固定化产酸酶的微环境pH变化规律、调控机制及两阶段催化策略的研究

项目名称： 固定化产酸酶的微环境pH变化规律、调控机制及两阶段催化策略的研究

项目编号： No.21476025

项目类型： 面上项目

立项/批准年度： 2015

项目学科： 有机化学

项目作者： 罗晖

作者单位： 北京科技大学

项目金额： 80万元

中文摘要： 固定化酶具有稳定性好、可重复使用、易与产物分离等优点，但通常存在着颗粒内部的传质阻力问题，尤其是对催化过程中pH发生变化的酶（主要是产酸酶），颗粒内部的传质阻力会导致微环境pH发生剧烈变化，对固定化酶的催化效率和稳定性都可能会造成不利影响，甚至限制了一些酶的工业应用。本课题针对固定化产酸酶在催化过程中微环境pH剧烈变化的问题，研究不同调控策略下的固定化酶微环境pH变化规律，并结合酶催化特性和稳定性数据建立相关的数学模型。在前期研究中发现，由于受微环境pH变化和底物/产物浓度影响，固定化产酸酶的催化过程普遍存在着反应前期稳定性差和后期催化效率低的问题。对此，本课题提出两阶段催化调控策略：通过一定手段适当减缓初期反应速度以提高酶的稳定性，加快后期反应速度以缩短催化总反应时间。在理论研究的基础上，本课题还将对2~3种有重要应用前景的产酸酶的固定化酶催化模式进行研究，以推动其在工业上的应用。

中文关键词： 生物催化；固定化酶；微环境pH；调控机制；两阶段催化

英文摘要： Immobilized enzymes offer considerable advantages such as facility of removal and reuse, increased shelf life and thermal stability. Unfortunately, most immobilized enzyme catalysts are porous and the internal diffusional resistance in particles remarkably affects the catalytic efficiency. For these enzymes catalyzing a proton-producing or consuming reaction, the internal diffusional resistance would result in a severe microenvironmental pH change in the immobilized enzymes, sometimes leading to a low operational stability. It is undoubtedly important that the effect of microenvironmental pH changes on the activity and stability of immobilized enzymes being investigated. In this proposal, the change of microenvironmental pH is discussed in the catalysis of immobilized enzymes by introducing a variety of regulation methods. A model describing the relationship between microenvironmental pH change and the activity and stability of the immobilized enzymes will be constructed. It was found in our previous work that the initial rate of the immobilisates catalyzed reaction is usually very high, which might result in a low stability, largely owing to the sharp intraparticle pH gradient. While in the late period of the catalysis, the biocatalyst shows a very low activity due to the low substrate concentration and high product accumulation. Accordingly, a novel strategy of two-stage regulated catalysis is developed to realize both high performance and operational stability of the biocatalyst. In the optimized biotransformation, the reaction rate of the first stage would be down regulated to preserve the enzymatic activity, and some methods would be used in the second stage to shorten the catalyzing duration. Furthermore, two or three enzymes with high potential application in industry (such as cephalosporin C acylase, acetyl esterase and nitrilase) would be used as the research objective to develop their catalyzing process. The results to be obtained in this work are critical for better understanding the relationship between microenvironmental pH change, efficiency and stability of the immobilized enzymes in proton-producing reactions. A guideline for selection of proper regulation protocols for an optimum catalysis of immobilized enzymes will be developed, and surely the output of this work will impel a more extensive and successful application of the immobilized enzymes in industry.

英文关键词： biocatalysis;immobilized enzyme;microenvironmental pH;regulation stratagy;two-stage catalysis