Complete biosynthesis of cannabinoids and their unnatural analogues in yeast
Complete biosynthesis of cannabinoids and their unnatural analogues in yeast
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DOI：
10.1038/s41586-019-0978-9
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发表时间：
2019-03-07
期刊：
影响因子：
64.8
通讯作者：
Keasling, Jay D.
中科院分区：
文献类型：
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作者：
Luo, Xiaozhou;Reiter, Michael A.;Keasling, Jay D.
几千年来，大麻因其药用特性而在全球范围内被种植和使用。一些大麻类化合物，大麻的标志性成分，及其类似物因其潜在的医疗应用而得到了广泛的研究(2)。某些大麻素制剂已在几个国家被批准为治疗一系列人类疾病的处方药(3)。然而，大麻类化合物的研究和药用一直受到大麻的合法调度、几乎所有几十种已知大麻类化合物的植物丰度低(4)及其结构复杂性的阻碍，这限制了大量的化学合成。本文报道了酿酒酵母中主要的大麻酚酸、Delta(9)-四氢大麻酚酸、大麻二酚酸、Delta(9)-四氢大麻黄酸和大麻地黄酸的完整生物合成。为了实现这一点，我们设计了天然的甲羟戊酸途径来提供高通量的焦磷酸香叶基，并引入了一种异源的、多生物衍生的己酰-辅酶A生物合成途径(5)。我们还介绍了编码参与橄榄酸生物合成的酶的大麻基因(6)，以及以前未发现的具有香叶焦磷酸的酶的基因：橄榄叶香叶转移酶活性和相应的大麻素合成酶基因(7，8)。此外，我们建立了一种生物合成方法，利用几个途径基因的混杂来产生大麻素类似物。将不同的脂肪酸喂给我们的工程菌株产生了大麻素类似物，并对分子中已知的改变受体结合亲和力和效力的部分进行了修改(9)。我们还证明，我们的生物系统可以通过简单的合成化学来补充，以进一步扩大可接近的化学空间。我们的工作为生产天然和非天然大麻素提供了一个平台，将允许对这些化合物进行更严格的研究，并可用于开发各种人类健康问题的治疗方法。
Cannabis sativa L. has been cultivated and used around the globe for its medicinal properties for millennia(1). Some cannabinoids, the hallmark constituents of Cannabis, and their analogues have been investigated extensively for their potential medical applications(2). Certain cannabinoid formulations have been approved as prescription drugs in several countries for the treatment of a range of human ailments(3). However, the study and medicinal use of cannabinoids has been hampered by the legal scheduling of Cannabis, the low in planta abundances of nearly all of the dozens of known cannabinoids(4), and their structural complexity, which limits bulk chemical synthesis. Here we report the complete biosynthesis of the major cannabinoids cannabigerolic acid, Delta(9)-tetrahydrocannabinolic acid, cannabidiolic acid, Delta(9)-tetrahydrocannabivarinic acid and cannabidivarinic acid in Saccharomyces cerevisiae, from the simple sugar galactose. To accomplish this, we engineered the native mevalonate pathway to provide a high flux of geranyl pyrophosphate and introduced a heterologous, multi-organism-derived hexanoyl-CoA biosynthetic pathway(5). We also introduced the Cannabis genes that encode the enzymes involved in the biosynthesis of olivetolic acid(6), as well as the gene for a previously undiscovered enzyme with geranylpyrophosphate: olivetolate geranyltransferase activity and the genes for corresponding cannabinoid synthases(7,8). Furthermore, we established a biosynthetic approach that harnessed the promiscuity of several pathway genes to produce cannabinoid analogues. Feeding different fatty acids to our engineered strains yielded cannabinoid analogues with modifications in the part of the molecule that is known to alter receptor binding affinity and potency(9). We also demonstrated that our biological system could be complemented by simple synthetic chemistry to further expand the accessible chemical space. Our work presents a platform for the production of natural and unnatural cannabinoids that will allow for more rigorous study of these compounds and could be used in the development of treatments for a variety of human health problems.