Proteomic Technologies for Complex Samples

蛋白质在生物过程中起着重要作用。这些作用通过蛋白质水平、相互作用、定位和翻译后修饰的变化来反映。因此，识别和监测细胞中蛋白质水平及其修饰变化的能力可以极大地帮助我们理解生物过程。研究蛋白质组（蛋白质的集合）的技术正在发展。不幸的是，这些技术并不能观察到所有的蛋白质。这导致蛋白质组学研究中重要蛋白质亚群的代表性不足。特别是，对于药物发现、细胞信号传导和其他生物过程至关重要的质膜蛋白，目前的蛋白质组学技术往往没有充分体现。更重要的是，即使通过全球蛋白质组学技术观察到它们，关于蛋白质水平变化和膜特异性修饰的信息也可能被掩盖。我们建议开发微流体技术，使基于细胞区室的亚蛋白质组学研究成为可能，最初的重点是质膜蛋白。我们的短期目标（1-3年）是开发新的膜增溶方案和一整套微流体技术，用于质谱法处理、鉴定和相对定量膜蛋白质组。我们目标的一个重要方面是为学生，博士后研究员和访问科学家提供有关微流控蛋白质组学技术和质谱学发展的培训。我们的中期目标（3-5年）是将这些技术整合到单个设备中，以证明它们在分析亚蛋白质组时的兼容性。我们的长期目标是应用这些微流体技术来研究膜蛋白质组的动态，使这些系统自动化，并探索其商业化。

Proteins play essential roles in biological processes. These roles are reflected through changes in levels, interactions, localization and post-translational modifications of proteins. Therefore, the ability to identify and monitor the level of proteins in the cell and the changes in their modifications can greatly help our understanding of biological processes. Technologies are being developed to study the proteome (ensembles of proteins). Unfortunately, these technologies are not able to look at all the proteins. This leads to under-representation of important subgroups of proteins in proteomic studies. In particular, plasma membrane proteins which are important for drug discovery, cell signaling, and other biological processes are often under-represented using current proteomic technologies. More importantly, even when they are observed by global proteomic technologies, the information about the changes in protein levels and modifications that are specific to the membrane can be overshadowed. We propose to develop microfluidic technologies that will allow the study of sub-proteomes based on cellular compartments with an initial focus on plasma membrane proteins. Our short term objectives (1-3 years) are to develop new membrane solubilization protocols and an ensemble of microfluidic technologies for processing, identifying and relative quantitating membrane proteomes by mass spectrometry. An important aspect of our objectives is to provide training for students, postdoctoral fellows, and visiting scientists on the development of microfluidic proteomic technologies and mass spectrometry. Our medium term objectives (3-5 years) are to integrate these technologies into single devices to demonstrate their compatibility for analyzing sub-proteomes. Our long term objective is to apply these microfluidic technologies to study the dynamicity of the membrane proteome, to automate these systems, and to explore their commercialization.