Abstract
Phage display is a powerful technique for rapid construction and screening of peptide libraries with over 109 sequence diversity. The M13 bacteriophage genome can be edited to incorporate randomized amino acids, which will be displayed on its minor coat protein (pIII). To enable screening of nonnatural cyclic peptides on phage, the minor coat protein can be modified with a chemical cross-linker. By taking advantage of the nucleophilicity and low abundance of free cysteines on phage, a variety of cysteine cross-linkers can be installed on the pIII protein. Here, we describe the construction of a chemically modified cyclic phage library through a cysteine cross-linking reagent, 1,3-dichloroacetone (DCA).
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Ahamad S, Bhat SA (2022) Recent update on the development of PCSK9 inhibitors for hypercholesterolemia treatment. J Med Chem 65(23):15513–15539. https://doi.org/10.1021/acs.jmedchem.2c01290
Tombling BJ, Lammi C, Lawrence N, Gilding EK, Grazioso G, Craik DJ, Wang CK (2021) Bioactive cyclization optimizes the affinity of a proprotein convertase subtilisin/kexin type 9 (PCSK9) peptide inhibitor. J Med Chem 64(5):2523–2533. https://doi.org/10.1021/acs.jmedchem.0c01766
Liu H, Zhao Z, Zhang L, Li Y, Jain A, Barve A, Jin W, Liu Y, Fetse J, Cheng K (2019) Discovery of low-molecular weight anti-PD-L1 peptides for cancer immunotherapy. J Immunother Cancer 7(1):270. https://doi.org/10.1186/s40425-019-0705-y
Zheng M, Haeffner F, Gao J (2022) N-terminal cysteine mediated backbone-side chain cyclization for chemically enhanced phage display. Chem Sci 13(28):8349–8354. https://doi.org/10.1039/D2SC03241D
Assem N, Ferreira DJ, Wolan DW, Dawson PE (2015) Acetone-linked peptides: a convergent approach for peptide macrocyclization and labeling. Angew Chem Int Ed 54(30):8665–8668. https://doi.org/10.1002/anie.201502607
Lin Q, Hopper D, Zhang H, Sfyris Qoon J, Shen Z, Karas JA, Hughes RA, Northfield SE (2020) 1,3-Dichloroacetone: a robust reagent for preparing bicyclic peptides. ACS Omega 5(4):1840–1850. https://doi.org/10.1021/acsomega.9b03152
Chen S, Lovell S, Lee S, Fellner M, Mace PD, Bogyo M (2021) Identification of highly selective covalent inhibitors by phage display. Nat Biotechnol 39(4):490–498. https://doi.org/10.1038/s41587-020-0733-7
Zheng M, Chen F-J, Li K, Reja RM, Haeffner F, Gao J (2022) Lysine-targeted reversible covalent ligand discovery for proteins via phage display. J Am Chem Soc 144(34):15885–15893. https://doi.org/10.1021/jacs.2c07375
Diderich P, Bertoldo D, Dessen P, Khan MM, Pizzitola I, Held W, Huelsken J, Heinis C (2016) Phage selection of chemically stabilized α-helical peptide ligands. ACS Chem Biol 11(5):1422–1427. https://doi.org/10.1021/acschembio.5b00963
Jo H, Meinhardt N, Wu Y, Kulkarni S, Hu X, Low KE, Davies PL, DeGrado WF, Greenbaum DC (2012) Development of α-helical calpain probes by mimicking a natural protein–protein interaction. J Am Chem Soc 134(42):17704–17713. https://doi.org/10.1021/ja307599z
Chen F-J, Zheng M, Nobile V, Gao J (2022) Fast and cysteine-specific modification of peptides, proteins and bacteriophage using chlorooximes. Chem Eur J 28(20):e202200058. https://doi.org/10.1002/chem.202200058
Ph.D.â„¢ Phage Display Libraries Instruction Manual (2018) New England Biolabs Inc. https://international.neb.com/-/media/nebus/files/manuals/manuale8100.pdf. Accessed 7 Jan 2023
Instruction Manual Ph.D.â„¢ Peptide Display Cloning System (2022) New England Biolabs Inc. https://www.neb.com/-/media/nebus/files/manuals/manuale8101.pdf?rev=f1e7f087fe7c44aba0e495aba9dd3d13&hash=591221E969ED4D29BF7BE8D9B194ED68. Accessed 20 Dec 2022
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2024 The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature
About this protocol
Cite this protocol
Chau, B., Liivak, K., Gao, J. (2024). Construction of Nonnatural Cysteine-Cross-Linked Phage Libraries. In: Tumban, E. (eds) Bacteriophages. Methods in Molecular Biology, vol 2738. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-3549-0_19
Download citation
DOI: https://doi.org/10.1007/978-1-0716-3549-0_19
Published:
Publisher Name: Humana, New York, NY
Print ISBN: 978-1-0716-3548-3
Online ISBN: 978-1-0716-3549-0
eBook Packages: Springer Protocols