Preparation of unnatural N-to-N and C-to-C protein fusions
Author(s)
Witte, Martin D.; Cragnolini, Juan J.; Dougan, Stephanie K.; Yoder, Nicholas C.; Ploegh, Hidde; Popp, Maximilian W.L.; ... Show more Show less
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Standard genetic approaches allow the production of protein composites by fusion of polypeptides in head-to-tail fashion. Some applications would benefit from constructions that are genetically impossible, such as the site-specific linkage of proteins via their N or C termini, when a remaining free terminus is required for biological activity. We developed a method for the production of N-to-N and C-to-C dimers, with full retention of the biological activity of both fusion partners and without inflicting chemical damage on the proteins to be joined. We use sortase A to install on the N or C terminus of proteins of interest the requisite modifications to execute a strain-promoted copper-free cycloaddition and show that the ensuing ligation proceeds efficiently. Applied here to protein–protein fusions, the method reported can be extended to connecting proteins with any entity of interest.
Date issued
2012-07Department
Massachusetts Institute of Technology. Department of Biology; Whitehead Institute for Biomedical ResearchJournal
Proceedings of the National Academy of Sciences of the United States of America
Publisher
National Academy of Sciences (U.S.)
Citation
Witte, M. D. et al. “Preparation of Unnatural N-to-N and C-to-C Protein Fusions.” Proceedings of the National Academy of Sciences 109.30 (2012): 11993–11998. Web. © 2012 National Academy of Sciences.
Version: Final published version
ISSN
0027-8424
1091-6490