β-葡萄糖苷酶高效分泌及催化合成稀有人参皂苷Compound K的机制研究

随着“健康中国”战略的实施，“防慢病，治未病，坚持预防为主，防治结合”的理念已成共识。稀有人参皂苷Compound K在众多疾病防治和治疗方面展现出巨大潜力，但自然来源受限、难以获取，因此如何高效低成本获得大量高纯度稀有人参皂苷Compound K是当前人参研究开发的热点之一。本项目致力于构建具有高效表达β-葡萄糖苷酶活性的酵母菌株，以高效、绿色、简洁的细胞表面展示技术制备固定化β-葡萄糖苷酶。在此基础上，基于ARTP技术的非理性诱变育种，结合液滴微流控技术具有的高通量筛选功能，极大的提高了获取目标突变株的速度。为进一步增强菌株的生产能力，突变菌株进行转录组学分析，结合反向工程手段挖掘与改造酵母关键作用靶点，强化催化表达系统。针对产物的合成，设计催化反应介质，建立高效催化合成体系。通过系统改造与优化固定化酶与反应体系的关键因素，以提升稀有人参皂苷Compound K的大规模生物制造能力。

With the implementation of the "Healthy China" strategy, the concept of "preventing chronic diseases, treatmenting predisease, adhering to the principle of prevention first and integrating prevention with control " has become a consensus. Rare ginsenoside CompoundK shows great potential in the prevention and treatment of a wide range of diseases. However, the natural source is limited and difficult to obtain. Therefore, how to obtain a large quantities of high-purity rare ginsenoside CompoundK with high efficiency and low cost is one of the hot spots in current ginseng research and development. This project aims to construct a yeast strain with high expression activity of β-glucosidase, and to prepare immobilized β-glucosidase by efficient, green and simple cell surface display technology. On this basis, irrational mutagenesis based on ARTP technology, combined with the high-throughput screening function of droplet microfluidic technology, can greatly improve the speed of acquiring target mutant strains. In order to further enhance the production capacity of the mutant strain, transcriptome analysis is performed, and the key action targets are excavated and modified with reverse engineering methods to strengthen the catalytic expression system. For the synthesis of CompoundK, the catalytic reaction medium is optimized and the efficient catalytic transformation system is established.