人类APPL2蛋白PTB结构域的表达纯化及其功能的研究

中国生物工程杂志

2009, Vol. 29

Issue (08): 1-7

研究报告

人类APPL2蛋白PTB结构域的表达纯化及其功能的研究

梁宁¹,石珠亮²,王东业²,魏国威¹,孔祥平³,陈昌友¹,刘劲松¹,徐爱民⁴,吴东海¹

1. 中国科学院广州生物医药与健康研究院
2. 中国科学技术大学生命科学学院
3. 解放军第458医院全军肝病中心
4. 香港大学药学院

Expression,Purification of PTB Domain of Human APPL2 and Screen for Its Peptide Ligands

LIANG Ning^1,2SHI Zhu-liang^1,2WANG Dong-ye¹WEI Guo-wei¹KONG Xiang-ping³
CHEN Chang-you^1 |LIU Jin-song¹XU Ai-min^1,4WU Dong-hai^1,2

1 Guangzhou Institute of Biomedicine and Health,Chinese Academy of Science,Guangzhou510663,China
2 Department of Life Science,University of Science and Technology of China,Hefei230026,China
3 Liver Disease Center of PLA,PLA 458 Hospital,Guangzhou510602,China
4 Department of Medicine,University of Hong Kong,Hong Kong999077,China

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摘要：

APPL蛋白，包括APPL1和APPL2，是细胞膜与细胞核之间重要的信息传递者，同时也为细胞增殖所必需。在大肠杆菌表达体系中，表达并纯化了人类APPL2蛋白PTB结构域，然后以此为靶标，利用噬菌体展示技术筛选相互作用肽段。经过3轮筛选，随机挑取48个噬菌体单克隆，进行ELISA分析。7肽序列ERLPFFY和YLTSPKH被证明能与PTB结构域特异地结合。对P3GST的野生型和将每个氨基酸突变为丙氨酸的突变型的ELISA检测，显示每个氨基酸均是结合所必需的。这些对有关APPL2的PTB结构域功能的进一步研究有参考意义。

关键词： APPL2; PTB; 表达; 纯化; 噬菌体展示

Abstract:

APPL proteins, including APPL1 and APPL2, are important messengers between cell membrane and cell nucleus and are essential to cell proliferation. In this study, we expressed and purified the PTB domain of human APPL2 (hAPPL2) in Escherichia coli. Phage display technology was then used to screen for the interacting peptides with this domain. After 3 rounds of panning, 48 single phage plaques were randomly picked and ELISA analysis was performed. Two 7-mer peptides, ERLPFFY and YLTSPKH were found to bind to the PTB domain of hAPPL2 in a specific manner. Both wild type and mutant forms of P3-GST fusion protein, with each amino acid substituted by alanine, were prepared and evaluated for binding capacity with hAPPL2 PTB by ELISA. Results showed that each residue was indispensible in the binding between this peptide to hAPPL2 PTB domain. This study shed light on structural interactions between the PTB domain of APPL2 and the binding proteins.

Key words: APPL2 PTB Expression Purification Phage Display

收稿日期: 2009-03-04 出版日期: 2009-07-28

ZTFLH:

Q819

基金资助:

国家“973”计划资助项目（2007CB914301,2006CB910202，2004CB720102）

通讯作者: 梁宁

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	梁宁
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	魏国威
	孔祥平
	陈昌友
	刘劲松
	徐爱民
	吴东海

引用本文:

梁宁,石珠亮,王东业,魏国威,孔祥平,陈昌友,刘劲松,徐爱民,吴东海. 人类APPL2蛋白PTB结构域的表达纯化及其功能的研究[J]. 中国生物工程杂志, 2009, 29(08): 1-7.

LIANG Ning- Dan-Zhu-Liang- Wang-Dong-Ye- Wei-Guo-Wei- Kong-Xiang-Beng- Chen-Chang-You- Liu-Jin-Song- Xu-Ai-Min- Tun-Dong-Hai. Expression,Purification of PTB Domain of Human APPL2 and Screen for Its Peptide Ligands. China Biotechnology, 2009, 29(08): 1-7.

链接本文:

https://manu60.magtech.com.cn/biotech/CN/ 或 https://manu60.magtech.com.cn/biotech/CN/Y2009/V29/I08/1

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