人肝癌细胞系HLE细胞表面抗原多肽的纳升电喷雾串联质谱从头测序分析

肿瘤细胞表面的抗原多肽能够被细胞毒T淋巴细胞特异性识别而引起免疫应答,因此有可能用于研制基于多肽的抗肿瘤疫苗。用弱酸将人肝癌细胞系HLE细胞表面抗原多肽和人正常肝细胞表面多肽洗脱后,经RP-HPLC分离,选择HLE细胞表面特异性多肽进行纳升电喷雾串联质谱(nanoESI-MS/MS)测序,共测定5个色谱峰中的20个多肽序列,分子量分布范围为1000~2000 Da。借助M asSeq软件分析出其中12个多肽的序列。经数据库查寻,其中的3个肽段分别来自钙调节蛋白、核蛋白S19和伴侣蛋白10。这些多肽的生物学功能及与肿瘤的关系值得深入研究。该研究表明nanoESI-MS/MS是测定微量混合多肽序列的最有效方法。

De Novo Sequencing of Antigen-derived Peptides Expressed on the Surface of Human Liver Cancer HLE Cells by Nanoelectrospray Tandem Mass Spectrometry

Antigen-derived peptides expressed at the cell surface of tumor cells may be recognized by melanoma-specific cytotoxic T lymphocytes (CTLs) and could be used in peptide-based melanoma vaccines.Such peptides from human liver cancer HLE cells surface were eluted by weak acid and were subsequently fractionated by reverse phase high performance liquid chromatography(RP-HPLC).The peptides from human liver cells were also eluted and then separated by RP-HPLC with the exact same methods.Five peaks specifically existed in HLE cells after comparing the RP-HPLC spectrum with the normal liver cells.Nanoelectrospray(tandem) mass spectrometry(nano ESI-MS/MS) was used to sequence and identify these peptide.Twenty(peptides) were sequenced and among these 12 got confirmed amino acid sequence after sequence analysis.The mass range of these peptides were 1000-2000 Da.After database search,three peptides were derived from protein calcium calmodulin,ribosomal protein S19 and chaperonin 10 respectively and the other nine peptides were new peptides.The biological function of these peptides and their relationship with tumor need be studied further.This study showed that nanoESI-MS/MS is a powerful tool for peptide sequencing from mixture with small amount.