Characterizing proteasome-substrate interactions by mass spectrometry proteomics

项目摘要 该应用程序的特点是生物分析（Reilly博士，印第安纳州大学）和生物 化学（Matouschek博士，得克萨斯大学，奥斯汀）的研究小组，旨在提高我们的理解 泛素-蛋白酶体蛋白质降解系统。调查人员提供补充性专门知识 在蛋白质组学，蛋白质生物信息学，分析化学等多个学科的经验 和仪器，到生物化学，分子生物学和细胞生物学。在印第安纳州大学， 质谱分析将与生物化学和遗传分析相结合，以表征 蛋白酶体及其底物，以提供蛋白酶体上新底物受体的候选物。在 得克萨斯大学奥斯汀分校的生物学相关性考生随后将进行生化亲测， teasome分析和遗传酵母实验。这些试验将确定新鉴定的 蛋白酶体对底物的识别和降解，并阐明其分子和结构。 底物选择和特异性的自然起源。 总的来说，我们认为，这一建议将导致重大进展的交联方法和我们的 了解蛋白酶体如何识别和消化底物。

PROJECT SUMMARY This application features a close collaboration between bioanalytical (Dr. Reilly, Indiana University) and bio- chemistry (Dr. Matouschek, University of Texas, Austin) research groups aimed at improving our understanding of the ubiquitin-proteasome protein degradation system. The investigators contribute complementary expertise and experience in a variety of disciplines ranging from proteomics, protein bioinformatics, analytical chemistry and instrumentation, to biochemistry, molecular biology and cell biology. At Indiana University, cross-linking mass spectrometry will be combined with biochemical and genetic assays to characterize the interaction between the proteasome and its substrates to provide candidates for novel substrate receptors on the proteasome. At The University of Texas at Austin, the biological relevance of candidates will then be tested in biochemical pro- teasome assays and genetic yeast experiments. These assays will define the contribution of newly identified components to substrate recognition and degradation by the proteasome and elucidate the molecular and struc- tural origin of substrate selection and specificity. Overall, we believe that this proposal will result in significant advances in cross-linking methodology and in our understanding of how the proteasome recognizes and digests substrates.