21ENGBIO: Engineering targeted activation of fungicides at the plant-pathogen interface

要保证足够养活全世界的粮食供应，就需要使用化学杀真菌剂来保护作物免受真菌和其他微生物的侵害。杀真菌剂及其生产行业在经济上至关重要，但大量使用杀真菌剂可能对环境造成风险。损害作物的真菌可以迅速进化出对个别化学品的抗性，随着真菌进一步适应，这就推动了喷洒到农田上的剂量的“逐步增加”。这个过程很快达到一个阶段，即使是最高剂量的杀真菌剂也变得无效。因此，需要不断开发新的杀菌剂化学品，以跟上抗药性现象的步伐，并最大限度地减少释放到环境中的杀菌剂的量。发现对真菌有毒的新化学品并不意味着它立即适合作为商业产品。一种“先导化合物”必须优化其效力，通过严格的安全测试，并具有合适的特性，以便分配到田间和作物中。对先导化合物进行修饰，使其对真菌更有效，但可能使其在其他方面（如溶解度）效果较差。制药行业在新药开发过程中也遇到了类似的问题。这个问题的一个解决方案是设计具有可溶性、毒性较小且适合递送到人体内的靶位点的化学结构的“前药”。然后，前体药物转化为药物形式;理想情况下，在需要的精确部位。我们建议将这一概念应用于农业杀菌剂，通过设计一种生物系统，可以在植物细胞和攻击真菌之间的界面上激活“前杀菌剂”。我们发现，可以将我们选择的酶（β-葡萄糖醛酸酶）专门发送到植物细胞表面上真菌试图攻击的位置。β-葡萄糖醛酸苷酶被人体内的一些前药利用，通过释放葡萄糖醛酸形成活性药物。当与前药连接时，葡糖醛酸化学基团支持化合物的溶解度，并且与药物形式相比可以降低前药活性。我们将优化我们的β-葡萄糖醛酸酶系统，并生产一种带有葡萄糖醛酸基团的前杀真菌剂。这将用于模式植物拟南芥的“概念验证”实验。我们将测量杀真菌剂前体渗透到植物中的能力的预测改善，并且我们还将测量我们的合成β-葡萄糖醛酸糖苷酶在需要的部位将杀真菌剂前体转化为杀真菌剂的效率。我们将联合收割机的专业知识，从多个科学领域（生物学，化学和物理学），以实现这一目标。这个工程生物学的例子将用于建立新的工业合作伙伴关系，以利用我们的亲杀真菌剂战略“释放”废弃或具有挑战性的先导化合物的全部潜力。我们还旨在激发新一代的想法，利用我们对植物免疫系统如何与致病微生物物理相互作用的不断增长的理解。

A secure food supply sufficient to feed the world requires the use of chemical fungicides to protect crops from fungi and other micro-organisms. Fungicides and the industry that produces them are economically critical, but intensive use of fungicides can pose a risk to the environment. The fungi that damage crops can quickly evolve resistance to individual chemicals and this drives the 'ratcheting-up' of doses sprayed onto fields as the fungi further adapt. This process quickly reaches a stage where even the highest doses of fungicide become ineffective. The constant development of new fungicidal chemicals is therefore required to keep pace with the phenomenon of resistance and to minimise the amount of fungicide released into the environment.Discovery of a new chemical that is toxic to fungi does not mean it is immediately suitable as a commercial product. A 'lead compound' must be optimised for potency and pass rigorous safety tests as well as have suitable characteristics for being distributed onto the field and into crops. Modifications to the lead compound that make it more effective against fungi can make it less effective in some other aspect such as its solubility. Similar problems are encountered by the pharmaceutical industry during new drug development. One solution to this problem is the design of 'pro-drugs' that have a chemical structure that is soluble, less toxic and suitable for delivery to the target site within the human body. The pro-drug then becomes converted to the drug form; ideally at the precise site where it is needed. We propose to adapt this concept to agricultural fungicides by engineering a biological system that can activate 'pro-fungicides' exactly where they are needed at interfaces between plant cells and attacking fungi.We have discovered that it is possible to send an enzyme of our choice (beta-glucuronidase) exclusively to the site on a plant cell surface where a fungus is attempting an attack. Beta-glucuronidase is exploited by some pro-drugs in the human body to form an active drug through the release of glucuronic acid. When attached to the pro-drug the glucuronic acid chemical group supports the solubility of the compound and can reduce the pro-drug activity compared to the drug form. We will optimise our beta-glucuronidase system and produce a pro-fungicide with an attached glucuronic acid group. This will be used in 'proof-of-concept' experiments with the model plant species Arabidopsis thaliana. We will measure the predicted improvement in the ability of the pro-fungicide to penetrate into the plant and we will also measure the efficiency of our synthetic beta-glucuronidase in converting pro-fungicide to fungicide at the site it is needed. We will combine expertise from multiple areas of science (biology, chemistry and physics) to achieve this. This example of engineered biology will be used to establish new industrial partnerships to 'unlock' the full potential of abandoned or challenging lead compounds using our pro-fungicide strategy. We also aim to inspire a new generation of ideas that utilise our growing understanding of how the plant immune system physically interacts with disease-causing microbes.