Generation and Proteome-Wide Screening of Hybrid Combinatorial Libraries

描述 摘要： 随着人类基因组测序和注释的完成，我们现在拥有了基因组中编码的所有人类蛋白质的完整目录。然而，大多数蛋白质的功能仍然未知。阐明蛋白质功能的一种方法是找到与感兴趣的蛋白质特异性结合并干扰其生物化学和细胞功能的小分子配体。今天化学生物学家面临的一个主要挑战是发现新蛋白质的新小分子探针，以促进其功能的阐明。最近蛋白质芯片的发展提供了一个令人兴奋的新机会，首次同时筛选几乎整个人类蛋白质组的化学库。然而，由于缺乏用于检测配体与芯片上的蛋白质的结合的通用读数，因此对于大多数（如果不是全部的话）现有的化学文库，人类蛋白质芯片的筛选尚不可行。我建议借用一个完善的蛋白质结合结构域的天然产物，既作为一个支架，目前组合肽和非肽库，并作为一个嵌入式通用标签库中的每个化合物。这样的杂交组合文库将适合于使用蛋白质芯片进行蛋白质组范围的筛选，通过利用衍生自天然产物的蛋白质结合结构域与其结合蛋白之间的相互作用，其可以用针对蛋白质的荧光标记的抗体检测。此外，已知的蛋白质结合结构域还可以赋予融合配体稳定性和细胞渗透性，增加来自这种杂交组合文库的命中将容易地适用于研究相关靶蛋白的细胞功能的可能性。这项工作，如果成功的话，可能会导致一种新的结构类型的配体，可用作蛋白质功能的探针。 公共卫生相关性： 该项目旨在产生一种新型的H

DESCRIPTION Abstract: With the completion of the sequencing and annotation of the human genome, we now have a complete catalog of all human proteins encoded in the genome. However, the functions of a majority of the proteins remain unknown. One way to elucidate the functions of proteins is to find small molecule ligands that specifically bind to the proteins of interest and perturb their biochemical and cellular functions. A major challenge for chemical biologists today is to discover new small molecule probes for new proteins to facilitate the elucidation of their functions. The recent advance in the development of protein chips has offered an exciting new opportunity to simultaneously screen chemical libraries against nearly the entire human proteome for the first time. However, screening of the human protein chips is not yet feasible with most, if not all, existing chemical libraries due to the lack of a universal readout for detecting the binding of a ligand to a protein on chips. I propose to borrow a well-established protein-binding domain of natural products to serve both as a scaffold to present combinatorial peptide and non-peptide libraries and as an embedded universal tag for each compound in the library. Such hybrid combinatorial libraries will be amenable to proteome-wide screening using protein chips by exploiting the interaction between the protein binding domain derived from natural products and its binding protein, which can be detected with a fluorescently labeled antibody against the protein. In addition, known protein-binding domain can also confer stability and cell permeability to the fused ligands, increasing the probability that hits from such hybrid combinatorial libraries will be readily applicable to study the cellular functions of the relevant target proteins. This work, if successful, may lead to a new structural class of ligands that can be used as probes of protein function. Public Health Relevance: This project is aimed at generating a new type of h