Abstract
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 were 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 elucidated 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.
| Original language | English |
|---|---|
| Pages (from-to) | 197-201 |
| Number of pages | 5 |
| Journal | Biochemical and Biophysical Research Communications |
| Volume | 386 |
| Issue number | 1 |
| DOIs | |
| State | Published - Aug 14 2009 |
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Keywords
- ORF phage display
- Phage display
- Phosphatidylcholine
- Phosphatidylserine
- Phosphatidylserine-binding protein
ASJC Scopus subject areas
- Biochemistry
- Biophysics
- Cell Biology
- Molecular Biology
Cite this
Efficient identification of phosphatidylserine-binding proteins by ORF phage display. / Caberoy, Nora B.; Zhou, Yixiong; Alvarado, Gabriela; Fan, Xianqun; Li, Wei.
In: Biochemical and Biophysical Research Communications, Vol. 386, No. 1, 14.08.2009, p. 197-201.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Efficient identification of phosphatidylserine-binding proteins by ORF phage display
AU - Caberoy, Nora B.
AU - Zhou, Yixiong
AU - Alvarado, Gabriela
AU - Fan, Xianqun
AU - Li, Wei
PY - 2009/8/14
Y1 - 2009/8/14
N2 - 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 were 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 elucidated 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.
AB - 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 were 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 elucidated 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.
KW - ORF phage display
KW - Phage display
KW - Phosphatidylcholine
KW - Phosphatidylserine
KW - Phosphatidylserine-binding protein
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U2 - 10.1016/j.bbrc.2009.06.010
DO - 10.1016/j.bbrc.2009.06.010
M3 - Article
C2 - 19520055
AN - SCOPUS:67649229597
VL - 386
SP - 197
EP - 201
JO - Biochemical and Biophysical Research Communications
JF - Biochemical and Biophysical Research Communications
SN - 0006-291X
IS - 1
ER -