Renewable Antibodies for Post Translational Modifications and Protease Activatio

用于翻译后修饰和蛋白酶激活的可再生抗体

基本信息

批准号：
8702418
负责人：
JAMES A WELLS
金额：
$ 79.75万
依托单位：
UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-09-24 至 2019-08-31
项目状态：
已结题

项目摘要

The long-term goal of the TR&D 3 project is to develop high-impact, renewable, and open-source antibody reagents for cellular signaling and biomedical research. Specifically, we plan to focus on signaling events marked by important protein post-translational modifications (PTMs) such as phosphorylation and proteolytic zymogen activation events and their attendant protein conformational changes. While some antibodies do exist for specific phosphorylation sites, these antibodies are often unreliable, rarely renewable, and only available to a fraction of the modified sites. In addition, detecting functional forms of specific proteases has not been routinely possible in cells or in vivo due to the lack of conformationally selective antibodies. Furthermore, the industrialization of high-throughput recombinant antibody (rAb) selection and preparation has been thwarted by the absence of reliable robotic expression platforms. Thus, our immediate goals are to generate high-quality, open-source antibody reagents, by using novel phage display technologies and libraries developed by the UCSF Antibiome Center, to PTMs including: (i) phosphorylation sites, (ii) proteolytic neo-epitopes, and their attendant conformational changes, and (iii) alternative rapid antigen expression technologies such as in vitro transcription and translation (IVTT) that support all three TR&D projects. These goals compliment TR&Ds 1 and 2 by focusing on PTMs within key transmembrane proteins

TR＆D 3项目的长期目标是开发用于细胞信号传导和生物医学研究的高影响力，可再生和开源抗体试剂。具体而言，我们计划专注于以重要蛋白质后翻译（PTM）为标志的信号事件，例如磷酸化和蛋白水解酶原激活事件及其随之而来的蛋白构象变化。尽管确实存在一些针对特定磷酸化位点的抗体，但这些抗体通常是不可靠的，很少可再生，并且仅适用于一部分修饰位点。此外，检测特定蛋白酶的功能形式尚未由于缺乏构象选择性抗体，在细胞或体内常规可能实现。此外，由于缺乏可靠的机器人表达平台而阻止了高通量重组抗体（RAB）选择和制备的工业化。因此，我们的近期目标是通过使用UCSF抗体中心开发的新型噬菌体展示技术和图书馆来生成高质量的开源抗体试剂，用于PTM，包括：（i）磷酸化位点，（i）（ii）蛋白水解的新型potepopes及其随身携带的型号，以及随身携带的（IIIII）的替代性及其（III III），以及（IIIIII），并具有（IIIII）的（III III），并具有（IVTT）支持所有三个TR＆D项目。这些目标通过关注密钥跨膜蛋白中的PTM来补充TR＆D 1和2