Extending the catalytic activity of enzymes to unnatural reactions

酶工程为可持续的化学合成创造了有用的催化剂，它也展示了对酶如何工作的理解。在这个项目中，蛋白质运动将被建模来识别赋予新能力的酶突变。这些突变将被实验评估以验证模型。将招募未被充分代表的少数民族学生参与该项目。我们将开发免费的蛋白质工程教材，以及蛋白质工程的网络工具。如果成功，该项目将使酶更广泛地应用于工业应用。蛋白质工程师目前无法设计出高效的酶。这个项目的重点是将天然酶的催化能力扩展到非自然反应的简单问题。这允许更精确的计算方法来测试假设，远残基有助于催化通过精确定位底物和催化基团。通过广泛的实验数据验证计算，将创建更现实的酶如何工作的模型。模型系统将是酯酶向羟基腈裂解酶的演化。这些酶之间交换其催化活性所需的氨基酸变化的最小数量将被确定。计算预测将通过构建酶变体和测量它们对催化的贡献进行实验测试。这些精细的计算模型接下来将用于预测哪些取代将把羟基腈裂解酶的催化活性扩展到硝基醇加成反应，这是一种非自然反应。对科学和工程的潜在贡献是第一个预测活性位点外残基如何促进催化的计算方法，以及第一个针对非自然反应的高效酶的合理设计。这些能力将使生物催化合成药物和精细化学品成为可能。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。

Engineering of enzymes creates useful catalysts for sustainable chemical synthesis, and it also demonstrates an understanding of how enzymes work. In this project, protein motion will be modeled to identify enzyme mutations that impart new abilities. These mutations will be evaluated experimentally to validate the model. Underrepresented minority students will be recruited to work on the project. A free textbook for protein engineering will be developed as well as web tools for protein engineering. If successful, the project will enable the broader application of enzymes for industrial applications. Protein engineers cannot currently design efficient enzymes. This project focuses on the simpler problem of extending a natural enzyme's catalytic abilities to unnatural reactions. This allows more accurate computational approaches to test the hypothesis that distant residues contribute to catalysis by precisely positioning the substrates and catalytic groups. More realistic models of how enzymes work will be created by validating computation with extensive experimental data. The model system will be the evolution of esterases into hydroxynitrile lyases. The minimum number of amino acid changes between these enzymes that are needed to exchange their catalytic activities will be identified. Computational predictions will be tested experimentally by constructing enzyme variants and measuring their contributions to catalysis. These refined computational models will next be used to predict which substitutions will extend the catalytic activity of hydroxynitrile lyases to nitroaldol addition, an unnatural reaction. The potential contributions to science and engineering are the first computational approach to predict how residues outside the active site contribute to catalysis, and the first rational design of efficient enzymes for unnatural reactions. These abilities will enable biocatalytic synthesis of pharmaceuticals and fine chemicals.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.