Institute of Chemical Technology Prague, Department of Biochemistry and Microbiology, Prague, Czech Republic.
Proteomics. 2011 Aug;11(16):3430-4. doi: 10.1002/pmic.201000603.
Plant seed oil bodies, subcellular lipoprotein inclusions providing storage reserves, are composed of a neutral lipid core surrounded by a phospholipid monolayer with several integrated proteins that play a significant role in stabilization of the particles and probably also in lipid mobilization. Oil bodies' proteins are generally very hydrophobic, due to the long uncharged sequences anchoring them into the lipid core, which makes them extremely difficult to handle and to digest successfully. Although oil bodies have been intensively studied during last decades, not all their proteins have been identified yet. To overcome the problems connected with their identification, a method based on SDS-PAGE, in-gel digestion and LC-MS/MS analysis was used. Digestion was carried out with trypsin and chymotrypsin, single or in combination, which increased significantly the number of identified peptides, namely the hydrophobic ones. Thanks to this methodology it was possible to achieve an extensive coverage of proteins studied, to analyze their N-terminal modifications and moreover, to detect four new oil bodies' protein isoforms, which demonstrates the complexity of oil bodies' protein composition.
植物种子油体是亚细胞脂蛋白内含物，为储存储备提供了场所，由一个中性脂质核心组成，周围环绕着一个磷脂单层，其中有几个整合蛋白，这些蛋白在稳定颗粒方面发挥着重要作用，可能在脂质动员中也起着重要作用。油体蛋白通常非常疏水，这是由于将它们锚定在脂质核心的长非带电序列所致，这使得它们极难成功处理和消化。尽管油体在过去几十年中得到了深入研究，但并非所有的蛋白质都已被鉴定出来。为了克服与鉴定相关的问题，使用了基于 SDS-PAGE、胶内消化和 LC-MS/MS 分析的方法。用胰蛋白酶和糜蛋白酶进行消化，单独或组合使用，这大大增加了鉴定肽的数量，即疏水性肽的数量。得益于这种方法，可以对所研究的蛋白质进行广泛的覆盖，分析它们的 N 端修饰，并且可以检测到四个新的油体蛋白同工型，这证明了油体蛋白组成的复杂性。
