I-Corps: Catalytic Artificial Self-Assemblies for the Biocatalytic Production of Small Molecules

I-Corps项目的更广泛影响/商业潜力是开发一类合成细胞，可以取代化学生产的传统生物催化过程。目前，实现小分子生物催化主要有两种方法：全细胞催化和无细胞催化，这两种方法都有各自的优势和挑战。全细胞催化在生产指标上受到毒素积累的限制。相比之下，无细胞系统可以支持更高的生产指标，但会受到酶降解的影响，这使得复杂化学品的生产变得困难，在经济上也不可行。提出的技术为缓解这些挑战提供了一条中间途径，并可用于生产目前由传统生物催化方法制造的小分子，如食品添加剂和香料，药物前体和生物燃料。例如，提出的技术可用于从木质纤维素中生产异丁醇，这被认为是下一代生物燃料。木质纤维素是最大的天然原料，而且不是从食物中提取的，这消除了人们对生物燃料与粮食生产竞争的担忧。与传统的乙醇生物燃料相比，异丁醇可以与汽油以更高的浓度混合，并直接用于现有的石油基础设施。由于与微生物相比，该技术的耐受性提高，预计将从糖化木质纤维素浓度中获得95%的异丁醇产量，并将温室气体排放量降低70%，这是传统方法无法实现的。这个I-Corps项目是基于胶体材料的开发，称为催化人工自组装（CASA），这是一种用于生物催化生产小分子的合成细胞。提出的技术使用复杂的凝聚原细胞制备和稳定使用低成本，市售聚合物。简化的复杂凝聚体原始细胞已被证明在其接近天然环境中保存酶，同时仍然提供无细胞系统的灵活性。复杂凝聚微滴乳剂的原细胞可将酶促反应速率提高25倍，并提供酶的长期稳定性（~4个月）以及细胞无法获得的加工灵活性。除了表现出改进的反应指标外，CASA对环境扰动具有鲁棒性，并克服了全细胞系统中细胞毒性和无细胞系统中酶稳定性的关键挑战。这可能允许更灵活和经济的生物反应器设计和扩大这些过程，其中提议的平台可以用作独立的方法或集成到现有的工业管道中，以降低化学品生产的成本。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

The broader impact/commercial potential of this I-Corps project is the development of a class of synthetic cells that can replace conventional biocatalytic processes for chemical production. Currently, biocatalysis of small molecules is implemented by two main methods: whole-cell catalysis and cell-free catalysis, both of which have their advantages and challenges. Whole-cell catalysis is limited in production metrics by the accumulation of toxins. In contrast, cell-free systems can support higher production metrics but suffer from enzyme degradation, which makes the manufacturing of complex chemicals difficult and economically unfeasible. The proposed technology provides an intermediate route to alleviate these challenges, and may be used to produce small molecules currently manufactured from conventional biocatalytic means, such as food additives and fragrances, drug precursors, and biofuels. For example, the proposed technology may be used to produce isobutanol from lignocellulose, which is considered the next generation of biofuels. Lignocellulose is the largest naturally available feedstock and is not derived from food sources, eliminating concerns about biofuel competition with food production. Compared with conventional ethanol biofuels, isobutanol may be blended with gasoline at higher concentrations and used directly in the existing petroleum infrastructure. The proposed technology is expected to achieve a 95% isobutanol yield from saccharified lignocellulose concentrations due to its improved tolerance compared to microbes, and lower greenhouse gas emissions by 70%, which cannot be achieved by conventional methods.This I-Corps project is based on the development of colloidal materials, called catalytic artificial self-assemblies (CASA), which are synthetic cells for the biocatalytic production of small molecules. The proposed technology uses complex coacervate protocells prepared and stabilized using low-cost, commercially available polymers. Simplified complex coacervate protocells have been shown to preserve enzymes in their near-native environments while still providing the flexibility of cell-free systems. Protocells of complex coacervate microdroplet emulsions improve enzymatic reaction rates by up to 25-fold and provide long-term stability (~4 months) to enzymes as well as processing flexibility not accessible in cells. In addition to showing improved reaction metrics, CASA is robust to environmental perturbations and overcomes key challenges concerning cell toxicity in whole cell systems, and enzyme stability in cell-free systems. This may allow a more flexible and economical bioreactor design and scaling up of these processes where the proposed platform may be used as a standalone method or integrated into existing industrial pipelines to reduce the cost of chemical production.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.