Collaborative Research: Engineering of Recoverable Cellulosomes for Bioconversion

可持续的、工业规模的从植物生物质中生产运输生物燃料依赖于一种叫做纤维素酶的酶，这种酶能将生物质的纤维素部分分解成糖，然后发酵成生物乙醇。这一过程的主要成本是纤维素酶，在当前一代的纤维素生物燃料设施中，纤维素酶在使用后被丢弃，因为它必须溶解在水中才能将纤维素分解成糖，并且不易回收。回收纤维素酶将降低生产纤维素生物燃料的成本。该项目的目标是开发一种将纤维素酶封装成可回收固体形式的新方法，以便在处理前多次收集和再利用。关键的创新是将纤维素酶结合到一种纳米大小的胶囊上，这种胶囊被称为纤维素体，它保留了纤维素转化酶的活性，并且有一个磁芯，有助于从稀释的水处理流中回收纤维素。与此项目相关的教育活动包括通过北达科他州立大学的培育美国部落本科生研究与教育（NATURE）项目协调指导本科生项目。自然产生纤维素酶的微生物将它们结合到细胞表面称为纤维素体的特殊结构中。由于这些酶在纤维素体中的接近性，纤维素和半纤维素材料的生物转化以高速度和高效率进行。然而，在纤维素酶的工业生产中，不存在纤维素体。这项研究将通过一种仿生方法来设计可回收的纤维素体，其中纤维素酶被封装在含有磁芯的纳米结构胶囊中。这种仿生纤维素体将包含许多限制在聚合物环境中的互补纤维素酶，并将被设计成模仿天然纤维素体的几个关键特性，包括水解活性的多样性，互补酶的接近性，以及仿生纤维素体与纤维素底物的强选择性结合。仿生纤维素的磁芯将有利于其从液体悬浮液中利用磁分离技术进行回收。为了探索仿生纤维素在生物质转化过程中的潜力，该研究有四个目标。第一个目标是制造一系列新的仿生纤维素，以增强水解酶的协同作用，将纤维素材料生物转化为糖。第二个目标是量化仿生纤维素体结构和固定化酶多样性对纤维素水解的影响，以获得对天然纤维素体中纤维素酶协同作用的基本理解。第三个目标是获得对纤维素体与纤维素底物和木质素相互作用的基本理解，第四个目标是增强仿生纤维素体的功能，以获得更广泛的应用。这些研究的预期结果包括对酶在工程纤维素体中的协同作用的基本理解，它们对不同形式底物的功效，以及它们的回收和再利用潜力。

The sustainable, industrial-scale production of transportation biofuels from plant biomass rely on enzymes called cellulases that break down the cellulosic fraction of biomass into sugars that can be fermented into bioethanol. The major cost of this process is the cellulase, which in the current generation of cellulosic biofuel facilities, is discarded after use because it must be dissolved in water to break down cellulose to sugars, and cannot be easily recovered. Recycling the cellulase enzyme would reduce the cost of cellulosic biofuels production. The goal of this project is to develop a new way of encapsulating the cellulase enzyme in recoverable solid form so that it can be collected reused multiple times before disposal. The key innovation is to bind the cellulase enzyme to nanometer-sized capsule called a cellulosome, which preserves the activity of the enzyme for cellulose conversion, and has a magnetic core which facilitates its recovery from dilute water processing streams. The educational activities associated with this project include mentoring of undergraduate student projects coordinated through the Nurturing American Tribal Undergraduate Research and Education (NATURE) program at North Dakota State University.Microorganisms that naturally produce cellulase enzymes bind them into special structures on the cell surface called cellulosomes. Due to the close proximity of these enzymes within the cellulosomes, the bioconversion of cellulosic and hemicellulosic materials proceeds with a high velocity and efficiency. However, in the industrial production of celluloytic enzymes, the cellulosome is not present. This research will engineer recoverable cellulosomes through a biomimetic approach where celluloytic enzymes are encapsulated within a nanstructured capsule containing a magnetic core. This biomimetic cellulosome will contain a number of complimentary celluloytic enzymes confined within a polymeric environment, and will be engineered to mimic several key properties of natural cellulosomes, including diversity of hydrolytic activity, close proximity of complementary enzymes, and strong and selective binding of the biomimetic cellulosome to cellulosic substrates. The magnetic core of the biomimetic cellulosome will facilitate its recovery from liquid suspension using magnetic separation techniques. To explore the potential of the biomimetic cellulosome for biomass conversion processes, the research has four objectives. The first objective is to make a series of new biomimetic cellulosomes to enhance synergism of hydrolytic enzymes for bioconversion of cellulosic materials to sugars. The second objective is to quantify the effects of the biomimetic cellulosome structure and immobilized enzyme diversity on cellulose hydrolysis to gain a fundamental understanding of cellulase synergism in natural cellulosomes. The third objective is to gain a fundamental understanding of cellulosome interaction with cellulosic substrates and lignin, and the fourth objective is to augment the functionality of the biomimetic cellulosome for broader applications. The anticipated outcomes from these studies include a fundamental understanding of the synergism of enzymes in engineered cellulosomes, their efficacy for different forms of substrates, and their potential for recovery and reuse.