Xenoprotein engineering via synthetic libraries
Chemical methods have enabled the total synthesis of protein molecules of ever-increasing size and complexity. However, methods to engineer synthetic proteins comprising noncanonical amino acids have not kept pace, even though this capability would be a distinct advantage of the total synthesis appr...
| Main Authors: | , , , , , , , , , , , , , , , , , |
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| Format: | Article |
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National Academy of Sciences (U.S.)
2019
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| Online Access: | http://hdl.handle.net/1721.1/120509 https://orcid.org/0000-0003-0588-6157 https://orcid.org/0000-0002-5508-0963 https://orcid.org/0000-0003-4995-1424 https://orcid.org/0000-0002-9383-2185 https://orcid.org/0000-0001-6226-8625 https://orcid.org/0000-0001-5474-4273 https://orcid.org/0000-0003-1632-5195 https://orcid.org/0000-0002-6214-7513 https://orcid.org/0000-0003-2315-6983 https://orcid.org/0000-0001-9021-0308 https://orcid.org/0000-0002-8434-0066 https://orcid.org/0000-0002-8601-7799 |
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| author | Gates, Zachary P Vinogradov, Alexander Alexandrovich Quartararo, Anthony James Bandyopadhyay, Anupam Choo, Zi-Ning Evans, Ethan Daniel Halloran, Kathryn Mijalis, Alexander James Mong, Surin Khai Simon, Mark Standley, Eric Alan Styduhar, Evan Tasker, Sarah Zinnen Touti, Faycal Weber, Jessica Marie Wilson, Jessica Laura Jamison, Timothy F Pentelute, Bradley L. |
| author2 | Massachusetts Institute of Technology. Department of Chemistry |
| author_facet | Massachusetts Institute of Technology. Department of Chemistry Gates, Zachary P Vinogradov, Alexander Alexandrovich Quartararo, Anthony James Bandyopadhyay, Anupam Choo, Zi-Ning Evans, Ethan Daniel Halloran, Kathryn Mijalis, Alexander James Mong, Surin Khai Simon, Mark Standley, Eric Alan Styduhar, Evan Tasker, Sarah Zinnen Touti, Faycal Weber, Jessica Marie Wilson, Jessica Laura Jamison, Timothy F Pentelute, Bradley L. |
| author_sort | Gates, Zachary P |
| collection | MIT |
| description | Chemical methods have enabled the total synthesis of protein molecules of ever-increasing size and complexity. However, methods to engineer synthetic proteins comprising noncanonical amino acids have not kept pace, even though this capability would be a distinct advantage of the total synthesis approach to protein science. In this work, we report a platform for protein engineering based on the screening of synthetic one-bead one-compound protein libraries. Screening throughput approaching that of cell surface display was achieved by a combination of magnetic bead enrichment, flow cytometry analysis of on-bead screens, and high-throughput MS/MS-based sequencing of identified active compounds. Direct screening of a synthetic protein library by these methods resulted in the de novo discovery of mirror-image miniprotein-based binders to a ∼150-kDa protein target, a task that would be difficult or impossible by other means. Keywords: xenoprotein; mirror-image miniprotein; D-protein; protein engineering; flow cytometry |
| first_indexed | 2024-09-23T12:57:11Z |
| format | Article |
| id | mit-1721.1/120509 |
| institution | Massachusetts Institute of Technology |
| last_indexed | 2024-09-23T12:57:11Z |
| publishDate | 2019 |
| publisher | National Academy of Sciences (U.S.) |
| record_format | dspace |
| spelling | mit-1721.1/1205092022-10-01T12:07:18Z Xenoprotein engineering via synthetic libraries Gates, Zachary P Vinogradov, Alexander Alexandrovich Quartararo, Anthony James Bandyopadhyay, Anupam Choo, Zi-Ning Evans, Ethan Daniel Halloran, Kathryn Mijalis, Alexander James Mong, Surin Khai Simon, Mark Standley, Eric Alan Styduhar, Evan Tasker, Sarah Zinnen Touti, Faycal Weber, Jessica Marie Wilson, Jessica Laura Jamison, Timothy F Pentelute, Bradley L. Massachusetts Institute of Technology. Department of Chemistry Gates, Zachary P Vinogradov, Alexander Alexandrovich Quartararo, Anthony James Bandyopadhyay, Anupam Choo, Zi-Ning Evans, Ethan Daniel Halloran, Kathryn Mijalis, Alexander James Mong, Surin Khai Simon, Mark Standley, Eric Alan Styduhar, Evan Tasker, Sarah Zinnen Touti, Faycal Weber, Jessica Marie Wilson, Jessica Laura Jamison, Timothy F Pentelute, Bradley L. Chemical methods have enabled the total synthesis of protein molecules of ever-increasing size and complexity. However, methods to engineer synthetic proteins comprising noncanonical amino acids have not kept pace, even though this capability would be a distinct advantage of the total synthesis approach to protein science. In this work, we report a platform for protein engineering based on the screening of synthetic one-bead one-compound protein libraries. Screening throughput approaching that of cell surface display was achieved by a combination of magnetic bead enrichment, flow cytometry analysis of on-bead screens, and high-throughput MS/MS-based sequencing of identified active compounds. Direct screening of a synthetic protein library by these methods resulted in the de novo discovery of mirror-image miniprotein-based binders to a ∼150-kDa protein target, a task that would be difficult or impossible by other means. Keywords: xenoprotein; mirror-image miniprotein; D-protein; protein engineering; flow cytometry United States. Defense Advanced Research Projects Agency (Award 023504-001) 2019-02-20T16:00:21Z 2019-02-20T16:00:21Z 2018-05 2017-12 2019-02-08T16:00:09Z Article http://purl.org/eprint/type/JournalArticle 0027-8424 1091-6490 http://hdl.handle.net/1721.1/120509 Gates, Zachary P. et al. “Xenoprotein Engineering via Synthetic Libraries.” Proceedings of the National Academy of Sciences 115, 23 (May 2018): E5298–E5306 © 2018 National Academy of Sciences https://orcid.org/0000-0003-0588-6157 https://orcid.org/0000-0002-5508-0963 https://orcid.org/0000-0003-4995-1424 https://orcid.org/0000-0002-9383-2185 https://orcid.org/0000-0001-6226-8625 https://orcid.org/0000-0001-5474-4273 https://orcid.org/0000-0003-1632-5195 https://orcid.org/0000-0002-6214-7513 https://orcid.org/0000-0003-2315-6983 https://orcid.org/0000-0001-9021-0308 https://orcid.org/0000-0002-8434-0066 https://orcid.org/0000-0002-8601-7799 http://dx.doi.org/10.1073/PNAS.1722633115 Proceedings of the National Academy of Sciences 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 | Gates, Zachary P Vinogradov, Alexander Alexandrovich Quartararo, Anthony James Bandyopadhyay, Anupam Choo, Zi-Ning Evans, Ethan Daniel Halloran, Kathryn Mijalis, Alexander James Mong, Surin Khai Simon, Mark Standley, Eric Alan Styduhar, Evan Tasker, Sarah Zinnen Touti, Faycal Weber, Jessica Marie Wilson, Jessica Laura Jamison, Timothy F Pentelute, Bradley L. Xenoprotein engineering via synthetic libraries |
| title | Xenoprotein engineering via synthetic libraries |
| title_full | Xenoprotein engineering via synthetic libraries |
| title_fullStr | Xenoprotein engineering via synthetic libraries |
| title_full_unstemmed | Xenoprotein engineering via synthetic libraries |
| title_short | Xenoprotein engineering via synthetic libraries |
| title_sort | xenoprotein engineering via synthetic libraries |
| url | http://hdl.handle.net/1721.1/120509 https://orcid.org/0000-0003-0588-6157 https://orcid.org/0000-0002-5508-0963 https://orcid.org/0000-0003-4995-1424 https://orcid.org/0000-0002-9383-2185 https://orcid.org/0000-0001-6226-8625 https://orcid.org/0000-0001-5474-4273 https://orcid.org/0000-0003-1632-5195 https://orcid.org/0000-0002-6214-7513 https://orcid.org/0000-0003-2315-6983 https://orcid.org/0000-0001-9021-0308 https://orcid.org/0000-0002-8434-0066 https://orcid.org/0000-0002-8601-7799 |
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