INTRACELLULAR MODIFICATIONS OF HUMAN APOLIPOPROTEIN E

这个子项目是利用由NIH/NCRR资助的中心拨款提供的资源的许多研究子项目之一。子项目和调查员(PI)可能从另一个NIH来源获得了主要资金，因此可能会出现在其他CRISE条目中。列出的机构是针对中心的，而不一定是针对调查员的机构。人血浆载脂蛋白E(ApoE)负责清除脂蛋白残基，与家族性III型高脂蛋白血症和阿尔茨海默病-S病等疾病有关。已知它在合成后最初是唾液酸化的，后来在血浆中需要分析。对HepG2细胞的apoE合成和细胞内修饰的研究表明，修饰后的apoE同种蛋白对应于含有两个、四个或六个唾液酸残基的sialo apoE形式。预期的O-糖基化位点需要确定，修饰的种类还需要用质谱学来表征。从apoE的胰酶消化中，LC-MS/MS可以在预期的位置检测到一个二唾液酸化的多糖，然后用标准的MALDI TOF MS也可以检测到它。振动冷却(VC)MALDI FT-MS可能有助于检测更高唾液酸化、更不稳定的同系物。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Human plasma apolipoprotein E (apoE) is responsible for the clearance of lipoprotein remnants and has been implicated in diseases like familial type III hyperlipoproteinemia and Alzheimer?s disease. It is known to be initially sialylated after synthesis and later desialylated in the plasma. Studies of synthesis and intracellular modification of apoE by HepG2 cells have shown modified apoE isoproteins corresponding to the sialo apoE forms with two, four or six sialic acid residues. The expected O-glycosylation site needs to be confirmed and the kind of modification to be characterized by mass spectrometry. From a tryptic digest of apoE, one disialylated glycan could be detected at the expected position by LC-MS/MS, and was then also found by standard MALDI TOF MS. It is planned to isolate the glycopepitde(s) with the help of lectins. Vibrationally cooled (VC) MALDI FT-MS might help to detect higher sialylated, more instable homologues.