Development of diagnostic platform for biomolecular interactions and biocatalysis using capillary electrophoresis and mass spectrometry

我们申请合作研究和发展基金有两个主要目标。第一个目标是开发一个基于毛细管电泳联用电喷雾电离质谱仪(MS)的筛选酶、底物和抑制剂的新平台。CE-MS只需要纳升的试剂溶液，并使用分离毛细管作为微反应器，允许同时检测多个底物。该方法可用于快速分析任何酶的底物专一性，并以自动方式测量稳态动力学。*第二个目标是开发一个新的诊断平台，基于动力学CE-MS研究蛋白质动力学和亲和力相互作用。动力学CE-MS建立了一种新的范式，即分离方法与MS一起作为蛋白质组学和基因组学中质量和结构分析的综合动力学工具。动力学CE-MS是研究蛋白质结构和功能的有价值的平台，包括构象的原位催化活性和它们对抑制剂结合的敏感性。*我们的目标是将加拿大Lumex仪器公司生产的CE仪器与质谱仪结合在一个诊断平台上。UOttawa和Lumex的这一合作项目的成功完成预计将产生基于毛细管电泳平台的新技术，这些技术将在诊断、药物发现、酶开发、抗体筛选中得到广泛应用，并将影响加拿大工业和加拿大人的福祉。*

There are two main goals of our application to the Collaborative Research and Development Grant. The first goal is to develop a new platform for screening enzymes, substrates and inhibitors based on capillary electrophoresis (CE) coupled to electrospray ionization mass spectrometry (MS). CE-MS requires only nanoliters of reagent solutions and uses the separation capillary as a microreactor, allowing multiple substrates to be assayed simultaneously. The method can be used to rapidly profile the substrate specificity of any enzyme and to measure steady-state kinetics in an automated fashion. ****The second goal is to develop a new diagnostic platform to study protein dynamics and affinity interactions based on kinetic CE-MS. Kinetic CE-MS establishes a new paradigm that separation methods together with MS are used as comprehensive kinetic tools with mass and structure elucidation in proteomics and genomics. Kinetic CE-MS serves as a valuable platform for studying protein structure and function, including the in situ catalytic activity of conformers and their susceptibility to inhibitor binding.****We aim to combine a CE instrument made by LUMEX Instruments Canada in Canada with a mass spectrometer in one diagnostic platform. The successful completion of this collaborative project between uOttawa and LUMEX is expected to generate new technologies based on the capillary electrophoresis platforms that will find wide application in diagnostics, drug discovery, enzyme development, antibody screening and will impact both Canadian industry and the well-being of Canadians.****