SBIR Phase I: Alcohol Resistant Enzymes through High-Throughput Combinatorial Protein-Polymer Conjugate Synthesis

这项小型企业创新研究（SBIR）项目的更广泛影响/商业潜力包括推进白色生物技术领域，该领域利用酶来创造有价值的工业产品。作为一种生物分子，酶比传统的化学物质更难处理，通常需要更广泛的开发才能适应工业或制药生产。这个SBIR项目将展示如何使用合成聚合物稳定化来改善酶的性能。酶具有精确、独特的结构和功能，这对它们在复杂化学反应中的催化作用至关重要。另一方面，合成聚合物，尽管结构不太精确，但可以合理地设计以承受或响应化学，热或生物条件。酶和合成聚合物的协同融合产生了先进的纳米装甲酶，具有改善的性能，如耐溶剂和耐温性，以及可调节的活性。创造这种新型稳定酶将导致酶催化更有效的商业利用，它需要更少的能量，使用更少的危险试剂，产生更少的废物，同时产生有价值的产品，如化学品，生物燃料和药品。这个SBIR一期项目建议开发一种组合合成设备，该设备可以提供高通量筛选具有所需性能（例如，温度，pH或有机溶剂稳定性）的酶-聚合物偶联物。迄今为止，只有低通量的合成和表征方法被应用于酶-聚合物偶联物的制备，限制了每个蛋白质的聚合物修饰类型的发展，并且依赖于随机猜测来选择测试的变体。因此，为了充分受益于目前市场上可获得的各种聚合物，人们必须考虑确定最佳性能酶-聚合物偶联物的方法。这将通过高通量合成酶-聚合物缀合物和高通量筛选所获得的性能来实现。本研究的初步目标应用是工业生物催化。高通量方法的应用不仅会加快研发周期，而且会加速我们对通过聚合物改性可以获得什么样的蛋白质性质的基础知识的发展，从而为工业应用建立这种方法。

The broader impact/commercial potential of this Small Business Innovation Research (SBIR) project includes advancement of the field of white biotechnology, which utilizes enzymes to create valuable industrial products. As biological molecules, enzymes are more difficult to work with than conventional chemicals and often need more extensive development before they can be adapted to industrial or pharmaceutical manufacturing. This SBIR project will demonstrate how enzymes performance can be improved using stabilization with synthetic polymers. Enzymes are characterized by precise, unique structure and function, which is in turn essential for their role in catalysis of complex chemical reactions. Synthetic polymers, on the other hand, despite being less precisely structured, can be rationally designed to withstand or respond to chemical, thermal or biological conditions. The synergistic fusion of enzymes and synthetic polymers results in advanced nano-armored enzyme with improved properties such as solvent and temperature resistance, and modulated activity. Creating such novel stabilized enzymes will result in more efficient commercial utilization of enzymatic catalysis which requires less energy, utilizes less hazardous reagents, and generates less waste while generating valuable products such as chemicals, biofuels, and pharmaceuticals.This SBIR Phase I project proposes to develop a combinatorial synthesis device that can feed high-throughput screening of enzyme-polymer conjugates with desired properties (for instance, temperature, pH- or organic solvent stability). To date, only low-throughput synthesis and characterization methods have been applied to the preparation of enzyme-polymer conjugates, limiting development to only few types of polymer modification per protein and depending on stochastic guesswork to select the variants tested. Thus, in order to fully benefit from the diverse set of polymers currently available on the market one has to consider methods of scaling the identification of optimally performing enzyme-polymer conjugates. This will be achieved through combination of high-throughput synthesis of enzyme-polymer conjugates and high-throughput screening of gained properties. The initial target application of the proposed research is focused on the industrial biocatalysis. Application of a high-throughput method will not only result in faster research and development cycles, but also will accelerate our development of fundamental knowledge of what kind of protein properties can be gained through polymer modification, thereby establishing this method for industrial applications.