质谱法分析大肠癌组织中踝蛋白的磷酸化修饰

引用本文: 孙莉, 赵海丰, 郭宏华, 扬光远, 何成彦, 师庆红, 文雪, 王小婷, 赵丽纯. 质谱法分析大肠癌组织中踝蛋白的磷酸化修饰[J]. 分析化学, 2012, 40(10): 1500-1506. doi: 10.3724/SP.J.1096.2012.20027 shu

Citation: SUN Li, ZHAO Hai-Feng, GUO Hong-Hua, YANG Guang-Yuan, HE Cheng-Yan, SHI Qing-Hong, WEN Xue, WANG Xiao-Ting, ZHAO Li-Chun. Mass Spectrometric Analysis of Phosphorylation Modification in Talin from Human Colorectal Carcinoma Tissues[J]. Chinese Journal of Analytical Chemistry, 2012, 40(10): 1500-1506. doi: 10.3724/SP.J.1096.2012.20027 shu

质谱法分析大肠癌组织中踝蛋白的磷酸化修饰

孙莉 ^1, ,
赵海丰 ^2, ,
郭宏华 ^1, ,
扬光远 ^2, ,
何成彦 ^1, ,
师庆红 ^1, ,
文雪 ^3, ,
王小婷 ^3, ,
赵丽纯 ^{3,
,}

通讯作者: 赵丽纯

基金项目:
本文是国家自然科学基金(No.21175055) (No.21175055)

吉林省科技厅项目(Nos.2011713,20110739)资助 (Nos.2011713,20110739)

摘要: 踝蛋白的磷酸化修饰,特别是肿瘤等病理条件下的踝蛋白磷酸化状态,与肿瘤的发病、转移机理密切相关。本研究采用盐析、离子交换层析和电泳分离并纯化了人大肠癌组织中的踝蛋白(Talin),经胰酶水解获得其肽段混合物,进一步分别利用固定化Fe³⁺亲和层析和TiO₂亲和层析在酸性条件下对其中磷酸化修饰肽段进行吸附,并以1%氨水进行洗脱。在Michrom Magic C₁₈色谱柱上, 以A:99%水+1%乙腈+0.1%甲酸和B:99%乙腈+1%水+0.1%甲酸两种流动相进行梯度洗脱分离,采用ESI质谱进行依赖数据的二级子离子扫描。结果显示,固定化Fe³⁺富集到8个磷酸化肽段而TiO₂富集到9个磷酸化肽段。本研究提供了一种快速、准确地鉴定从人大肠癌组织中分离表征踝蛋白的方法。

关键词:

质谱
/ 踝蛋白
/ 大肠癌
/ 磷酸化修饰

English

Mass Spectrometric Analysis of Phosphorylation Modification in Talin from Human Colorectal Carcinoma Tissues

1.
¹ China-Japan Union Hospital, Jilin University, Changchun 130033, China;

Corresponding author: 赵丽纯

Received Date: 10 January 2012
Fund Project:
本文是国家自然科学基金(No.21175055)

吉林省科技厅项目(Nos.2011713,20110739)资助

Abstract: The phosphorylation modification of talin, especially the phosphorylation state of talin in pathological environment such as carcinoma, is closely relevant to carcinogenesis and metastasis process. In this study, talin protein was isolated from human colorectal carcinoma tissues by salt fractionation and ion exchange chromatography, followed by further purification by electrophoresis. The purified talin was subject to tryptic digestion. Either immobilized Fe³⁺ affinity chromatography or TiO₂ affinity chromatography was used to absorb the phosphorylated peptides under acidic condition, which were then eluted with 1% ammonium hydroxide. The separation was performed on a Michrom Magic C₁₈ column with gradient elution using two mobile phase solutions (A:99% water+1% ACN+0.1% formic acid; B:99% ACN+1% water+0.1% formic acid). ESI mass spectrometer was used to detect the product ions of the eluted peaks under data-dependent acquisition mode. The results indicated that 8 phosphorylated peptides were captured by the immobilized Fe³⁺ affinity chromatography, whereas 9 phosphorylated peptides were captured by TiO₂ affinity chromatography. The present study provides a rapid, accurate method for characterizing talin protein isolated from human colorectal carcinoma tissues.

Key words:

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通讯作者: 陈斌, bchen63@163.com

1.
沈阳化工大学材料科学与工程学院沈阳 110142

质谱法分析大肠癌组织中踝蛋白的磷酸化修饰

通讯作者: 赵丽纯

English

Mass Spectrometric Analysis of Phosphorylation Modification in Talin from Human Colorectal Carcinoma Tissues

Corresponding author: 赵丽纯

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通讯作者: 陈斌, bchen63@163.com

中国化学会秘书处

质谱法分析大肠癌组织中踝蛋白的磷酸化修饰

通讯作者: 赵丽纯

English

Mass Spectrometric Analysis of Phosphorylation Modification in Talin from Human Colorectal Carcinoma Tissues

Corresponding author: 赵丽纯

CCS Chemistry：嵌段共聚物自组装成 “三明治”二维有机材料，实现纳米级压力传感功能

CCS Chemistry：颜徐州、鲍哲南： 你的皮肤可能是一片SPMs

CCS Chemistry：工作高效不疲劳，只须让催化剂动起来

CCS Chemistry：静电组装导向聚合- 聚电解质纳米凝胶量化制备新策略

CCS Chemistry：金黄色葡萄球菌醛基脱氢酶是如何识别特异性底物的？

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通讯作者: 陈斌, bchen63@163.com

中国化学会秘书处

CCS Chemistry：颜徐州、鲍哲南：你的皮肤可能是一片SPMs