Refining Oxidative Enzyme Systems from Talented Microorganisms for Industrial Biocatalysis.

微生物产生可用于广泛工业应用的酶，但一个特别具有挑战性的领域是正在开发的药物的氧化代谢物的产生以及农用化学品代谢物的环境归宿。前者是由人类的肝酶产生的，它们的特性需要了解，它们的存在需要在临床试验中监测。后者是在环境中产生的，它们的性质也需要了解，因为它们可以持续很长一段时间。这两种代谢物类别可能非常难以化学合成。人类药物代谢物可以使用哺乳动物组织制剂生产，而农业化学品代谢物可以在环境模型中生产，但在新产品开发期间生产所需的数量可能具有挑战性。微生物系统提供了一个非常有用的替代方案。菌丝发现已经组装了一组野生型细菌，其已被证明通过全细胞微生物培养生物转化在产生一组>100个工业相关小有机分子的目标氧化代谢物方面非常有效，并且与目前市售的基于单一类型酶的基因工程微生物酶制剂相比，表现出显著更好的性能。菌丝发现小组还证明了在制造早期药物先导化合物的新衍生物方面的有效性，这些化合物具有改善的性质，特别是溶解度，这对药物生物利用度很重要。然而，这种全培养物生物转化方法在目标代谢物的生产速度及其生产到数克和最终千克数量的可扩展性方面具有局限性。该项目旨在通过识别编码负责在菌丝微生物组中六种最有才华的细菌中产生氧化代谢物的单个酶的基因来应对这一挑战，克隆并将这些基因引入特征良好的宿主细菌中，这些宿主细菌可以在标准条件下在小规模或大规模的实验室培养中生长。这项工作将与伦敦大学学院约翰·沃德教授的小组合作进行，他们率先提出了适当的程序。这项工作将包括对这些生物的整个基因组进行测序，鉴定感兴趣的酶的序列，并进行克隆，以产生表达感兴趣的酶的宿主菌株的基因工程衍生物，沿着其完全功能活性所需的辅因子，具有扩大规模的潜力。这项新的合作希望在氧化工业生物技术的应用方面取得重大进展，并以双方以往的高度互补的经验为基础。这是英国学术和工业机构提供相互支持的一个例子，最终应该有利于制药和农用化学品行业及其全球客户。

Microorganisms produce enzymes useful for a wide range of industrial applications but one particularly challenging area is the production of oxidized metabolites of drugs that are being developed and also the environmental fate of metabolites of agrochemicals. The former are produced in humans by liver enzymes and their properties need to be understood, and their presence monitored in clinical trials. The latter are produced in the environment and their properties again need to be understood as they can persist for extended periods. Both metabolite categories can be very difficult to synthesise chemically. Human drug metabolites can be produced using mammalian tissue preparations, and agrochemical metabolites in environmental models, but producing either in the quantities required during new product development can be challenging. Microbial systems offer a very useful alternative. Hypha Discovery has assembled a panel of wild-type bacteria that has proved highly effective in producing target oxidized metabolites for a set of >100 industrially-relevant small organic molecules by whole-cell microbial culture biotransformation, and shows significantly better performance compared to the current commercially available genetically-engineered microbial enzyme preparations which are based on a single type of enzyme. The Hypha Discovery panel has also proved effective in making new derivatives of early-stage pharmaceutical lead compounds with improved properties, in particular solubility, which is important for drug bioavailability. This whole-culture biotransformation approach has limitations, however, with regard to the speed of production of the target metabolite and the scalability of its production to multi-gramme and, eventually, kilogramme quantities. This project aims to address the challenge by identifying the genes encoding individual enzymes responsible for producing oxidized metabolites in the six most talented bacteria from Hypha's organism panel, cloning and introducing these genes into well-characterised host bacteria that can be grown in laboratory cultures under standard conditions at small- or large-scale. This work will be done in collaboration with Professor John Ward's group at University College, London, who have pioneered appropriate processes. The work will involve sequencing the whole genomes of these organisms, identifying the sequences for the enzymes of interest, and undertaking the cloning required to produce genetically-engineered derivatives of the host strains expressing the enzymes of interest along with co-factors required for their full functional activity, with scale-up potential.This new collaboration hopes to deliver a significant advance in the application of oxidative industrial biotechnology, and builds on the previous, and highly complementary, experience of both partners. It is an example of UK academic and industrial institutions providing mutual support that should eventually be of benefit to the pharmaceutical and agrochemical industries, and their customers, worldwide.