泡沫分离-低温凝聚相分离两相传质分离过程集成优化及其对多功能融合酶的选择性分离纯化

工业酶制剂是生物制造的核心，突破微生物酶大规模分离纯化的关键技术高度契合国家战略需求。本课题以工业应用为导向，开发适合规模化生产的“泡沫分离-低温凝聚相分离”集成技术，通过构建多功能融合酶来解决下游分离纯化的瓶颈问题，主要研究内容如下：①将液-气（泡沫分离）与液-液（低温凝聚相分离）两种不同分离机制的两相分离法集成优化，在无化学药剂添加下实现目标酶的选择性分离纯化，解决传统分离纯化方法存在的产品收率低、酶活损失大和生产成本高的问题；②在泡沫分离过程中，通过BBP标签强化酶在气-液界面的定向吸附，解决无捕收剂时泡沫分离效率和选择性不高，以及酶在界面容易聚集失活的问题；③在低温凝聚相分离过程中，通过RLP标签赋予酶上临界溶解温度特性，解决非温敏特性酶直接从消泡液中快速分离的难题。相关科学问题的解决可以为生物制品大规模分离纯化技术的突破，以及新技术的应用提供理论指导。

Industrial enzymes are the core of bio-manufacturing, and the breakthrough of the large-scale separation and purification technology for microbial enzymes highly corresponds to the need of national strategy. This research is industrial application-oriented to develop an integrated technology of “foam separation & cold coacervation-phase separation” that is suitable for scale production, and a kind of multifunctional fusion enzyme is to be constructed to solve the bottleneck problems of the downstream separation and purification processes. The main research contents are as follows. First, two types of two-phase separation methods based on different separation mechanism are integrated and optimized, and the selective separation and purification of the target enzyme can be realized without the addition of chemical reagents, solving the problems of low yield, activity loss and high production cost induced by the traditional separation and purification process. Secondly, in the process of foam separation, the oriented adsorption of enzyme can be enhanced on the gas-liquid interface by the BBP-tag to solve the problems of low foam separation efficiency and selectivity, as well as the tendency of enzyme molecules to aggregate and deactivate at the interface. Thirdly, in the process of cold coacervation-phase separation, the target enzyme can be vested with an upper critical solution temperature characteristic of the RLP-tag, and so the problem of the direct and fast separation of enzyme without inherent thermosensitivity from defoaming liquid can be solved. The settlement of the related scientific problems can provide the theoretical guidance for the breakthrough of large-scale separation and purification technology of biological products and the application of new technology.