Quantum Dot Targeting with Lipoic Acid Ligase and HaloTag for Single-Molecule Imaging on Living Cells
We present a methodology for targeting quantum dots to specific proteins on living cells in two steps. In the first step, Escherichia coli lipoic acid ligase (LplA) site-specifically attaches 10-bromodecanoic acid onto a 13 amino acid recognition sequence that is genetically fused to a protein of in...
| Main Authors: | , , , , |
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| Format: | Article |
| Language: | en_US |
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American Chemical Society (ACS)
2015
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| Online Access: | http://hdl.handle.net/1721.1/95744 https://orcid.org/0000-0003-4994-2392 https://orcid.org/0000-0002-8277-5226 |
| _version_ | 1826202224903061504 |
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| author | Liu, Daniel S. Phipps, William S. Loh, Ken H. Howarth, Mark Ting, Alice Y. |
| author2 | Massachusetts Institute of Technology. Department of Chemistry |
| author_facet | Massachusetts Institute of Technology. Department of Chemistry Liu, Daniel S. Phipps, William S. Loh, Ken H. Howarth, Mark Ting, Alice Y. |
| author_sort | Liu, Daniel S. |
| collection | MIT |
| description | We present a methodology for targeting quantum dots to specific proteins on living cells in two steps. In the first step, Escherichia coli lipoic acid ligase (LplA) site-specifically attaches 10-bromodecanoic acid onto a 13 amino acid recognition sequence that is genetically fused to a protein of interest. In the second step, quantum dots derivatized with HaloTag, a modified haloalkane dehalogenase, react with the ligated bromodecanoic acid to form a covalent adduct. We found this targeting method to be specific, fast, and fully orthogonal to a previously reported and analogous quantum dot targeting method using E. coli biotin ligase and streptavidin. We used these two methods in combination for two-color quantum dot visualization of different proteins expressed on the same cell or on neighboring cells. Both methods were also used to track single molecules of neurexin, a synaptic adhesion protein, to measure its lateral diffusion in the presence of neuroligin, its trans-synaptic adhesion partner. |
| first_indexed | 2024-09-23T12:04:09Z |
| format | Article |
| id | mit-1721.1/95744 |
| institution | Massachusetts Institute of Technology |
| language | en_US |
| last_indexed | 2024-09-23T12:04:09Z |
| publishDate | 2015 |
| publisher | American Chemical Society (ACS) |
| record_format | dspace |
| spelling | mit-1721.1/957442022-10-01T07:54:43Z Quantum Dot Targeting with Lipoic Acid Ligase and HaloTag for Single-Molecule Imaging on Living Cells Liu, Daniel S. Phipps, William S. Loh, Ken H. Howarth, Mark Ting, Alice Y. Massachusetts Institute of Technology. Department of Chemistry Ting, Alice Y. Liu, Daniel S. Phipps, William S. Loh, Ken H. Howarth, Mark We present a methodology for targeting quantum dots to specific proteins on living cells in two steps. In the first step, Escherichia coli lipoic acid ligase (LplA) site-specifically attaches 10-bromodecanoic acid onto a 13 amino acid recognition sequence that is genetically fused to a protein of interest. In the second step, quantum dots derivatized with HaloTag, a modified haloalkane dehalogenase, react with the ligated bromodecanoic acid to form a covalent adduct. We found this targeting method to be specific, fast, and fully orthogonal to a previously reported and analogous quantum dot targeting method using E. coli biotin ligase and streptavidin. We used these two methods in combination for two-color quantum dot visualization of different proteins expressed on the same cell or on neighboring cells. Both methods were also used to track single molecules of neurexin, a synaptic adhesion protein, to measure its lateral diffusion in the presence of neuroligin, its trans-synaptic adhesion partner. National Institutes of Health (U.S.) (R01 GM072670) Camille & Henry Dreyfus Foundation Massachusetts Institute of Technology. Computational and Systems Biology Program. MIT-Merck Postdoctoral Fellowship 2015-03-03T15:58:26Z 2015-03-03T15:58:26Z 2012-11 2012-10 Article http://purl.org/eprint/type/JournalArticle 1936-0851 1936-086X http://hdl.handle.net/1721.1/95744 Liu, Daniel S., William S. Phipps, Ken H. Loh, Mark Howarth, and Alice Y. Ting. “Quantum Dot Targeting with Lipoic Acid Ligase and HaloTag for Single-Molecule Imaging on Living Cells.” ACS Nano (December 5, 2012): 11080-11087. https://orcid.org/0000-0003-4994-2392 https://orcid.org/0000-0002-8277-5226 en_US http://dx.doi.org/10.1021/nn304793z ACS Nano Creative Commons Attribution-Noncommercial-Share Alike http://creativecommons.org/licenses/by-nc-sa/4.0/ application/pdf American Chemical Society (ACS) PMC |
| spellingShingle | Liu, Daniel S. Phipps, William S. Loh, Ken H. Howarth, Mark Ting, Alice Y. Quantum Dot Targeting with Lipoic Acid Ligase and HaloTag for Single-Molecule Imaging on Living Cells |
| title | Quantum Dot Targeting with Lipoic Acid Ligase and HaloTag for Single-Molecule Imaging on Living Cells |
| title_full | Quantum Dot Targeting with Lipoic Acid Ligase and HaloTag for Single-Molecule Imaging on Living Cells |
| title_fullStr | Quantum Dot Targeting with Lipoic Acid Ligase and HaloTag for Single-Molecule Imaging on Living Cells |
| title_full_unstemmed | Quantum Dot Targeting with Lipoic Acid Ligase and HaloTag for Single-Molecule Imaging on Living Cells |
| title_short | Quantum Dot Targeting with Lipoic Acid Ligase and HaloTag for Single-Molecule Imaging on Living Cells |
| title_sort | quantum dot targeting with lipoic acid ligase and halotag for single molecule imaging on living cells |
| url | http://hdl.handle.net/1721.1/95744 https://orcid.org/0000-0003-4994-2392 https://orcid.org/0000-0002-8277-5226 |
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