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

这个小企业创新研究（SBIR）项目的更广泛的影响/商业潜力包括白色生物技术领域的进步，该领域利用酶来创造有价值的工业产品。作为生物分子，酶比传统化学品更难使用，在它们适用于工业或制药生产之前通常需要更广泛的开发。该SBIR项目将展示如何使用合成聚合物稳定酶的性能。酶的特点是精确的，独特的结构和功能，这反过来又是必不可少的，它们在催化复杂的化学反应的作用。另一方面，合成聚合物尽管结构不太精确，但可以合理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导致先进的纳米装甲酶具有改进的性能，如耐溶剂性和耐温性，以及调节的活性。创造这种新型的稳定化酶将导致酶催化的更有效的商业利用，其需要更少的能量，使用更少的危险试剂，并产生更少的废物，同时产生有价值的产品，如化学品，生物燃料和药物。SBIR第一阶段项目提出开发一种组合合成装置，可以高通量筛选具有所需性质的酶-聚合物缀合物（例如，温度、pH或有机溶剂稳定性）。到目前为止，只有低通量的合成和表征方法已被应用到酶-聚合物缀合物的制备，限制了发展，只有少数类型的聚合物修饰每个蛋白质和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聚合物的不同集合，必须考虑缩放最佳性能的酶-聚合物缀合物的鉴定的方法。这将通过酶-聚合物缀合物的高通量合成和获得的性质的高通量筛选的组合来实现。该研究的初步目标应用集中在工业生物催化上。高通量方法的应用不仅会导致更快的研究和开发周期，而且还将加速我们对通过聚合物修饰可以获得何种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.