key biosynthetic step of the requisite tropane skeleton (methylecgonone) from the identified intermediate 4-(1-methyl-2-pyrrolidinyl)-3-oxobutanoic acid (MPOA) has remained, until this point, unknown. Herein, we identify two missing enzymes (EnCYP81AN15 and EnMT4) necessary for the biosynthesis of the tropane skeleton in cocaine by transient expression of the candidate genes in Nicotiana benthamiana. Cytochrome P450 EnCYP81AN15 was observed to selectively mediate the oxidative cyclization of S-MPOA to yield the unstable intermediate ecgonone, which was then methylated to form optically active methylecgonone by methyltransferase EnMT4 in Erythroxylum novogranatense. The establishment of this pathway corrects the long-standing (but incorrect) biosynthetic hypothesis of MPOA methylation first and oxidative cyclization second. Notably, the de novo reconstruction of cocaine was realized in N. benthamiana with the two newly identified genes, as well as four already known ones. This study not only reports a near-complete biosynthetic pathway of cocaine and provides new insights into the metabolic networks of tropane alkaloids (cocaine and hyoscyamine) in plants but also enables the heterologous synthesis of tropane alkaloids in other (micro)organisms, entailing significant implications for pharmaceutical production.

DOI: 10.1021/jacs.2c09091

Source: https://pubs.acs.org/doi/10.1021/jacs.2c09091