CAREER: Towards the Rational Control of Redox Potential and Catalytic Activity of Designed Functional Membrane Proteins

在化学学部有机动力学项目的支持下，亚利桑那州立大学的Giovanna Ghirlanda教授将设计和研究具有可调特性的新型功能膜蛋白。作为这个方向的第一步，血红素结合位点已经被设计成一种天然的膜蛋白，糖蛋白a，它通过其跨膜螺旋结构域的结合形成稳定的二聚体。最初设计的ME1以高亲和力结合铁原卟啉IX，在天然膜血红素结合蛋白范围内表现出氧化还原电位，并显示出初步的催化活性。在资助期间，该系统将被进一步研究，并作为设计大型突变文库的模板，明确设计用于优化血红素结合和控制氧化还原电位。该文库将用于分析血红素环境的重要性，其疏水性和芳香残基的存在，以确定ME1支架内的中点电位和结合亲和力。在这些结果的基础上，ME1将进化成一种具有单氧化酶和过氧化物酶活性的人工酶。有机和大分子化学项目支持亚利桑那州立大学的Giovanna Ghirlanda教授，他对膜相关蛋白的研究是生物传感器和生物催化剂应用的一个相对未开发的资源。对膜环境中蛋白质结构和功能之间关系的更深入理解将允许制备适合特定生物和非生物应用的人工小型化蛋白质。Ghirlanda博士将与亚利桑那州立大学现有的一个项目合作，实施一种综合的方法，将K-12学校的课堂演示、本科培训和专门的教师研讨会结合起来。参加这项服务学习实习和推广计划的本科生将在PI的实验室接受培训，进行课堂实验，旨在激发学生对生物技术的兴趣，并以简单但科学合理的方式解释基本概念。该项目旨在让来自不同背景的K-12学生接触大学经历，并通过促进与大学生的接触提供榜样。

With the support of the Organic Dynamics Program in the Chemistry Division, Professor Giovanna Ghirlanda of Arizona State University will design and study novel functional membrane proteins with tunable properties. As a first step in this direction, a heme-binding site has been engineered into a natural membrane protein, Glycophorin A, which forms stable dimers by the association of its transmembrane helical domain. The initial design, ME1, binds ferric protoporphyrin IX with high affinity, exhibits a redox potential within the range of natural membrane heme-binding proteins, and shows nascent catalytic activity. During the funding period this system will be further investigated and serve as the template for the design of a large library of mutants explicitly designed to optimize heme binding and controlling the redox potential. The library will be used to analyze the importance of the heme environment, its hydrophobicity and the presence of aromatic residues in determining the midpoint potential and the binding affinity within the context of the ME1 scaffold. Building on these results ME1 will be evolved into an artificial enzyme for monooxigenase and peroxidase reactivity.The Organic and Macromolecular Chemistry Program supports Professor Giovanna Ghirlanda of Arizona State University whose research on membrane-associated proteins is a relatively untapped resource for applications as biosensors and biocatalysts. A deeper understanding of the relationship between protein structure and function in the membrane environment will permit the preparation of artificial miniaturized proteins tailored to specific biological and non-biological applications. Dr. Ghirlanda will partner with an existing program at ASU to implement a comprehensive approach that integrates classroom demonstrations in K-12 schools, undergraduate training, and specialized workshops for teachers. Undergraduates participating in this service learning internship and outreach program will be trained in the PI's lab to perform classroom experiments designed to spark an interest in biotechnology and to explain the underlying concepts in a simple but scientifically sound manner. The program aims at exposing K-12 students from diverse backgrounds to the college experience and providing role models by facilitating contact with college students.