Kinetic Controlled Tag-Catcher Interactions for Directed Covalent Protein Assembly.
Over the last few years, a number of different protein assembly strategies have been developed, greatly expanding the toolbox for controlling macromolecular assembly. One of the most promising developments is a rapid protein ligation approach using a short polypeptide SpyTag and its partner, SpyCatc...
| Main Authors: | , , |
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| Format: | Article |
| Language: | English |
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Public Library of Science (PLoS)
2016-01-01
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| Series: | PLoS ONE |
| Online Access: | http://europepmc.org/articles/PMC5082641?pdf=render |
| _version_ | 1818675211133779968 |
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| author | Lee Ling Tan Shawn S Hoon Fong T Wong |
| author_facet | Lee Ling Tan Shawn S Hoon Fong T Wong |
| author_sort | Lee Ling Tan |
| collection | DOAJ |
| description | Over the last few years, a number of different protein assembly strategies have been developed, greatly expanding the toolbox for controlling macromolecular assembly. One of the most promising developments is a rapid protein ligation approach using a short polypeptide SpyTag and its partner, SpyCatcher derived from Streptococcus pyogenes fibronectin-binding protein, FbaB. To extend this technology, we have engineered and characterized a new Tag-Catcher pair from a related fibronectin-binding protein in Streptococcus dysgalactiae. The polypeptide Tag, named SdyTag, was constructed based on the native Cna protein B-type (CnaB) domain and was found to be highly unreactive to SpyCatcher. SpyCatcher has 320-fold specificity for its native SpyTag compared to SdyTag. Similarly, SdyTag has a 75-fold specificity for its optimized Catcher, named SdyCatcherDANG short, compared to SpyCatcher. These Tag-Catcher pairs were used in combination to demonstrate specific sequential assembly of tagged proteins in vitro. We also demonstrated that the in vivo generation of circularized proteins in a Tag-Catcher specific manner where specific Tags can be left unreacted for use in subsequent ligation reactions. From the success of these experiments, we foresee the application of SdyTags and SpyTags, not only, for multiplexed control of protein assembly but also for the construction of novel protein architectures. |
| first_indexed | 2024-12-17T08:23:58Z |
| format | Article |
| id | doaj.art-caf79bf74ad24edc8ea6275ef9ddde97 |
| institution | Directory Open Access Journal |
| issn | 1932-6203 |
| language | English |
| last_indexed | 2024-12-17T08:23:58Z |
| publishDate | 2016-01-01 |
| publisher | Public Library of Science (PLoS) |
| record_format | Article |
| series | PLoS ONE |
| spelling | doaj.art-caf79bf74ad24edc8ea6275ef9ddde972022-12-21T21:56:50ZengPublic Library of Science (PLoS)PLoS ONE1932-62032016-01-011110e016507410.1371/journal.pone.0165074Kinetic Controlled Tag-Catcher Interactions for Directed Covalent Protein Assembly.Lee Ling TanShawn S HoonFong T WongOver the last few years, a number of different protein assembly strategies have been developed, greatly expanding the toolbox for controlling macromolecular assembly. One of the most promising developments is a rapid protein ligation approach using a short polypeptide SpyTag and its partner, SpyCatcher derived from Streptococcus pyogenes fibronectin-binding protein, FbaB. To extend this technology, we have engineered and characterized a new Tag-Catcher pair from a related fibronectin-binding protein in Streptococcus dysgalactiae. The polypeptide Tag, named SdyTag, was constructed based on the native Cna protein B-type (CnaB) domain and was found to be highly unreactive to SpyCatcher. SpyCatcher has 320-fold specificity for its native SpyTag compared to SdyTag. Similarly, SdyTag has a 75-fold specificity for its optimized Catcher, named SdyCatcherDANG short, compared to SpyCatcher. These Tag-Catcher pairs were used in combination to demonstrate specific sequential assembly of tagged proteins in vitro. We also demonstrated that the in vivo generation of circularized proteins in a Tag-Catcher specific manner where specific Tags can be left unreacted for use in subsequent ligation reactions. From the success of these experiments, we foresee the application of SdyTags and SpyTags, not only, for multiplexed control of protein assembly but also for the construction of novel protein architectures.http://europepmc.org/articles/PMC5082641?pdf=render |
| spellingShingle | Lee Ling Tan Shawn S Hoon Fong T Wong Kinetic Controlled Tag-Catcher Interactions for Directed Covalent Protein Assembly. PLoS ONE |
| title | Kinetic Controlled Tag-Catcher Interactions for Directed Covalent Protein Assembly. |
| title_full | Kinetic Controlled Tag-Catcher Interactions for Directed Covalent Protein Assembly. |
| title_fullStr | Kinetic Controlled Tag-Catcher Interactions for Directed Covalent Protein Assembly. |
| title_full_unstemmed | Kinetic Controlled Tag-Catcher Interactions for Directed Covalent Protein Assembly. |
| title_short | Kinetic Controlled Tag-Catcher Interactions for Directed Covalent Protein Assembly. |
| title_sort | kinetic controlled tag catcher interactions for directed covalent protein assembly |
| url | http://europepmc.org/articles/PMC5082641?pdf=render |
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