Immunoproteomics

描述（由申请方提供）：拟定研究旨在评价使用质谱读数提供选定蛋白质灵敏检测的新方法，以及监测靶抗原翻译后修饰事件的方法。拟定的测定涉及由固定化抗体介导的抗原捕获步骤（免疫）和涉及结合抗原质谱分析的分析步骤（蛋白质组学）。该项目的目标是开发从不相关蛋白质的复杂混合物中捕获抗原的快速灵敏方法，该方法允许随后使用飞行时间质谱法精确测定结合抗原的分子量。区分靶向抗原的大小的细微变化的能力将允许实现对任何靶向抗原的翻译后修饰事件的分析。此外，将严格评价基于质谱读数上特定分子量峰下面积获得半定量数据的能力。概念验证研究将集中于分泌的链球菌半胱氨酸蛋白酶SpeB的许多性质，已知其对表面抗吞噬M蛋白进行后修饰，并降解分泌的细菌纤溶酶原激活剂SK。

DESCRIPTION (provided by applicant): The proposed studies are designed to evaluate new methods using a mass spectral readout to provide sensitive, detection of selected proteins, as well as methods to monitor post-translational modification events of targeted antigens. The proposed assay involves an antigen capture step mediated by immobilized antibody (immuno) and an analytical step involving mass spectral analysis of bound antigen (proteomics). The goal of the project is to develop rapid sensitive methods of antigen capture from complex mixtures of unrelated proteins in a maimer that permits the subsequent precise molecular weight determination of the bound antigen using time of flight mass spectrometry. The ability to distinguish subtle variation in the size of a targeted antigen will allow analysis of post-translational modification events for any targeted antigen to be achieved. In addition, the ability to obtain semi-quantitative data based on the area under a specific molecular weight peak on the mass spectral read-out will be critically evaluated. The proof of concept studies will focus on a number of properties of the secreted streptococcal cysteine protease SpeB which is known to post-translationally modify the surface anti-phagocytic M protein, and degrade the secreted bacterial plasminogen activator SK.