Proteome Analysis of 14-3-3 Targets in Tomato Fruit Tissues
Proteome Analysis of 14-3-3 Targets in Tomato Fruit Tissues
复制标题
番茄果实组织中 14-3-3 靶标的蛋白质组分析
DOI：
10.1007/978-1-0716-0528-8_21
复制
发表时间：
2020
期刊：
影响因子：
--
通讯作者：
Sato Takeo
中科院分区：
文献类型：
--
作者：
Luo Yongming;Lu Yu;Yamaguchi Junji;Sato Takeo
番茄是一种主要的农作物，也是人类饮食的重要组成部分。番茄果实具有肉质果实和从部分光合作用到完全异养代谢的相变等独特特征，使其成为果实发育研究的模式系统。虽然番茄基因组已经完全测序，但功能蛋白质组学研究仍处于起步阶段。蛋白质组学技术，特别是多种方法的结合，提供了一个非常强大的工具，以准确地识别功能蛋白质和更详细地研究某些蛋白质组。植物14-3-3蛋白与其多个靶蛋白的直接结合调节后者的功能，表明这些14-3-3蛋白直接参与各种生理途径。本章概述了番茄果实组织中14-3-3蛋白复合物的鉴定方法。这些方法包括蛋白质提取、免疫共沉淀、SDS-PAGE、SYPRO Ruby染色、凝胶内胰蛋白酶消化和14-3-3相互作用组的LC-MS/MS分析的详细方案。
Tomato is a major crop plant and an important constituent of the human diet. Exclusive features such as bearing fleshy fruits and undergoing a phase transition from partially photosynthetic to fully heterotrophic metabolism make tomato fruit a model system for fruit development studies. Although the tomato genome has been completely sequenced, functional proteomics studies are still at their starting stage. Proteomics technologies, especially the combination of multiple approaches, provide a very powerful tool to accurately identify functional proteins and investigate certain sets of proteins in more detail. The direct binding of plant 14-3-3 proteins to their multiple target proteins modulates the functions of the latter, suggesting that these 14-3-3 proteins are directly involved in various physiological pathways. This chapter outline methods for the identification of 14-3-3 protein complexes in tomato fruit tissues. These methods include detailed protocols for protein extraction, coimmunoprecipitation, SDS-PAGE, SYPRO Ruby staining, in-gel trypsin digestion, and LC-MS/MS analysis for 14-3-3 interactomics.