Glisovic-Aplenc Tina, Gill Saar, Spruce Lynn A, Smith Ian R, Fazelinia Hossein, Shestova Olga, Ding Hua, Tasian Sarah K, Aplenc Richard, Seeholzer Steven H
Division of Oncology, Center for Childhood Cancer Research, The Children's Hospital of Philadelphia, Philadelphia, PA, USA.
Department of Pediatrics, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA, USA.
Proteomics. 2017 Apr;17(7). doi: 10.1002/pmic.201600344. Epub 2017 Mar 6.
The proteins of the cellular plasma membrane (PM) perform important functions relating to homeostasis and intercellular communication. Due to its overall low cellular abundance, amphipathic character, and low membrane-to-cytoplasm ratio, the PM proteome has been challenging to isolate and characterize, and is poorly represented in standard LC-MS/MS analyses. In this study, we employ sucrose gradient ultracentrifugation for the enrichment of the PM proteome, without chemical labeling and affinity purification, together with GeLCMS and use subsequent bioinformatics tools to select proteins associated with the PM/cell surface, herein referred to as the surfaceome. Using this methodology, we identify over 1900 cell surface associated proteins in a human acute myeloid leukemia cell line. These surface proteins comprise almost 50% of all detected cellular proteins, a number that substantially exceeds the depth of coverage in previously published studies describing the leukemia surfaceome.
细胞质膜（PM）的蛋白质执行与体内平衡和细胞间通讯相关的重要功能。由于其在细胞内的总体丰度较低、具有两亲性且膜与细胞质的比例较低，PM蛋白质组的分离和表征具有挑战性，并且在标准的液相色谱-串联质谱（LC-MS/MS）分析中代表性不足。在本研究中，我们采用蔗糖梯度超速离心法富集PM蛋白质组，无需化学标记和亲和纯化，并结合凝胶内消化液相色谱-质谱联用（GeLCMS），随后使用生物信息学工具选择与PM/细胞表面相关的蛋白质，在此称为表面蛋白质组。使用这种方法，我们在人急性髓系白血病细胞系中鉴定出超过1900种细胞表面相关蛋白。这些表面蛋白几乎占所有检测到的细胞蛋白的50%，这一数字大大超过了先前描述白血病表面蛋白质组的研究中的覆盖深度。
