Efficient identification of phosphatidylserine-binding proteins by ORF phage display.
Efficient identification of phosphatidylserine-binding proteins by ORF phage display.
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DOI：
10.1016/j.bbrc.2009.06.010
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发表时间：
2009-08-14
影响因子：
3.1
通讯作者：
Li, Wei
中科院分区：
文献类型：
--
作者：
Caberoy, Nora B.;Zhou, Yixiong;Alvarado, Gabriela;Fan, Xianqun;Li, Wei
关键词：
为了有效地阐明磷脂酰丝氨酸（PS）的生物学作用，我们开发了开放阅读框（ORF）噬菌体展示来鉴定PS结合蛋白。利用表达ps结合蛋白MFG-E8的噬菌体克隆优化噬菌体筛选程序。利用ORF噬菌体展示cDNA文库进行3轮噬菌体筛选，使其ps结合活性提高了约300倍。共鉴定出17个ps结合噬菌体克隆。与传统cDNA文库的噬菌体展示不同，所有17个ps结合克隆都是编码13种真实蛋白质的orf。序列分析表明，所有鉴定的ps特异性噬菌体克隆均含有二聚体碱性氨基酸残基。GST融合蛋白表达了3种PS结合蛋白，并验证了它们与PS脂质体的结合活性，但不与磷脂酰胆碱脂质体结合。这些结果阐明了以前未知的PS结合蛋白，并证明ORF噬菌体展示是一种多功能技术，能够有效地识别PS等非蛋白分子的结合蛋白。
To efficiently elucidate the biological roles of phosphatidylserine (PS), we developed open-reading-frame (ORF) phage display to identify PS-binding proteins. The procedure of phage panning was optimized with a phage clone expressing MFG-E8, a well-known PS-binding protein. Three rounds of phage panning with ORF phage display cDNA library resulted in ~300-fold enrichment in PS-binding activity. A total of 17 PS-binding phage clones were identified. Unlike phage display with conventional cDNA libraries, all 17 PS-binding clones are ORFs encoding 13 real proteins. Sequence analysis revealed that all identified PS-specific phage clones had dimeric basic amino acid residues. GST fusion proteins were expressed for 3 PS-binding proteins and verified for their binding activity to PS liposomes, but not phosphatidylcholine liposomes. These results elucidate previously unknown PS-binding proteins and demonstrated that ORF phage display is a versatile technology capable of efficiently identifying binding proteins for non-protein molecules like PS.
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影响因子：
4.4
作者：
Gustincich, S;Vatta, P;Schneider, C
通讯作者：
Schneider, C
DOI：
10.1016/s1570-9639(02)00402-8
发表时间：
2002-09-23
影响因子：
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作者：
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通讯作者：
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影响因子：
64.8
作者：
Park, Daeho;Tosello-Trampont, Annie-Carole;Ravichandran, Kodi S.
通讯作者：
Ravichandran, Kodi S.