Third‐Generation Covalent TMP‐Tag for Fast Labeling and Multiplexed Imaging of Cellular Proteins

Mo, Jiaming; Chen, Jingting; Shi, Yabo; Sun, Jingfu; Wu, Yunxiang; Liu, Tianyan; Zhang, Junwei; Zheng, Yan‐Zhen; Li, Yulong; Chen, Zhuo

doi:10.1002/anie.202207905

Cited by 13 publications

(17 citation statements)

References 63 publications

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“…Confocal imaging of FKBP-fused COX8A colocalized with Mito-Tracker Green (Figure S24a). However, fusion of COX8A to HaloTag did not result in correct localization to the mitochondria (Figure S24b). Nuclear reader protein YTHDC1 undergoes liquid–liquid phase separation, distributed as nuclear puncta in living cells. , We fused YTHDC1 to different tags to examine whether tag size could affect the distribution of YTHDC1.…”

Section: Resultsmentioning

confidence: 99%

“…With the development of versatile protein-labeling techniques, live-cell fluorescence imaging has emerged as a routine tool to investigate protein function and localization. Recent resurgence of small molecular dyes − and the development of self-labeling tags, which leverage the site-specific conjugation of fluorophores, − have expanded the fluorescence imaging toolbox. Compared to widely adopted fluorescent proteins, self-labeling tags provide a more flexible choice of the spectral characteristics of the dyes, higher brightness, and photostability.…”

Section: Introductionmentioning

confidence: 99%

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Self-Renewable Tag for Photostable Fluorescence Imaging of Proteins

Du,

Wang,

Luo

et al. 2023

J. Am. Chem. Soc.

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We report the development of a self-renewable tag (srTAG) for protein fluorescence imaging. srTAG leverages the "on-protein" fluorophore equilibrium between the fluorescent zwitterion and non-fluorescent spirocyclic form and the reversible fluorescence labeling to enable self-recovery of fluorescence after photobleaching. This small-sized srTAG allows 2−6 times longer imaging duration compared to other commonly used self-labeling tags and is compatible with fluorophores with different spectral properties. This study provides a new strategy for fine tuning of self-labeling tags.

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Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Self-Renewable Tag for Photostable Fluorescence Imaging of Proteins

Du,

Wang,

Luo

et al. 2023

J. Am. Chem. Soc.

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“…[47] TMP, acrylamide, and the fluorophore are all components of this third-generation tag with close distance. [47] On the one hand, the linker between the dye and adapter is kept to a minimum because modern spironolactone/lactam rhodamine renders fluorogenicity by using residues on the surface of the protein. [48] On the other hand, TMP was functionalized with a two-carbon amine, and then immediately acrylate was attached to this nitrogen.…”

Section: Snap/clip Tagmentioning

confidence: 99%

Chemical tags and beyond: Live‐cell protein labeling technologies for modern optical imaging

Saimi,

Chen

2023

Smart Molecules

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Imaging proteins with high resolution is crucial for studying cellular physiology and pathology. Fluorescence imaging is a privileged method to visualize proteins with subcellular precision in live cells. In recent years, there has been a tremendous advance in the field of fluorescent dyes that are optically more sophisticated than genetically‐encodable fluorescent proteins. In this review, we aim to discuss modern bioconjugation methods to specifically incorporate these dyes into protein‐of‐interests. We focus on advances in live‐cell labeling strategies and fluorescent probes, especially the HaloTag, SNAP‐tag, TMP‐tag, and unnatural amino acid systems and their applications. These protein labeling methods, along with cutting‐edge dyes and novel microscopy methods, have become the infrastructure for biological research in the era of super‐resolution imaging.

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“…Protein recognition, particularly protein isoform-specific recognition, is of great significance for accurate clinical diagnosis and academic research. Among many biological analysis methods, organic fluorescent probes are widely used to recognize proteins in living simples due to their high selectivity, remarkable sensitivity, excellent spatial resolution, noninvasiveness, and fast and easy operation. − …”

mentioning

confidence: 99%

Molecular Design Strategy of Protein Isoform-Specific Fluorescent Probes by Considering Molecule in Its Entirety

Zang,

Wang,

Zhang

et al. 2023

Anal. Chem.

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Generally, different isoforms of proteins exert separate biological functions. However, due to similar structures and identical catalysis functions, distinguishing isoforms is challenging. Summarizing a molecular design strategy has great significance in developing a protein-specific fluorescent probe. Usually, recognition of a group was deemed to be the key to a protein isoform-specific response. However, some novel literature reported that fluorophore could play a vital role in the protein isoform-specific response. It means that any part of the fluorescent probe could affect the detected properties. In this work, we report the generation of the first probe to specifically recognize HexA(β-N-acetylhexosaminidase A), Hex-C4, by adjusting the length of the linker. Hex-C4 exhibits specific recognition of HexA both in vitro and in living cells. The integration of the fluorescent spectrum and the MD (molecular dynamics) results provide two factors for the molecular design of isoform-specific fluorescent probes. One is the interaction between tetraphenyl ethylene (AIE fluorogen) and amino acid residues, and the other is the interaction between amino acid residues and the binding group. In this work, a powerful tool to detect HexA in living cells is reported for the first time. Further, a workable molecular design strategy for protein isoform-specific fluorescent probes is summarized.

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Third‐Generation Covalent TMP‐Tag for Fast Labeling and Multiplexed Imaging of Cellular Proteins

Cited by 13 publications

References 63 publications

Self-Renewable Tag for Photostable Fluorescence Imaging of Proteins

Self-Renewable Tag for Photostable Fluorescence Imaging of Proteins

Chemical tags and beyond: Live‐cell protein labeling technologies for modern optical imaging

Molecular Design Strategy of Protein Isoform-Specific Fluorescent Probes by Considering Molecule in Its Entirety

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