Abstract
Molecular evolution of binding affinity of artificial peptides by random mutation and affinity selection was performed. DNA fragments encoding ganglioside Galβ1-3GalNAcβ1-4(Neu5Acα2-3)Galβl-4Glcβ1- 1′Cer (GM1)-binding peptides were amplified by error-prone polymerase chain reaction with single strand DNA of phages. This peptide contains artificial zinc-binding sequence to restrict peptide conformation. The amplified DNA fragments were digested with Acc65 I and Eag I, and the fragment was ligated into the M13KE phagemid vector. The vector was used to transform host cells (ER2738) by electroporation and the corresponding phage particles were generated. After two cycles of an affinity selection against GM1 monolayer, three positive phage clones were isolated. Phage ELSIA indicated that these clones had affinity for GM1 at 0.01 nM. The binding affinity of an ee2-1 mutant, ee2-15, altered at His-3 to Leu was found to be enhanced. We showed the binding affinity of peptides for target sugar chain was improved by molecular evolution process.
| Original language | English |
|---|---|
| Title of host publication | Polymer Preprints, Japan |
| Pages | 1824 |
| Number of pages | 1 |
| Volume | 55 |
| Edition | 1 |
| Publication status | Published - 2006 |
| Event | 55th SPSJ Annual Meeting - Nagoya, Japan Duration: 2006 May 24 → 2006 May 26 |
Other
| Other | 55th SPSJ Annual Meeting |
|---|---|
| Country | Japan |
| City | Nagoya |
| Period | 06/5/24 → 06/5/26 |
Fingerprint
Keywords
- Beta structure
- Ganglioside
- GM1
- Molecular evolution
- Peptide library
- Phage display method
ASJC Scopus subject areas
- Engineering(all)
Cite this
Selection of sugar-binding peptides from a scaffold-containing library. / Matsubara, Teruhiko; Sato, Toshinori.
Polymer Preprints, Japan. Vol. 55 1. ed. 2006. p. 1824.Research output: Chapter in Book/Report/Conference proceeding › Conference contribution
}
TY - GEN
T1 - Selection of sugar-binding peptides from a scaffold-containing library
AU - Matsubara, Teruhiko
AU - Sato, Toshinori
PY - 2006
Y1 - 2006
N2 - Molecular evolution of binding affinity of artificial peptides by random mutation and affinity selection was performed. DNA fragments encoding ganglioside Galβ1-3GalNAcβ1-4(Neu5Acα2-3)Galβl-4Glcβ1- 1′Cer (GM1)-binding peptides were amplified by error-prone polymerase chain reaction with single strand DNA of phages. This peptide contains artificial zinc-binding sequence to restrict peptide conformation. The amplified DNA fragments were digested with Acc65 I and Eag I, and the fragment was ligated into the M13KE phagemid vector. The vector was used to transform host cells (ER2738) by electroporation and the corresponding phage particles were generated. After two cycles of an affinity selection against GM1 monolayer, three positive phage clones were isolated. Phage ELSIA indicated that these clones had affinity for GM1 at 0.01 nM. The binding affinity of an ee2-1 mutant, ee2-15, altered at His-3 to Leu was found to be enhanced. We showed the binding affinity of peptides for target sugar chain was improved by molecular evolution process.
AB - Molecular evolution of binding affinity of artificial peptides by random mutation and affinity selection was performed. DNA fragments encoding ganglioside Galβ1-3GalNAcβ1-4(Neu5Acα2-3)Galβl-4Glcβ1- 1′Cer (GM1)-binding peptides were amplified by error-prone polymerase chain reaction with single strand DNA of phages. This peptide contains artificial zinc-binding sequence to restrict peptide conformation. The amplified DNA fragments were digested with Acc65 I and Eag I, and the fragment was ligated into the M13KE phagemid vector. The vector was used to transform host cells (ER2738) by electroporation and the corresponding phage particles were generated. After two cycles of an affinity selection against GM1 monolayer, three positive phage clones were isolated. Phage ELSIA indicated that these clones had affinity for GM1 at 0.01 nM. The binding affinity of an ee2-1 mutant, ee2-15, altered at His-3 to Leu was found to be enhanced. We showed the binding affinity of peptides for target sugar chain was improved by molecular evolution process.
KW - Beta structure
KW - Ganglioside
KW - GM1
KW - Molecular evolution
KW - Peptide library
KW - Phage display method
UR - http://www.scopus.com/inward/record.url?scp=33749817995&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=33749817995&partnerID=8YFLogxK
M3 - Conference contribution
AN - SCOPUS:33749817995
VL - 55
SP - 1824
BT - Polymer Preprints, Japan
ER -