UNS: Engineered Protein-Inorganic Self-Assembly to Control Enzyme Performance and Recovery

小行星1510551 酶在各种载体结构上的固定化用于提高酶的稳定性、活性、回收率和经济性。 这项工作的目标是创造一种新的酶固定化模式，每体积含有高水平的酶，最重要的是，保持或提高酶的活性。通过在生物相容性材料存在下使用蛋白质自组织，将产生含有活性酶的多孔、高表面积颗粒。作为测试情况，产生药物前体的酶及其再生伴侣酶都将固定在颗粒中。将检测颗粒的产品生产、随时间推移的稳定性以及在多个反应循环中收集和重复使用的能力。这项研究的结果将不仅是一种改进的酶固定化方法，但将产生如何在固定化结构的伴侣酶的位置可以提高其性能和生产力的基础知识。这里开发的颗粒平台是模块化的，可以很容易地应用于制造重要产品的各种酶。PI将把这项研究与中学到研究生院的活动结合起来，以招募和留住STEM领域的女性。这项研究的目标是创造生物催化，自组装材料，能够提供所需的微米和纳米结构环境，以支持和增强工业相关应用的酶活性。传统上，酶通过共价相互作用或吸附固定在固体合成载体上，以提高活性、回收率和/或寿命。在这里，酶被设计成与互补蛋白质构建块组装，以创建生物催化材料。这提供了一个局部蛋白质拥挤的环境，并消除了对共价固定或非特异性吸附的需要，共价固定或非特异性吸附通常会导致结构和功能的损失，并缺乏对特定酶位置或密度的控制。该项目结合了蛋白质工程，生产含有酶和自组装结构域的蛋白质，以及蛋白质-无机自组装，以构建适当的结构来控制酶的性能和回收。自组装首先发生在纳米级，然后发生在微米级，以产生具有内部结构复杂性的分级结构的超颗粒，以实现高表面积和孔隙率。该研究将为蛋白质-无机自组装与双酶耦合体系的概念验证提供基础知识，并用于辅助因子再生手性胺的生产。所采用的设计策略是模块化的，因此可以在相同的超颗粒中同时使用多种酶，均匀分散或空间分离。模块化设计还设置了与酶的物理性质无关的材料结构，并且允许使用几乎任何酶。将评估超微粒固定化对酶活性和协同作用的影响，并将提供该系统在工业反应过程中使用的实用性的实际评估。此外，这项工作还产生了关于蛋白质与无机组分自组装的基础知识，以及关于酶位置的空间控制如何控制动力学参数和活性的见解。这项工作将证明酶的蛋白质-无机自组装可以提供一个通用的模板，以及所需的化学和物理环境，以支持复杂的工业生物催化。这项由CBET部门生物技术和生物化学工程计划授予的奖项由材料研究部门生物材料计划共同资助。

1510551 Champion, Julie Immobilization of enzymes on various support structures is used to enhance enzyme stability, activity, recovery, and economics. The goal of this work is to create a new mode of enzyme immobilization that contains high enzyme levels per volume, and most importantly, retains or improves enzyme activity. Through the use of protein self-organization in the presence of biocompatible materials, a porous, high surface area particle containing active enzyme will be created. As a test case, an enzyme that produces pharmaceutical precursors and its regenerating partner enzyme will both be immobilized in the particles. The particles will be tested for production of product, stability over time, and the ability to collect and reuse over multiple reaction cycles. The results of this research will not only be an improved method of enzyme immobilization, but will generate fundamental knowledge of how placement of partner enzymes in immobilized structures can improve their performance and productivity. The particle platform developed here is modular and can be easily applied to a wide variety of enzymes that make important products. The PIs will integrate the research with activities for middle to graduate school to recruit and retain women in STEM.The goal of this research is to create bio-catalytic, self-assembled materials capable of providing the required micro and nano-structured environment to support and enhance enzymatic activity for industrially relevant applications. Traditionally, enzymes are immobilized on solid synthetic supports via covalent interactions or adsorption in order to enhance activity, recovery, and/or lifetime. Here, enzymes are engineered to assemble with complementary protein building blocks to create biocatalytic materials. This provides a local protein-crowded environment and eliminates the need for covalent immobilization or non-specific adsorption which often leads to loss of structure and function, and lack control over specific enzyme placement or density. This project combines protein engineering, to produce proteins containing both enzymatic and self-assembly domains, and protein-inorganic self-assembly, to build appropriate structures to control enzyme performance and recovery. Self-assembly occurs first at the nanoscale and then at the microscale to produce hierarchically structured supraparticles that have internal structural complexity to achieve high surface area and porosity. The proposed research will provide basic knowledge and proof of concept of protein-inorganic self-assembly with a coupled two enzyme system for production of chiral amines with co-factor regeneration. The employed design strategy is modular, so that multiple enzymes can be used simultaneously in the same supraparticles, either dispersed homogenously or segregated spatially. The modular design also sets the material structure independently of the physical properties of the enzyme(s), and allows almost any enzyme(s) to be used. The impact of supraparticle immobilization on enzyme activity and synergy will be evaluated, and also a practical assessment of the utility of this system to be used in industrial reaction processes will be provided. Additionally, this work also generates fundamental knowledge on protein self-assembly with inorganic components and insight as to how spatial control of enzyme placement can give control of kinetic parameters and activity. This work will demonstrate that protein-inorganic self-assembly of enzymes can provide a general template, as well as the required chemical and physical environment, to support complex industrial biocatalysis.This award by the Biotechnology and Biochemical Engineering Program of the CBET Division is co-funded by the Biomaterials Program of the Division of Materials Research.