New Catalysts by Cofactor Modulation

在化学系生命过程化学（CLP）项目的支持下，约翰霍普金斯大学的Steven Rokita教授正在研究基于含有天然辅助因子的合成类似物的蛋白质的新型催化剂的开发。蛋白质及其辅助因子之间的复合物在促进对我们自身新陈代谢至关重要的反应方面非常有效。然而，它们独特的特性阻碍了它们在工业上的广泛应用。通常可以对蛋白质进行工程改造，以增强它们在此类应用中的催化能力。辅助因子工程为蛋白质工程提供了一个强大的替代方案，但迄今为止很少受到关注。本项目将利用有机化学合成新的辅因子，结合其蛋白支架，应用于绿色化学和生物修复。该项目将提供跨越化学和生物化学的高级培训，并将为学生未来在酶优化和生物催化过程化学中的应用（可持续化学的新兴领域）做出贡献奠定基础。通过约翰霍普金斯大学化学系、生物物理系和生物系之间的合作，化学操纵生物系统的力量也将被引入与创建生物化学专业相关的新课程中。黄素依赖性硝基还原酶超家族的酶促进了广泛的化学转化，有望在工业生物催化过程中得到应用。尽管如此，对自然变异的研究和酶库的构建尚未从最具特征的硝基还原酶和碘酪氨酸脱碘酶亚群中产生令人满意的候选者。在这个项目下，罗基塔实验室的研究将转而关注通过修改促进催化所必需的辅助因子来控制特异性和化学。黄素结构的微小变化可以引起催化化学的巨大变化，例如5-去氮黄素和6-羧基黄素。例如，天然黄素的还原能力不足以使硝基芳烃完全还原为胺类产物，但含有5-去氮黄素的硝基还原酶应该具有足够的还原能力来产生胺类产物。作为补充，6-羧基黄素形成比天然辅因子更稳定的半醌中间体的能力将被整合到还原性脱卤化学的生物催化剂开发中。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。

With the support of the Chemistry of Life Processes (CLP) program in the Division of Chemistry, Professor Steven Rokita from Johns Hopkins University is studying the development of new catalysts based on proteins containing synthetic analogues of natural cofactors. Complexes between proteins and their cofactors are highly efficient at promoting reactions that are fundamental to our own metabolism. However, their exquisite specificity precludes their widespread application for industry. Proteins can often be engineered to enhance their catalytic repertoire for such applications. Cofactor engineering offers a powerful alternative to protein engineering but has received little attention to date. This project will use organic chemistry to synthesize new cofactors that, in combination with their protein scaffolds, may be applied to green chemistry and bioremediation. The project will provide advanced training, spanning chemistry and biochemistry, and will provide students with a foundation for future contributions to enzyme optimization and application in biocatalytic process chemistry, an emerging area in sustainable chemistry. The power of chemistry to manipulate biological systems will also be introduced into new courses associated with the creation of a biochemistry major through a collaboration between the Departments of Chemistry, Biophysics, and Biology at Johns Hopkins University. Enzymes of the flavin-dependent nitroreductase superfamily promote a wide range of chemical transformations with promise for deployment in industrial biocatalytic processes. Still, the search for natural variants and the construction of enzyme libraries has yet to produce satisfactory candidates from the best characterized subgroups of nitroreductases and iodotyrosine deiodinases. Under this project, research in the Rokita laboratory will instead focus on controlling specificity and chemistry by modifying the cofactors necessary to promote catalysis. Only minor changes to the flavin structure can engender large changes in catalytic chemistry as illustrated by examples of 5-deazaflavin and 6-carboxyflavin. The reducing power of native flavin is not sufficient to drive the full reduction of nitroaromatics to their amine products, but a nitroreductase containing 5-deazaflavin should have sufficient reducing capacity to produce the amines, for example. In a complementary aim, the ability of 6-carboxyflavin to form a more stable semiquinone intermediate than the native cofactor will be integrated into biocatalyst development for reductive dehalogenation chemistry.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.