Abstract
Covalently linking together different proteins can enhance functionality for a range of applications. We have developed the SnoopLigase peptide–peptide conjugation method to easily and specifically link proteins fused to the peptides SnoopTagJr or DogTag via an isopeptide bond. SnoopLigase conjugation has been applied for enhancing enzyme resilience and for antigen oligomerization to enhance vaccine efficacy. Following conjugation, SnoopLigase and unreacted substrates can be removed by solid-phase immobilization of SnoopLigase, yielding purified protein–protein conjugates. Here, we describe procedures for designing tag-fused proteins, SnoopLigase purification, and ligation of SnoopTagJr and DogTag. We further define steps for the purification of the ligated product and quantification of ligation success.
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Acknowledgments
Can M. Buldun and Irsyad N.A. Khairil Anuar contributed equally to this work.
Funding for C.M.B. was provided by the Engineering and Physical Sciences Research Council (EPSRC) and Corpus Christi College Oxford. Funding for I.N.A.K.A. was provided by Yayasan Khazanah, Oxford Centre for Islamic Studies, and St. John’s College Oxford. M.H. was funded by the Biotechnology and Biological Sciences Research Council (BBSRC, BB/S007369/1).
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Buldun, C.M., Khairil Anuar, I.N.A., Howarth, M. (2021). SnoopLigase-Mediated Peptide–Peptide Conjugation and Purification. In: Ryadnov, M. (eds) Polypeptide Materials. Methods in Molecular Biology, vol 2208. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-0928-6_2
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DOI: https://doi.org/10.1007/978-1-0716-0928-6_2
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