二维电泳和质谱技术鉴定肝癌血清差异表达的N-连接糖蛋白

Proteomic Profiling of N-linked Glycoproteins Involved in Hepatocellular Carcinoma by Two-dimensional Electrophoresis and Mass Spectrometry

缺乏有效的早期诊断方法是导致肝细胞癌(hepatocellular carcinoma,HCC)预后极差的主要原因之一.蛋白质异常糖基化与恶性肿瘤细胞侵袭、转移等生物学过程关系密切,人体内至少有50%的蛋白质发生了糖基化修饰.本实验采用IgY12去除血清高丰度蛋白、多植物凝集素亲和层析技术分别从20例肝癌和年龄、性别匹配的20例非癌慢性肝病患者血清中纯化N-连接糖蛋白、二维电泳分析差异表达的蛋白质斑点,质谱检测、生物信息学等技术鉴定了18个差异表达的糖蛋白和/或其异质体(12种高表达和6种低表达).ExPASy数据库比对结果表明,本实验鉴定的糖蛋白质分子含有至少1个已报道的N-糖基化位点.这些差异表达的糖蛋白属于急性期反应蛋白,分别具有蛋白酶抑制、生物转运、凝血和纤溶等功能,表明肝癌的发生发展过程中机体产生的急性期反应物可能是潜在的肝癌血清标志物.

The shortage of effective early diagnosis methods for early hepatocellular carcinoma （HCC） represents one of the main contributors for worse prognosis of HCC. Glycoproteins comprise nearly half of the human proteome, and aberrant protein glycosylation is closely correlated with invasion and metastasis of malignant cells during carcinogenesis. In the present study, N-linked glycoproteins were isolated from the immunodepleted sera of high abundant proteins from 20 HCC patients and 20 controls or subjects of chronic liver disease with matched age and sex. By two dimensional electorphoresis-based proteomics, mass spectrometry and matching against the ExPASy database, a total of 18 differential glvconroteins and/or glycoprotein fragments were identified, including 12 up-regulated and 6 down-regulated l＇^-llnkecl glycoproteins, each of which contained at least one N-glycosylation site. These differentially expressed glycoproteins belonging to acute-phase proteins that play multiple functions, such as proteinase inhibition, substrate transport, coagulation, and fibrinolysis, etc. , which might become potential serum biomarkers for HCC diagnosis.