Harnessing enzymes from plants for selective functionalisation of triterpenoid scaffolds

植物是出类拔萃的化学工程师。它们生产的天然产品在制药、农用化学品和工业生物技术行业具有巨大价值。然而，由于难以获得来源物种、产量低、纯化问题以及对环境可持续性的担忧，此类化合物的可获得性受到限制。绝大多数植物天然产物具有高度复杂的结构，超出了化学合成的范围，这一事实使这一问题进一步复杂化。这意味着，潜在的新药和其他有价值的化学品的巨大储藏库仍然被锁在生产它们的工厂内。越来越多的植物基因组序列的可获得性使人们有可能从这些序列中‘挖掘’编码产生这些不同化学物质的酶的基因。如果这些酶能够被利用并组装成一个工具包，使普通核心支架能够进行生物合成和系统多样化，这将使人类能够模仿和扩大植物王国中发现的化学多样性，例如，制造效率更高、副作用更少的新药。我们特别感兴趣的是一类非常大且结构复杂的植物天然产物，称为三萜类化合物。这类20,000种天然产物都来自相同的化学简单的共同前体，显示出巨大的结构多样性，是生物活性分子的丰富来源。众所周知的例子包括人参皂苷和甘草酸苷，人参皂苷和甘草酸苷是由甘草产生的甜味剂和抗炎物质。人们对三萜类化合物的药学性质很感兴趣，最近已有几种化合物作为抗炎/抗癌药物进入临床试验(甘草次酸和半合成的三萜类化合物巴度松龙甲基)。在这项提案中，我们描述了我们将如何发现植物如何合成和多样化复杂的三萜类支架。我们将获得催化这些过程的酶，并将它们部署到一种经过优化的工程面包酵母中，以支持三萜类支架的多样化。然后，我们将研究这些分子的特征，这些分子决定了它们的生物学活性。

Plants are chemical engineers par excellence. The natural products that they produce have tremendous value in the pharmaceutical, agrochemical and industrial biotechnology industries. However, the availability of such compounds is limited by difficulties in accessing source species, low yield, purification problems, and concerns about environmental sustainability. This problem is further confounded by the fact that the vast majority of plant natural products have highly complex structures and are beyond the reach of chemical synthesis. This means that a vast reservoir of potential new drugs and other valuable chemicals remain locked up inside the plants that produce them. The availability of a growing number of plant genome sequences is now making it possible to 'mine' these sequences for genes encoding the enzymes that make these diverse chemicals. If these enzymes can be harnessed and assembled into a toolkit that would enable biosynthesis and systematic diversification of common core scaffolds this would enable humans to mimic and expand on the chemical diversity found in the plant kingdom and, for example, to make new drugs with improved efficacy and reduced side effects. We are especially interested in a very large and structurally complex class of plant natural products known as triterpenoids. This class of >20,000 natural products, all derived from the same chemically simple common precursor display enormous structural diversity and are a rich source of bioactive molecules. Well known examples include ginsenosides, associated with the 'feel good factor' of ginseng, and glycyrrhizin, a sweetener and anti-inflammatory produced by liquorice. There is considerable interest in the pharmaceutical properties of triterpenoids and several have recently entered clinical trials as anti-inflammatory/anti-cancer drugs (glycyrrhetinic acid and the semi-synthetic triterpenoid bardoxolone methyl). In this proposal, we describe how we will discover how plants synthesise and diversify complex triterpenoid scaffolds. We will garner the enzymes that catalyse these processes and deploy them into a strain of engineered baker's yeast optimised to support triterpene scaffold diversification. We will then investigate the features of these molecules that determine their biological activities.