Human IgA-binding Peptides Selected from Random Peptide Libraries AFFINITY MATURATION AND APPLICATION IN IGA PURIFICATION
- Authors
- Hatanaka, Takaaki; Ohzono, Shinji; Park, Mirae; Sakamoto, Kotaro; Tsukamoto, Shogo; Sugita, Ryohei; Ishitobi, Hiroyuki; Mori, Toshiyuki; Ito, Osamu; Sorajo, Koichi; Sugimura, Kazuhisa; Ham, Sihyun; Ito, Yuji
- Issue Date
- Dec-2012
- Publisher
- AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC
- Citation
- JOURNAL OF BIOLOGICAL CHEMISTRY, v.287, no.51, pp 43126 - 43136
- Pages
- 11
- Journal Title
- JOURNAL OF BIOLOGICAL CHEMISTRY
- Volume
- 287
- Number
- 51
- Start Page
- 43126
- End Page
- 43136
- URI
- https://scholarworks.sookmyung.ac.kr/handle/2020.sw.sookmyung/11789
- DOI
- 10.1074/jbc.M112.389742
- ISSN
- 0021-9258
1083-351X
- Abstract
- Phage display system is a powerful tool to design specific ligands for target molecules. Here, we used disulfide-constrained random peptide libraries constructed with the T7 phage display system to isolate peptides specific to human IgA. The binding clones (A1-A4) isolated by biopanning exhibited clear specificity to human IgA, but the synthetic peptide derived from the A2 clone exhibited a low specificity/affinity (K-d = 1.3 mu M). Therefore, we tried to improve the peptide using a partial randomized phage display library and mutational studies on the synthetic peptides. The designed Opt-1 peptide exhibited a 39-fold higher affinity (K-d = 33 nM) than the A2 peptide. An Opt-1 peptide-conjugated column was used to purify IgA from human plasma. However, the recovered IgA fraction was contaminated with other proteins, indicating nonspecific binding. To design a peptide with increased binding specificity, we examined the structural features of Opt-1 and the Opt-1-IgA complex using all-atom molecular dynamics simulations with explicit water. The simulation results revealed that the Opt-1 peptide displayed partial helicity in the N-terminal region and possessed a hydrophobic cluster that played a significant role in tight binding with IgA-Fc. However, these hydrophobic residues of Opt-1 may contribute to nonspecific binding with other proteins. To increase binding specificity, we introduced several mutations in the hydrophobic residues of Opt-1. The resultant Opt-3 peptide exhibited high specificity and high binding affinity for IgA, leading to successful isolation of IgA without contamination.
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