Collaborative Research: Engineering of Recoverable Cellulosomes for Bioconversion

从植物生物质中可持续地、工业化地生产运输生物燃料依赖于一种名为纤维素酶的酶，这种酶能将生物质中的纤维素部分分解成糖，这些糖可以发酵成生物乙醇。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将纤维素分解为糖，并且不容易回收。回收纤维素酶将降低生产纤维素生物燃料的成本。本项目的目标是开发一种新的方法，将纤维素酶以可回收的固体形式包裹起来，以便在处置之前可以多次收集和重复使用。关键的创新是将纤维素酶结合到名为纤维素体的纳米胶囊上，该胶囊保留了纤维素酶的活性以进行纤维素转化，并具有一个磁芯，便于从稀释水处理流中回收纤维素酶。与这个项目相关的教育活动包括指导本科生项目，通过北达科他州立大学的培育美国部落本科生研究和教育(自然)计划进行协调。自然产生纤维素酶的微生物将它们结合到细胞表面的特殊结构中，称为纤维素体。由于这些酶在纤维素体中非常接近，纤维素和半纤维素材料的生物转化速度和效率都很高。然而，在纤维素酶的工业化生产中，纤维素体是不存在的。这项研究将通过仿生方法设计可回收的纤维素体，将纤维素酶封装在含有磁芯的纳米结构胶囊中。这种仿生纤维素体将在聚合物环境中包含许多免费的纤维素酶，并将被设计成模拟天然纤维素体的几个关键性质，包括水解性的多样性，互补酶的紧密接近，以及仿生纤维素体与纤维底物的强烈和选择性结合。仿生纤维素体的磁芯将有助于利用磁分离技术从液体悬浮液中恢复。为了探索仿生纤维素体在生物质转化过程中的潜力，本研究有四个目标。第一个目标是制造一系列新的仿生纤维素体，以增强纤维素类物质生物转化为糖的水解酶的协同作用。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酶降解的影响，以期从根本上了解纤维素酶在天然纤维素体中的协同作用。第三个目标是对纤维素体与纤维素底物和木质素的相互作用有一个基本的了解，第四个目标是增强仿生纤维素体的功能，使其具有更广泛的应用。这些研究的预期结果包括对工程纤维素体中酶的协同作用的基本了解，它们对不同形式底物的有效性，以及它们回收和再利用的潜力。

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.