南方医科大学学报

南方医科大学学报 ›› 2005, Vol. 25 ›› Issue (09): 1152-1154,1157.

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小鼠肝脏磷酸化蛋白质组的二维液相色谱分离

黎永明, 陈腾祥, 杨丽萍, 刘亚伟, 姜勇   

  1. 南方医科大学病理生理学教研室, 广东, 广州, 510515
  • 出版日期:2005-09-20 发布日期:2005-09-20
  • 基金资助:
    收稿日期:2005-4-25。
    基金项目:国家杰出青年科学基金(39925014);国家自然科学基金(30030060);重大基础理论研究(973)计划项目(2002CB513000);国家高技术研究发展计划(863)课题(2001AA234061)
    作者简介:黎永明(1978- ),女,2002年毕业于河北医科大学,在读硕士研究生,电话:(020)61640114-89103,E-mail:gslym@etang.com.
    通讯作者:姜勇,电话:(020)61648231,E-mail:yjiang@public;guangzhou.gd.cn.

Two-dimensional liquid chromatographic fractionation of mouse liver phosphoproteome

LI Yong-ming, CHEN Teng-xiang, YANG Li-ping, LIU Ya-wei, JIANG Yong   

  1. 南方医科大学病理生理学教研室, 广东, 广州, 510515
  • Online:2005-09-20 Published:2005-09-20

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摘要： 目的 利用二维液相色谱法分离小鼠肝脏磷酸化蛋白质组。方法 取正常小鼠肝脏,裂解肝脏后利用金属磷酸盐亲和层析树脂提取磷酸化蛋白。将磷酸化蛋白用初始缓冲液置换后,进行一维色谱聚焦分离,再将一维收集的pH值在8.5至4.0之间的组分分别进行二维无孔硅胶反相高效液相色谱分离。最后利用ProteoVue软件将二维UV图转换成胶图进行分析。结果 成功提取了小鼠肝脏磷酸化蛋白,并在浓缩除盐后通过二维液相色谱分离成功建立小鼠肝脏磷酸化蛋白质组pI/UV图谱。其中,一维色谱聚焦分离pH值在8.5至4.0之间共收集16个组分,每个组分的二维UV图转换成pI/UV胶图。结论 磷酸化蛋白纯化技术与二维液相色谱技术的结合是研究磷酸化蛋白质组的有效方法,为下一步鉴定和研究磷酸化蛋白的功能打下坚实的基础。

Abstract: Objective To fractionate mouse liver phosphoproteome by two-dimensional liquid chromatography. Methods The phosphoproteins were extracted from the lysates of normal mouse livers with phosphate metal affinity chromatography resin. The phosphoproteins were exchanged by the initial buffer and separated by chromatofocusing in the first dimension. The fractions with pH value between 8.5 and 4.0 were separated by non-porous silica reverse-phase high-performance liquid chromatography. Finally, the UV maps were converted into gel maps by ProteoVue software. Results The phosphoproteins of mouse liver were successfully extracted and fractionated by two-dimensional liquid chromatographic fractionation after concentration and desalting. The pI/UV map of mouse liver phospho-proteomic was obtained successfully. There were 16 fractions with pH between 8.5 and 4.0 after chromatofocusing in the first dimension and the UV maps of each fraction were converted into pI/UV gel maps. Conclusion Combination of phosphoprotein enrichment and two-dimensional liquid chromatographic fractionation is an effective approach of phosphoproteomic studies.

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