RUI: Learning the Rules that Govern the Folding and Stability of Coiled Coils

本项目的目的是探索调节短肽聚合成α-螺旋聚合结构的因素。对这种生物聚合物过程的更好的理解将有助于推进许多不同的活跃研究领域，包括：(1)阐明蛋白质折叠的一般规则及其与聚合的关系；(2)了解这种过程如何导致错误折叠和聚集；以及(3)设计新的生物材料。这些螺旋聚合物的形成能力取决于螺旋肽之间设计的交错相互作用，螺旋肽的结构基于众所周知的盘绕结构基序。多肽序列将被突变，以测试疏水相互作用在聚合物组装和结构中的作用。还将进行突变，以测试金属结合对聚合的调节，以及一种能够引起局部结构变化的新型光敏开关。为该项目制作的一种特殊的序列变体被发现形成包含a-螺旋或b-链构象的聚合物。研究人员将测试谷氨酸在影响形成这两种聚合物结构的倾向方面所起的作用。利用圆二色谱和傅里叶变换红外光谱对聚合物的二级结构、含量和稳定性进行了定量研究。聚合物的大小和形状将通过分析超速离心法、原子力显微镜和动态光散射进行跟踪。虽然该项目专注于基础研究问题，但也将影响实验室中的另一个项目，该项目的目标是创造生物电子材料，涉及哈弗福德和宾夕法尼亚大学的教职员工。这里描述的工作将主要由本科生推动，因为哈弗福德学院是一所本科院校。学生通过撰写论文提案和期末论文来完成主修课学分，在小组会议上展示他们的工作，并在进行实验的同时向整个系展示他们的工作。研究人员已经为化学和生物专业的初级实验室课程制定了这项研究计划的内容，并正在开发与生物材料设计相关的想法，以纳入化学生物学和生物化学的初级讲座课程。调查员长期致力于改善代表性不足群体获得研究的机会。除了长期致力于为来自费城的K-12学生提供科学和写作方面的现有推广计划外，PI还经常在夏天在他的研究实验室为至少一名自认为属于代表性不足群体的学生预留空间。在一个较新的方向上，这位研究人员还在去年夏天招待了一名来自费城的高中教师和学生到他的实验室工作，在那里他们学习了合成和提纯对这个项目目标至关重要的多肽。

The objective of this project is to explore factors that regulate the polymerization of short peptides into a-helical polymeric structures. An improved understanding of such biopolymer processes will contribute to the advancement of a number of different active fields of research, including: (1) elucidating general protein folding rules and how they relate to polymerization; (2) understanding how such processes can lead to misfolding and aggregation; and (3) designing novel biomaterials. The ability of these helical polymers to form depends on designed staggered interactions between helical peptides, whose structure is based on the well-known coiled-coil structural motif. The peptide sequence will be mutated to test the role of the hydrophobic interaction in polymer assembly and structure. Mutations will also be made to test regulation of polymerization by metal binding and a novel photosensitive switch capable of inducing local structural changes. One particular sequence variant made for this project was discovered to form polymers containing either a-helical or b-strand conformations. The investigator will test the role of glutamines in influencing the propensity to form these two polymer structures. Circular dichroism and Fourier transform infrared spectroscopy will be used to quantify secondary structure content and stability of the polymers. The size and shape of the polymers will be tracked by analytical ultracentrifugation, atomic force microscopy, and dynamic light scattering.Although focusing on basic research questions, this project will also impact a different project in the lab whose goal is to create bio-electronic materials, involving faculty at Haverford and at U. Pennsylvania. The work described here will be driven primarily by undergraduates, since Haverford College is an undergraduate institution. Students fulfill major course credit by writing a thesis proposal and a final thesis paper, presenting their work at group meetings and to the Department as a whole, while carrying out their experiments. Elements of this research program have been developed by the investigator for a junior laboratory course to be taught to chemistry and biology majors, and ideas relevant to biomaterial design are being developed for inclusion in a junior-level lecture course in chemical biology and biochemistry. The investigator has a long-standing commitment to improving access to research for under-represented groups. In addition to a long-term commitment to an existing outreach program for science and writing for K-12 students from Philadelphia, The PI regularly reserved space in his research lab over the summer for at least one student self-identified as belonging to an under-represented group. In a newer direction, the investigator has also hosted a high school teacher and student from Philadelphia to work in his lab this past summer, in which they learned to synthesize and purify peptides critical for the goals of this project.