Regulatory Protein-DNA Interactions in vivo Analyzed by Ultrafast Photochemical Crosslinking

康奈尔大学应用和工程物理学院的Watt W.Webb，分子生物学和遗传学系的John T.Lis，化学和化学生物学系的Tadhg P.Begley，分子生物学和遗传学系的Bik-quon Tye，分子生物学和遗传学系的Eric E.Alani和应用和工程物理学院的Alexander L.Gaeta，都是由化学合作研究计划资助的，资金来自化学系、物理系、分子细胞生物学系和多学科活动办公室。这个多方面的团队将使用超快的紫外线激光交联，在细胞周期中的功能激活过程中，瞬间并共价地将蛋白质锁定到它们的核酸目标上。然后，通过最先进的分析蛋白质和核酸生物化学和结构分析来解析这些分子复合体，以确定交联型氨基酸及其核苷酸结合部位。这些研究人员将利用新技术研究调控基因转录、复制和修复的复杂DNA-蛋白质相互作用网络，这些新技术允许对活细胞中难以捉摸的DNA-蛋白质相互作用进行高时间和空间分辨率的生化分析。这项研究的最终目标是发展在原核和真核系统的活细胞上进行功能基因组实验的方法，以建立DNA结合蛋白的全基因组图谱，从而跟踪细胞生长周期中的全球基因调控，并为破译基因调控网络的复杂性提供有效的工具。在本研究中，将采用一种合作的方法来同时针对DNA结合蛋白的光化学和功能基因组学。科学思想将被整合到三个学科--生物学、化学和物理学--联合研究DNA-蛋白质相互作用的分子生物学家；研究DNA光化学的化学家；应用超短激光脉冲光学的物理学家；以及为跨学科研究提供接口的生物物理学家。学生将接受从高中到博士后的所有级别的培训。

Watt W. Webb, School of Applied and Engineering Physics, John T. Lis, Department of Molecular Biology and Genetics, Tadhg P. Begley, Department of Chemistry and Chemical Biology, Bik-Kwoon Tye, Department of Molecular Biology and Genetics, Eric E. Alani, Department of Molecular Biology and Genetics, and Alexander L. Gaeta, School of Applied and Engineering Physics, all of Cornell University are funded with an award from the Collaborative Research in Chemistry Program with funds provided by the Division of Chemistry, the Division of Physics, the Division of Molecular, Cellular Biology and the Office of Multidisciplinary Activities. This multifaceted team will employ ultrafast UV laser crosslinking to lock proteins to their nucleic acid targets instantaneously and covalently during functional activation as it occurs during the cell cycle. These molecular complexes are then resolved by state-of-the-art analytical protein and nucleic acid biochemistry and structural analysis to identify crosslinked amino acids and their nucleotide binding-sites. These researchers will investigate the network of complex DNA-protein interactions regulating gene transcription, replication, and repair using new technologies that allow high temporal and spatial resolution biochemical analysis of elusive DNA-protein interactions in living cells. The ultimate goal of this research is to develop the methods to perform functional genomic experiments on living cells of prokaryotic and eukaryotic systems to establish genome-wide maps of DNA-bound proteins, thus following global gene regulation control through a cell's growth cycle and providing efficient tools for deciphering the complexity of gene regulator networks.In this research, a collaborative approach will be employed to target both the photochemistry and the functional genomics of DNA-binding-proteins. Scientific ideas will be integrated across three disciplines - biology, chemistry, and physics - uniting molecular biologists investigating DNA-protein interactions; chemists studying the photochemistry of DNA; physicists applying ultrashort laser pulse optics; and biophysicists providing the interface for the interdisciplinary research. Students will receive training at all levels from high school to postdoctoral.