Collaborative Research: Development of 2D IR Spectroscopy as a Quantitative Probe of Protein Structure, with Applications to Membrane and Aggregated Proteins

威斯康星大学麦迪逊分校的Martin Zanni教授和合作者James Skinner（威斯康星大学麦迪逊分校），Daniel Raleigh（纽约州立大学石溪分校）和Sean Decatur（奥伯林学院）在化学合作研究项目的支持下，设计和测试了基于二维红外（2D IR）光谱的方法，用于表征蛋白质的结构和功能。这些技术对于研究核磁共振和x射线晶体学难以应用的系统尤其重要，例如细胞膜中的蛋白质和蛋白质聚集体。这项工作汇集了一个多样化和互补的研究团队，通过理论、实验和合成的新组合，为从原子结构模型计算二维红外光谱制定了严格的协议。为了开发理论方法，在单个残基上进行同位素标记或突变，从而可以用二维红外光谱测量特定残基的线宽和/或偶联。虽然这项工作主要是针对开发二维红外作为蛋白质动力学的工具，但它也可以应用于其他系统。参与本研究的本科生、研究生和博士后正在接受培训，以便在跨学科和跨机构的协作环境中工作。代表性不足的群体的参与源于奥伯林大学、霍华德大学、石溪大学和威斯康星大学之间的交流项目。网络基础设施的组成部分包括讨论研究进展的网络会议和用于光谱理论计算的用户友好软件的传播。一个本科物理化学实验室正在进行实验，以展示如何将FTIR光谱用作蛋白质二级结构测定的工具；该实验将在威斯康星大学、石溪大学、奥伯林大学和霍华德大学的物理化学实验课程中进行测试，然后提供给更广泛的学术界。缺乏所需仪器的机构将能够利用正在开发的基于网络的远程访问功能，将这个实验纳入他们的课程。

Prof. Martin Zanni at the University of Wisconsin in Madison, and collaborators James Skinner (UW-Madison), Daniel Raleigh (SUNY at Stony Brook), and Sean Decatur (Oberlin College) are supported by the Collaborative Research in Chemistry Program to devise and test methods based on two-dimensional infrared (2D IR) spectroscopy for characterization of protein structure and function. The techniques are particularly important for studying systems for which NMR and x-ray crystallography are difficult to apply, such as proteins in cell membranes and protein aggregates. The work brings together a diverse and complementary team of researchers to develop a rigorous protocol for calculating 2D IR spectra from atomistic structural models through a novel combination of theory, experiment, and synthesis. To develop the theoretical methodology, well-characterized proteins are either isotope-labeled or mutated at individual residues so that the linewidths and/or couplings of specific residues can be measured with 2D IR spectroscopy. While the work is primarily directed toward developing 2D IR as a tool for protein kinetics, it can be applied to other systems as well. Undergraduate and graduate students and postdocs involved in this study are being trained to work in a collaborative interdisciplinary and inter-institutional environment. Participation of underrepresented groups derives from exchange programs among Oberlin, Howard University, Stony Brook, and Wisconsin. Cyber-infrastructure components include web-conferences to discuss research progress and dissemination of user-friendly software for the theoretical calculation of spectra. An undergraduate physical chemistry lab experiment is being developed to show how FTIR spectroscopy can be used as a tool for protein secondary structure determination; the experiment will be tested in the physical chemistry lab courses at Wisconsin, Stony Brook, Oberlin, and Howard, and then made available to the broader academic community. Institutions that lack the needed instrumentation will be able to incorporate this experiment into their curriculum by taking advantage of the web-based remote access features being developed.