Identifying the targets of oncogenic/tumor-suppressive F box proteins

 描述（由申请人提供）：泛素生物学代表细胞调节的主要模式。泛素化的最后一步涉及通过“泛素连接酶”识别底物。人类编码超过700种这样的连接酶，显著超过由基因组编码的激酶的数量。在这些泛素连接酶基因中有许多癌基因和肿瘤抑制基因。在许多情况下，这些连接酶的底物尚未被很好地定义，部分原因是缺乏用于鉴定这些底物的稳健技术。我们已经开发了一种方法，我们称之为“连接酶捕获”，它捕获新生的泛素化底物的泛素连接酶，并允许其共纯化和鉴定。在这里，我们将应用这项技术的泛素连接酶特异性亚基称为F盒蛋白。人类编码大约80种这样的蛋白质，我们将重点关注9种已经被证明在肿瘤中突变或错误表达的蛋白质，或者具有与肿瘤发生相关的功能的靶点（如细胞周期调节因子和DNA损伤反应蛋白）。此外，我们将使用这种技术来表征这些底物粘合剂的结合表面。总之，这些数据将允许识别潜在的药物靶点，更重要的是，将允许产生F盒蛋白本身的抑制剂，从而影响药物的作用。 最相关底物的周转率。

 DESCRIPTION (provided by applicant): Ubiquitin biology represents a major mode of cellular regulation. The final step of ubiquitination involves the recognition of a substrate by a "ubiquiti ligase". Humans encode over 700 such ligases, significantly exceeding the number of kinases encoded by the genome. Among these ubiquitin ligase genes are many oncogenes and tumor suppressors. In many cases the substrates of these ligases have not been well defined, in part due to a lack of robust technologies for identifying these substrates. We have developed a method, which we call "Ligase Trapping" which traps the nascent ubiquitinated substrate on the ubiquitin ligase and allows for its co-purification and identification. Here, we will apply this technology to a group of ubiquitin ligase specificity subunits called F box proteins. Human's encode about 80 such proteins, and we will focus on nine that have either been shown to be mutated or mis-expressed in tumors, or have targets whose functions have been linked to tumorigenesis (such as cell cycle regulators and DNA damage response proteins). In addition, we will use this technology to characterize the binding surfaces of these substrate binders. Together, these data will allow the identification of potential drug targets, and, more importantly will allow the generation of inhibitors of the F box proteins themselves in such a way as to affect the turnover of the most relevant substrates.