Improved Stabilities of Labeling Probes for the Selective Modification of Endogenous Proteins in Living Cells and In Vivo

Lin, Kuan‐Yu; Lam, Chak Hin; Lin, Xin-Hui; Hsu, Jung-I; Fan, Syuan-Yun; Gupta, Nitesh K.; Lin, Yu‐Chun; Tee, Boon Khoon; Li, Jui-Ping; Chen, Jen‐Kun; Tan, Kui‐Thong

doi:10.1002/asia.202100060

Cited by 5 publications

(4 citation statements)

References 49 publications

(17 reference statements)

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“…To achieve efficient labeling of the cell surface hCA proteins, the probes were functionalized with the difluorophenyl ester reactive group, which was reported previously by our group (Figure S2). , The detailed synthetic schemes to prepare these labeling probes are described in the Supporting Information (Schemes S1–S5). By using MALDI-TOF and in-gel Western blot analysis, we confirmed that 1 can be labeled efficiently to the purified hCAI and transmembrane hCAIX from MCF7 cells, respectively (Figure S3).…”

Section: Resultsmentioning

confidence: 99%

Small-Molecule Modulated Affinity-Tunable Semisynthetic Protein Switches

Shao-jie

et al. 2022

ACS Sens.

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Engineered protein switches have been widely applied in cell-based protein sensors and point-of-care diagnosis for the rapid and simple analysis of a wide variety of proteins, metabolites, nucleic acids, and enzymatic activities. Currently, these protein switches are based on two main types of switching mechanisms to transduce the target binding event to a quantitative signal, through a change in the optical properties of fluorescent molecules and the activation of enzymatic activities. In this paper, we introduce a new affinity-tunable protein switch strategy in which the binding of a small-molecule target with the protein activates the streptavidin–biotin interaction to generate a readout signal. In the absence of a target, the biotinylated protein switch forms a closed conformation where the biotin is positioned in close proximity to the protein, imposing a large steric hindrance to prevent the effective binding with streptavidin. In the presence of the target molecule, this steric hindrance is removed, thereby exposing the biotin for streptavidin binding to produce strong fluorescent signals. With this modular sensing concept, various sulfonamide, methotrexate, and trimethoprim drugs can be selectively detected on the cell surface of native and genetically engineered cells using different fluorescent dyes and detection techniques.

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Section: Resultsmentioning

confidence: 99%

Small-Molecule Modulated Affinity-Tunable Semisynthetic Protein Switches

Shao-jie

et al. 2022

ACS Sens.

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“…Nucleic acid fluorescent probe–based imaging technology has attracted widespread attention, which can display quantitative maps according to the concentrations of target molecules in living samples. Although numerous nucleic acid probes have sufficient sensitivity and selectivity for in vitro imaging of various targets, there are several scientific and technical challenges to in situ and in vivo fluorescence imaging: i) the complexity of tumor microenvironment (TME) might cause damage to normal cells and non-target molecules ( Lin et al, 2021b ) and ii) the high background of enzymatic catalysis dramatically decreases the feasibility of in vivo fluorescence imaging ( Ferrero et al, 2021 ). Thus, novel fluorophores with high quantum yield need to be explored for eliminating the background signal and reducing the perturbation to normal biological processes, which is vital for monitoring the enzymatic processes with greater temporal and spatial resolution.…”

Section: Discussionmentioning

confidence: 99%

The Application of Nucleic Acid Probe–Based Fluorescent Sensing and Imaging in Cancer Diagnosis and Therapy

Huang

Wang

et al. 2021

Front. Chem.

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It is well known that cancer incidence and death rates have been growing, but the development of cancer theranostics and therapeutics has been a challenging work. Recently, nucleic acid probe–based fluorescent sensing and imaging have achieved remarkable improvements in a variety of cancer management techniques, credited to their high sensitivity, good tolerance to interference, fast detection, and high versatility. Herein, nucleic acid probe–based fluorescent sensing and imaging are labeled with advanced fluorophores, which are essential for fast and sensitive detection of aberrant nucleic acids and other cancer-relevant molecules, consequently performing cancer early diagnosis and targeted treatment. In this review, we introduce the characteristics of nucleic acid probes, summarize the development of nucleic acid probe–based fluorescent sensing and imaging, and prominently elaborate their applications in cancer diagnosis and treatment. In discussion, some challenges and perspectives are elaborated in the field of nucleic acid probe–based fluorescent sensing and imaging.

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“…After the first success using tosylate, several cleavable electrophiles have been developed as reactive modules to date as shown in Figure 2A, including phenyl ester, [27,28] acyl imidazole, [29,30] dibromophenyl benzoate, [31] N-acyl-N-alkyl-sulfonamide (these undergo acyl transfer), [32,33] N-sulfonyl pyridone (protein sulfonation), [34] sulfone azide, (Cu 2 + catalyzed diazotransfer on Lys) [35] Nsulfanylethylanilide (SEAlide) (environment responsive N-to-S acyl transfer followed by acyl transfer on the reactive amino acid), [36] O-nitrobenzoxadiazole (O-NBD: SN aromatic reaction), [37][38][39] TCC probe (Nucleophilic addition to alkyne), [40] and others (Figure 2A and Figure S1). [41][42][43][44] It is noted that each electrophile showed distinct amino acids selectivity in the labeling reaction and varied labeling kinetics. These progresses are valuable to expand the availability/flexibility of LDchem for targeting diverse POIs in a variety of research objectives.…”

Section: Principle Of Ligand-directed Chemistry For Protein Labelingmentioning

confidence: 99%

Ligand‐Directed Chemistry for Protein Labeling for Affinity‐Based Protein Analysis

Sakamoto

Hamachi

2023

Israel Journal of Chemistry

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Structural and functional analyses of proteins-ofinterest (POI) in multimolecular crowding conditions (mMCC) such as live cells and tissues are regarded as inevitable challenges for in depth understanding of the real shapes of POIs in their existing natural environments. Activity-based protein profiling (ABPP) is a definitely powerful tool capable of analyzing a proteome possessing a particular activity under mMCC. While ABPP usually targets a proteome of interest, study of a particular protein in mMCC is also valuable. Although activity-based probes (ABPs) are often used for this aim, most of conventional ABPs cause the loss of original activities, and therefore are not perfectly suitable for functional analysis of labeled proteins. Liganddirected chemistry (LDchem) developed by our group is an alternative approach of ABPs, that can modify a surface of POI rather than its active site using a cleavable electrophile in a traceless manner. LDchem thus enables the POI labeling with a synthetic fluorophore with no or minimal effects on the original functions of POIs even in mMCC. In this review, we briefly describe a principle of LDchem for native protein labeling and summarize its recent chemical biology applications such as the imaging-based biological analysis of POI functions and construction of POI-based biosensors.

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Improved Stabilities of Labeling Probes for the Selective Modification of Endogenous Proteins in Living Cells and In Vivo

Cited by 5 publications

References 49 publications

Small-Molecule Modulated Affinity-Tunable Semisynthetic Protein Switches

Small-Molecule Modulated Affinity-Tunable Semisynthetic Protein Switches

The Application of Nucleic Acid Probe–Based Fluorescent Sensing and Imaging in Cancer Diagnosis and Therapy

Ligand‐Directed Chemistry for Protein Labeling for Affinity‐Based Protein Analysis

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