| 改进的~(18)O同位素标记法标记蛋白质多肽 |
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| 引用本文： | 赵焱,卢庄,贾伟,应万涛,钱小红. 改进的~(18)O同位素标记法标记蛋白质多肽[J]. 分析化学, 2010, 38(1) |
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| 作者姓名： | 赵焱 卢庄 贾伟 应万涛 钱小红 |
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| 作者单位： | 蛋白质组学国家重点实验室,北京蛋白质组研究中心,北京,102206;军事医学科学院放射与辐射医学研究所,北京,100850;蛋白质组学国家重点实验室,北京蛋白质组研究中心,北京,102206 |
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| 基金项目： | 国家自然科学基金(Nos.20635010,20735005);;国家重点基础研究发展规划项目(No.2007CB914104)资助 |
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| 摘 要： | 针对18O同位素标记反应两个重要影响因素——肽段分散度和胰酶灭活方法,进行了标记条件的改进和灭活方法的优化。在H218O中加入RapigestTM SF助溶剂并微波辅助加热,使α-酪蛋白胰酶酶切肽段的标记效率得到明显改进(18O/16O峰面积比值>99%)。标记后,对胰酶进行还原烷基化化学修饰彻底灭活,使标记后的肽段稳定性显著提高,放置6d不发生回交反应。对标准蛋白质甲状腺球蛋白酶切肽段混合物标记后的质谱实验结果表明:优化的标记方法能快速稳定地标记蛋白质酶切多肽。
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| 关 键 词： | (18)O同位素标记 多肽 蛋白质组 |
Peptide Labeling with Improved ~(18)O Incorporation Method |
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| ZHAO Yan,LU Zhuang,JIA Wei,YING Wan-Tao,QIAN Xiao-Hong. Peptide Labeling with Improved ~(18)O Incorporation Method[J]. Chinese Journal of Analytical Chemistry, 2010, 38(1) |
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| Authors: | ZHAO Yan LU Zhuang JIA Wei YING Wan-Tao QIAN Xiao-Hong |
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| Affiliation: | State Key Lab of Proteomics;Beijing Proteome Research Center;Beijing 102206;Beijing Institute of Radiation Medicine;Beijing 100850 |
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| Abstract: | In order to optimize the ~(18)O labeling method, two key aspects, peptide dispersion and trypsin deac tivation were discussed o The addition of Rapigest SF in H_2~('8)O and microwave heating enhanced labeling efficiency of α-casein digested peptides(~(18)O/~(16)O) ratio >99%).Chemical modification with tris(2-carboxyeth yl) phosphine (TCEP) and iodoacetamide (IAA) resulted in trypsin deactivated completely.No significant back-exchange from ~(18)O to ~(16)O was observed after labeling in 6 days.The experiment result with peptide mixture from showed that the improved method could be effectively used to label protein and peptide. |
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| Keywords: | 18O stable isotope labeling Peptide Proteome |
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