高等学校化学学报 ›› 2013, Vol. 34 ›› Issue (12): 2704.doi: 10.7503/cjcu20130824

• 分析化学 • 上一篇    下一篇

基于化学衍生和CID碎裂模式鉴定蛋白精氨酸-ADP-核糖基化的新方法

赵志强1,2, 邱辉华1, 姚军1,2, 陈永峰1   

  1. 1. 台州学院医学院, 台州 318000;
    2. 肿瘤研究所, 台州 318000
  • 收稿日期:2013-08-27 出版日期:2013-12-10 发布日期:2013-10-23
  • 作者简介:赵志强,男,博士,副教授,主要从事蛋白组学和生物质谱方法学研究. E-mail:xiaosazhaocn@163.com
  • 基金资助:

    浙江省自然科学基金(批准号:LY12C05002)资助.

Novel Method for Identification of Protein Arginine-ADP-ribosylation Based on Chemical Derivatization and CID Fragmentation

ZHAO Zhi-Qiang1,2, QIU Hui-Hua1, YAO Jun1,2, CHEN Yong-Feng1   

  1. 1. Medical School, Taizhou 318000, China;
    2. Institute of Tumor, Taizhou University, Taizhou 318000, China
  • Received:2013-08-27 Online:2013-12-10 Published:2013-10-23

摘要:

建立了一种新的基于碰撞诱导解离(CID)碎裂模式鉴定精氨酸-腺苷二磷酸(ADP)-核糖基化多肽的新方法. 首先,在碱性条件下将精氨酸-ADP-核糖基化血管紧张素-Ⅰ转变为鸟氨酸化血管紧张素-Ⅰ,或在磷酸二酯酶和碱性磷酸酶处理下水解为精氨酸核糖基化血管紧张素-Ⅰ,然后对上述2种衍生物进行基于CID碎裂模式的串联质谱分析. 结果表明,与未衍生的精氨酸-ADP-核糖基化血管紧张素-Ⅰ相比,在鸟氨酸化血管紧张素-Ⅰ和精氨酸核糖基化血管紧张素-Ⅰ的质谱图上发现大部分来自于肽骨架碎裂的离子峰,可提供足够的序列信息以确定精氨酸-ADP-核糖基化位点.

关键词: 血管紧张素-Ⅰ, 化学衍生, 碰撞诱导解离, 质谱, 精氨酸-ADP-核糖基化

Abstract:

High performance liquid chromatography coupled to electrospray ionization mass spectrometry(MS) was widely used to map protein posttranslational modifications in complex biological samples. But frequently-used collision induced dissociation(CID) was unsuccessful in fragmentizing arginine adenosine diphosphate(ADP)-ribosylated peptides because the spectra from CID fragmentation were dominated by ADP-ribose fragment ions at the expense of the fragments generated from peptide backbone, making the assignment of the peptide sequence very difficult. In this work, a novel method was developed for identification of arginine ADP-ribosylated peptides based on CID fragmentation. Arginine ADP-ribosylated angiotensin-Ⅰ was either converted to ornithinylated angiotensin-Ⅰ under alkaline condition or hydrolyzed to arginine ribosylated angiotensin-Ⅰ by treatment with phosphodiesterase and alkaline phosphatase. The two derivatives were then subjected to CID fragmentation during tandem MS analysis. Compared with unmodified ADP-ribosylated angiotensin-Ⅰ, the two derivatives generated more sequence specific ions in the spectra, giving enough information to localize the site of arginine ADP-ribosylation.

Key words: Angiotensin-Ⅰ, Chemical derivatization, Collision induced dissociation, Mass spectrometry, Arginine adenosine diphosphate(ADP)-ribosylation

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