Unlocking the Next-Generation Protein Expression Systems for Animal-Free Cheese Production

由于收入增加、人口增加和饮食变化，全球对乳制品的需求正在增长。令人担忧的是，乳制品生产是世界上最大的甲烷排放源之一。乳制品的无动物替代品有可能通过提供可持续的生产实践来减少污染。然而，尽管植物基牛奶替代品的价格、口味和便利性几乎与动物源性牛奶相当，但植物基奶酪仍然未能提供足够的消费者体验。在过去的10年里，精确发酵的潜在用途--一种将转基因微生物用作细胞工厂以生产所需生物分子的过程，例如药品和生物燃料--已经被研究用于食品蛋白质生产。虽然在原则上精确发酵提供了一个可持续的和道德的方式生产食品蛋白质，到目前为止，没有主要的替代蛋白质公司已经能够提供负担得起的和可扩展的解决方案，酪蛋白-主要的奶酪蛋白- production.We认为，缺乏进展主要是由于试图基因工程传统的工业微生物，如_E。coli和S.酿酒酵母需要多次修饰，以利用廉价的食物来源，承受生物反应器中的环境压力，并产生复杂的蛋白质。事实上，自2014年以来，在将优化工作扩大到不太受欢迎但科学上成熟的巴斯德毕赤酵母和T之后，替代蛋白质领域已经取得了很大的进步。reesei_.我们建议通过采用可能更适合食品生产但目前缺乏必要的基因工程工具的非常规酵母物种可以实现更大的收益。我们的目标是使用非常规微生物物种开发下一代微生物工厂。我们的方法将在中试规模的生物反应器中进行验证，以评估用于酪蛋白工业生产的新型菌株。该原理验证项目将通过大规模扩大蛋白质表达系统的选择，为整个精密发酵行业提供服务，为工业级可扩展性量身定制。

The demand for dairy products is growing globally due to the rising income, increasing population, and dietary changes. Concerningly, dairy production is one of the biggest methane emitters in the world. Animal-free alternatives for dairy products have the potential to reduce pollution by offering sustainable production practices. However, while plant-based alternatives for milk have nearly reached price, taste, and convenience parity to animal-derived milk, plant-based cheese still fails to offer an adequate consumer experience.In the last 10 years, the potential use of precision fermentation -- a process where genetically modified microorganisms are used as cell factories to produce desired biomolecules, such as pharmaceuticals and biofuels -- has been investigated for food protein production. While in principle precision fermentation offers a sustainable and ethical way of producing food proteins, to date no major alternative protein company has been able to offer affordable and scalable solutions to casein -- the main cheese protein -- production.We argue that the lack of progress is mainly due to the attempts to genetically engineer conventional industrial microorganisms such as _E. coli_ and _S. cerevisiae_ that require multiple modifications to utilise cheap food sources, withstand environmental pressures in bioreactors, and produce complex proteins. Indeed, since 2014 the alternative protein sector already saw great improvements after expanding optimisation efforts to less popular but scientifically well-established species of _P. pastoris_ and _T. reesei_. We propose that even larger gains can be achieved by adopting unconventional yeast species that might be naturally more suitable for food production but currently lack the necessary genetic engineering tools.We aim to develop the next-generation microbial factories using unconventional microbial species. Our methodology will be validated in pilot-scale bioreactors evaluating the novel strains for the industrial production of casein proteins. This proof-of-principle project will serve the whole precision fermentation sector by massively expanding the choice of protein expression systems, tailored for industrial level scalability.