Preparation of unnatural N-to-N and C-to-C protein fusions
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...
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| Format: | Article |
| Language: | en_US |
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National Academy of Sciences (U.S.)
2013
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| Online Access: | http://hdl.handle.net/1721.1/76755 https://orcid.org/0000-0002-1090-6071 |
| _version_ | 1811090437827461120 |
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| author | Witte, Martin D. Cragnolini, Juan J. Dougan, Stephanie K. Yoder, Nicholas C. Ploegh, Hidde Popp, Maximilian W.L. |
| author2 | Massachusetts Institute of Technology. Department of Biology |
| author_facet | Massachusetts Institute of Technology. Department of Biology Witte, Martin D. Cragnolini, Juan J. Dougan, Stephanie K. Yoder, Nicholas C. Ploegh, Hidde Popp, Maximilian W.L. |
| author_sort | Witte, Martin D. |
| collection | MIT |
| description | 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. |
| first_indexed | 2024-09-23T14:45:21Z |
| format | Article |
| id | mit-1721.1/76755 |
| institution | Massachusetts Institute of Technology |
| language | en_US |
| last_indexed | 2024-09-23T14:45:21Z |
| publishDate | 2013 |
| publisher | National Academy of Sciences (U.S.) |
| record_format | dspace |
| spelling | mit-1721.1/767552022-09-29T10:23:02Z Preparation of unnatural N-to-N and C-to-C protein fusions Witte, Martin D. Cragnolini, Juan J. Dougan, Stephanie K. Yoder, Nicholas C. Ploegh, Hidde Popp, Maximilian W.L. Massachusetts Institute of Technology. Department of Biology Whitehead Institute for Biomedical Research Ploegh, Hidde Witte, Martin D. Cragnolini, Juan J. Dougan, Stephanie K. Yoder, Nicholas C. Popp, Maximilian W. 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. Netherlands Organization for Scientific Research Human Frontier Science Program (Strasbourg, France) National Institutes of Health (U.S.) (R01 Award AI033456) National Institutes of Health (U.S.) (R01 Award AI087879) 2013-02-06T20:25:01Z 2013-02-06T20:25:01Z 2012-07 2012-03 Article http://purl.org/eprint/type/JournalArticle 0027-8424 1091-6490 http://hdl.handle.net/1721.1/76755 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. https://orcid.org/0000-0002-1090-6071 en_US http://dx.doi.org/10.1073/pnas.1205427109 Proceedings of the National Academy of Sciences of the United States of America Article is made available in accordance with the publisher's policy and may be subject to US copyright law. Please refer to the publisher's site for terms of use. application/pdf National Academy of Sciences (U.S.) PNAS |
| spellingShingle | Witte, Martin D. Cragnolini, Juan J. Dougan, Stephanie K. Yoder, Nicholas C. Ploegh, Hidde Popp, Maximilian W.L. Preparation of unnatural N-to-N and C-to-C protein fusions |
| title | Preparation of unnatural N-to-N and C-to-C protein fusions |
| title_full | Preparation of unnatural N-to-N and C-to-C protein fusions |
| title_fullStr | Preparation of unnatural N-to-N and C-to-C protein fusions |
| title_full_unstemmed | Preparation of unnatural N-to-N and C-to-C protein fusions |
| title_short | Preparation of unnatural N-to-N and C-to-C protein fusions |
| title_sort | preparation of unnatural n to n and c to c protein fusions |
| url | http://hdl.handle.net/1721.1/76755 https://orcid.org/0000-0002-1090-6071 |
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