Enzyme Cascades and Synthetic Biology Routes to Non-Natural Alkaloids

几个世纪以来，植物和来自植物的药用化合物已用于治疗疼痛、感染、哮喘、癌症，并且这些植物天然产物中的许多用于现代医学，例如：镇痛剂吗啡和可待因;抗疟疾奎宁用于疟疾;长春碱和高三尖杉酯碱用于癌症治疗，以及麻黄碱用于哮喘。生物碱家族的天然化合物超过14，000种，许多具有生物活性。最近，人们对使用微生物生产这些天然化合物产生了浓厚的兴趣，而不是从世界上难以获得的植物中分离它们。合成生物学的方法来实现这一点被证明是非常强大的，最近的文章已经强调了从酵母生产吗啡的可能性。在以前的工作中，我们已经调查了一些早期的步骤和酶，使用氨基酸和执行转换到一个关键的中间体的途径，植物用来合成苄基异喹啉生物碱，一个家庭的生物碱超过2500。我们现在的目标是使用这些酶，与其他酶不用于植物途径，和新的酶，已经很少探索在生物催化应用。这些酶将用于实验室细菌中，以产生自然界中未发现的各种结构多样的化合物。将筛选化合物的抗菌性能。酶将在体外级联中用于产生三种、四种、五种或更多种酶途径，并且通过在细菌中建立这些途径，我们将能够使用可再生资源以可再现的方式产生新化合物。

Plants and medicinal compounds from plants have been used for centuries to treat pain, infections, asthma, cancer and many of these plant natural products are used in modern medicine, for example: the analgesics morphine and codeine; anti-malarial quinine for malaria; vinblastine and homoharringtonine for cancer treatment, and ephedrine for asthma. There are over 14,000 natural compounds of the alkaloid family and many possess biological activities. Recently, there has been intensive interest in producing some of these natural compounds using microbes, rather than isolating them from plants in parts of the world that are difficult to access. Synthetic biology approaches to achieve this are proving to be extremely powerful and recent articles have highlighted the possibility of producing morphine from yeast.In previous work we have investigated some of the early steps and enzymes that use amino acids and perform transformations towards a key intermediate in the pathways that plants use to synthesise the benzylisoquinoline alkaloids, a family of over 2500 alkaloids. We now aim to use these enzymes, together with other enzymes not used in the plant pathway, and new enzymes that have been little explored in biocatalytic applications. The enzymes will be used in a laboratory bacterium to generate a wide range of structurally diverse compounds that are not found in nature. Compounds will screened for antibacterial properties. Enzymes will be used in in vitro cascades to generate three, four, five, or more multiple enzyme pathways, and also by establishing the pathways in bacteria we will be able to generate new compounds in a reproducible way using a renewable resource.