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Biology 2015, 4(1), 161-172; doi:10.3390/biology4010161

Easy and Rapid Binding Assay for Functional Analysis of Disulfide-Containing Peptides by a Pull-Down Method Using a Puromycin-Linker and a Cell-Free Translation System

Graduate School of Science and Engineering, Saitama University, Sakura-ku, Saitama 338-8570, Japan
These authors are equally contributed to this work.
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Academic Editor: Thorsten Berg
Received: 20 December 2014 / Revised: 11 February 2015 / Accepted: 17 February 2015 / Published: 2 March 2015
(This article belongs to the Special Issue Protein-Protein Interactions)
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Abstract

Constrained peptides are an attractive class as affinity reagents or drug leads owing to their excellent binding properties. Many kinds of these peptides, such as cyclic peptides containing disulfide bridges, are found in nature or designed artificially by directed evolution. However, confirming the binding properties of the disulfide-rich peptides can be generally difficult, because of oxidative folding problems in the preparation steps. Therefore, a method for evaluating the binding properties of such peptides rapidly and easily is required. Here, we report an easy and rapid method for preparing biotin-attached peptides containing disulfide bridges or a chemical cross-linker using a cell-free translation system and a puromycin-linker, which is applicable to pull-down assays for protein (or peptide) molecular interaction analysis. View Full-Text
Keywords: molecular interaction analysis; pull-down assay; cell-free translation system; puromycin; constrained peptide; cyclic peptide; disulfide-rich peptide; cross-linking; in vitro selection; directed evolution molecular interaction analysis; pull-down assay; cell-free translation system; puromycin; constrained peptide; cyclic peptide; disulfide-rich peptide; cross-linking; in vitro selection; directed evolution
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This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. (CC BY 4.0).

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MDPI and ACS Style

Tanemura, Y.; Mochizuki, Y.; Kumachi, S.; Nemoto, N. Easy and Rapid Binding Assay for Functional Analysis of Disulfide-Containing Peptides by a Pull-Down Method Using a Puromycin-Linker and a Cell-Free Translation System. Biology 2015, 4, 161-172.

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