纳升液相色谱-飞行时间质谱鉴定HaCaT细胞膜蛋白  

Nano High Performance Liquid Chromatography Coupling with Time-of-flight Mass Spectrometer for Identification of Membrane Proteins in HaCaT Cells

作　　者：曾卓颖[1,2] 张航[2] 黄爱博 夏菠[1,2] 洪文旭[2,3] 刘建军[2] 郭时印[1] 

机构地区：[1]湖南农业大学食品科学技术学院,湖南长沙410128 [2]深圳市疾病预防控制中心,深圳市现代毒理学重点实验室,广东深圳518055 [3]深圳市人口和计划生育科学研究所,广东深圳518040

出　　处：《安徽农业科学》2013年第28期11293-11294,11335,共3页Journal of Anhui Agricultural Sciences

基　　金：国家自然科学基金(21102094);广东省医学科研基金(A2012577);深圳市重点实验室提升项目(ZDSY2-0120615085804889);深圳市科技创新委基础研究项目(JCYJ20130329161137543)

摘　　要：[目的]探讨通过纳升液相色谱-飞行时间质谱检测人永生化表皮细胞的膜蛋白质的方法。[方法]对人永生化表皮细胞进行贴壁培养,然后对处于对数生长期细胞的膜蛋白进行提取、富集和酶解,用nano-HPLC-TOF/TOF-MS对蛋白质进行鉴定,最后用生物信息学技术对鉴定蛋白进行分析和归类。[结果]在鉴定的15个蛋白中,与细胞膜相关的蛋白有9个,占比例的60%。生物信息学分析表明,这些蛋白在分子功能上主要涉及催化,结构分子活性和结合;在生物学进程上,这些蛋白主要归类为代谢,细胞成分形成和细胞生物进程等。[结论]纳升色谱-飞行时间质谱是鉴定膜蛋白的有效方法。[ Objective] To apply the nano high performance liquid chromatography coupling with time-of-flight mass spectrometer （nano- HPLC-TOF-MS） to detect the membrane proteins in HaCaT cells. [ Methods] HaCaT cells were collected after the cell confluence reached up to 70% -80% by membrane extraction kit. Then, the enzymatic hydrolysis was conducted on the membrane protein comPlexes, and followed by separation and detection with nano-HPLC-TOF-MS. The identified proteins were classified by bioinformatics analysis. [ Result] There are 9 membrane proteins of the identified 15 proteins, which accounted for the proportion of sixty percents. The bioinformatics analysis indicated that these membrane proteins mainly involved into molecular functions such as catalysis, structure molecular activity and combination. For the bio- logical process, these proteins mainly involved with metabolism and cell components formation. [ Conclusion ] Nano-HPLC-TOF-MS is effective to separate.and detect membrane proteins in HaCaT cells.

关 键 词：人永生化表皮细胞 膜蛋白质 纳升液相色谱 飞行时间质谱 

分 类 号：S188[农业科学—农业基础科学]