Optical Fluorescence Imaging of Native Proteins Using a Fluorescent Probe with a Cell-Membrane-Permeable Carboxyl Group
Although various methods for selective protein tagging have been established, their ap plications are limited by the low fluorescent tagging efficiency of specific terminal regions of the native proteins of interest (NPIs). In this study, the highly sensitive fluorescence imaging of single NPIs was...
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| Format: | Article |
| Language: | English |
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MDPI AG
2022-05-01
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| Series: | International Journal of Molecular Sciences |
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| Online Access: | https://www.mdpi.com/1422-0067/23/10/5841 |
| _version_ | 1797499128271339520 |
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| author | Jung Min Kim Young-Mi Kang |
| author_facet | Jung Min Kim Young-Mi Kang |
| author_sort | Jung Min Kim |
| collection | DOAJ |
| description | Although various methods for selective protein tagging have been established, their ap plications are limited by the low fluorescent tagging efficiency of specific terminal regions of the native proteins of interest (NPIs). In this study, the highly sensitive fluorescence imaging of single NPIs was demonstrated using a eukaryotic translation mechanism involving a free carboxyl group of a cell-permeable fluorescent dye. In living cells, the carboxyl group of cell-permeable fluorescent dyes reacted with the lysine residues of acceptor peptides (AP or AVI-Tag). Genetically encoded recognition demonstrated that the efficiency of fluorescence labeling was nearly 100%. Nickel-nitrilotriacetic acid (Ni-NTA) beads bound efficiently to a single NPI for detection in a cell without purification. Our labeling approach satisfied the necessary conditions for measuring fluorescently labeled NPI using universal carboxyl fluorescent dyes. This approach is expected to be useful for resolving complex biological/ecological issues and robust single-molecule analyses of dynamic processes, in addition to applications in ultra-sensitive NPIs detection using nanotechnology. |
| first_indexed | 2024-03-10T03:43:01Z |
| format | Article |
| id | doaj.art-f4a7557fc9da4a60a461c75aebaead82 |
| institution | Directory Open Access Journal |
| issn | 1661-6596 1422-0067 |
| language | English |
| last_indexed | 2024-03-10T03:43:01Z |
| publishDate | 2022-05-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | International Journal of Molecular Sciences |
| spelling | doaj.art-f4a7557fc9da4a60a461c75aebaead822023-11-23T11:29:23ZengMDPI AGInternational Journal of Molecular Sciences1661-65961422-00672022-05-012310584110.3390/ijms23105841Optical Fluorescence Imaging of Native Proteins Using a Fluorescent Probe with a Cell-Membrane-Permeable Carboxyl GroupJung Min Kim0Young-Mi Kang1BK21 FOUR R&E Center for Environmental Science and Ecological Engineering, Korea University, 145 Anam-ro, Seongbuk-gu, Seoul 02842, KoreaDepartment of Orthopaedic Surgery, Yonsei University College of Medicine, 50-1 Yonsei-ro, Seodaemun-gu, Seoul 03722, KoreaAlthough various methods for selective protein tagging have been established, their ap plications are limited by the low fluorescent tagging efficiency of specific terminal regions of the native proteins of interest (NPIs). In this study, the highly sensitive fluorescence imaging of single NPIs was demonstrated using a eukaryotic translation mechanism involving a free carboxyl group of a cell-permeable fluorescent dye. In living cells, the carboxyl group of cell-permeable fluorescent dyes reacted with the lysine residues of acceptor peptides (AP or AVI-Tag). Genetically encoded recognition demonstrated that the efficiency of fluorescence labeling was nearly 100%. Nickel-nitrilotriacetic acid (Ni-NTA) beads bound efficiently to a single NPI for detection in a cell without purification. Our labeling approach satisfied the necessary conditions for measuring fluorescently labeled NPI using universal carboxyl fluorescent dyes. This approach is expected to be useful for resolving complex biological/ecological issues and robust single-molecule analyses of dynamic processes, in addition to applications in ultra-sensitive NPIs detection using nanotechnology.https://www.mdpi.com/1422-0067/23/10/5841carboxyl fluorescent dyesingle native proteins of interesthighly efficient fluorescence labelingcell-permeable fluorescent dyebioorthogonal reactionsnickel-nitrilotriacetic acid bead assay |
| spellingShingle | Jung Min Kim Young-Mi Kang Optical Fluorescence Imaging of Native Proteins Using a Fluorescent Probe with a Cell-Membrane-Permeable Carboxyl Group International Journal of Molecular Sciences carboxyl fluorescent dye single native proteins of interest highly efficient fluorescence labeling cell-permeable fluorescent dye bioorthogonal reactions nickel-nitrilotriacetic acid bead assay |
| title | Optical Fluorescence Imaging of Native Proteins Using a Fluorescent Probe with a Cell-Membrane-Permeable Carboxyl Group |
| title_full | Optical Fluorescence Imaging of Native Proteins Using a Fluorescent Probe with a Cell-Membrane-Permeable Carboxyl Group |
| title_fullStr | Optical Fluorescence Imaging of Native Proteins Using a Fluorescent Probe with a Cell-Membrane-Permeable Carboxyl Group |
| title_full_unstemmed | Optical Fluorescence Imaging of Native Proteins Using a Fluorescent Probe with a Cell-Membrane-Permeable Carboxyl Group |
| title_short | Optical Fluorescence Imaging of Native Proteins Using a Fluorescent Probe with a Cell-Membrane-Permeable Carboxyl Group |
| title_sort | optical fluorescence imaging of native proteins using a fluorescent probe with a cell membrane permeable carboxyl group |
| topic | carboxyl fluorescent dye single native proteins of interest highly efficient fluorescence labeling cell-permeable fluorescent dye bioorthogonal reactions nickel-nitrilotriacetic acid bead assay |
| url | https://www.mdpi.com/1422-0067/23/10/5841 |
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