A photocleavable peptide-tagged mass probe for chemical mapping of epidermal growth factor receptor 2 (HER2) in human cancer cells

Liu, Liang; Kuang, Yuqiong; Wang, Zhongcheng; Chen, Yun

doi:10.1039/d0sc04481d

Cited by 12 publications

(9 citation statements)

References 40 publications

(37 reference statements)

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“…Among them, well-designed peptides can have both sequence uniqueness to avoid overlapping with endogenous peptides and sufficient mass spectrometric response for quantification . Notably, several peptide mass tags have been frequently employed in our previous work. − In this study, the peptide AVLGDPFR was selected as the mass tag. The product ion spectrum and LC–MS/MS chromatogram of AVLGDPFR are exhibited in Figure S5.…”

Section: Resultsmentioning

confidence: 99%

Construction of a Mass-Tagged Oligo Probe Set for Revealing Protein Ratiometric Relationship Associated with EGFR–HER2 Heterodimerization in Living Cells

Sun

Zhu

et al. 2022

Anal. Chem.

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Protein dimerization, as the most common form of protein−protein interaction, can manifest more significant roles in cellular signaling than individual monomers. For example, excessive formation of EGFR−HER2 dimer has been implicated in cancer development and therapeutic resistance in addition to the overexpression of EGFR and HER2 proteins. Thus, quantitative evaluation of these heterodimers in living cells and revelation of their ratiometric relationship with protein monomers in dimerization may provide insights into clinical cancer management. To achieve this goal, the prerequisite is protein heterodimer quantification. Given the current lack of quantitative methods, we constructed a mass-tagged oligo nanoprobe set for quantification of EGFR−HER2 dimer in living cells. The mass-tagged oligo nanoprobe set contained two targeting probes (nucleic acid aptamers), a connector probe, a hairpin probe, and a photocleavable mass-tagged probe. Two distinct aptamers can recognize target protein monomers and initiate the subsequent hybridization cascade involving binding to the connector probe, formation of an initiator strand, opening of a hairpin probe, and ensuing hybridization with a photocleavable mass-tagged probe. Ultimately, the mass tag was released under ultraviolet light and then subjected to mass spectrometric analysis. In this way, the information regarding the interaction between two protein monomers was successfully converted to the quantitative signal of the mass tag. Using the assay, the expression level of EGFR−HER2 dimer and its relationship with individual protein monomers were determined in four breast cancer cell lines. We are among the first to obtain the absolute level of protein heterodimer, and this quantitative information may be vital in understanding the molecular basis of cancer.

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Section: Resultsmentioning

confidence: 99%

Construction of a Mass-Tagged Oligo Probe Set for Revealing Protein Ratiometric Relationship Associated with EGFR–HER2 Heterodimerization in Living Cells

Sun

Zhu

et al. 2022

Anal. Chem.

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“… 29 In this study, an o -nitrobenzyl derivative PL was employed as the linker, which is UV light sensitive. 30 The photo-cleavable mechanism of the PL is illustrated in Fig. S5.…”

Section: Resultsmentioning

confidence: 99%

Ultrasensitive detection of β-lactamase-associated drug-resistant bacteria using a novel mass-tagged probe-mediated cascaded signal amplification strategy

et al. 2022

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“…To further understand the role of HER2 homodimer in HER2-positive breast cancer, the ratiometric relationship of the homodimer with the total protein was studied in three HER2-positive cell lines (SK-BR-3 cells, BT474 cells, and HCC1954 cells). Using the previously developed HER2 quantitative assay, 46 the total amount of HER2 was determined and is presented in Figure 6E. The percent HER2 homodimer out of the HER2 monomer is shown in Figure 6F.…”

Section: Optimization and Validation Of The Probe Set-based Approachmentioning

confidence: 99%

Steric Hindrance On–Off Mass-Tagged Probe Set Enables Detection of Protein Homodimer in Living Cells

Zhu

Shi

et al. 2022

ACS Appl. Mater. Interfaces

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The major challenge in the detection of protein homodimers is that the identical monomers in a homodimer are indistinguishable using most conventional methods and cannot be sequentially recognized. In this study, a steric hindrance on−off mass-tagged probe set strategy was developed for the quantification of HER2 homodimer in living cells. The probe set contained a hindrance probe and a detection probe. The hindrance probe had a DNA dendrimer as a hindrance group to achieve the steric hindrance on−off function and thus the assignment of monomer identity. The detection probe contained a mass tag released for mass spectrometric quantification. Using the steric hindrance on−off mass-tagged probe set, the level of HER2 homodimer in various breast cancer cell lines was quantified. This is the first report to determine the quantity of protein homodimers, and the steric hindrance on−off probe set developed herein can facilitate the illustration of protein function in cancer.

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A photocleavable peptide-tagged mass probe for chemical mapping of epidermal growth factor receptor 2 (HER2) in human cancer cells

Abstract: An integrated approach based on a photocleavable peptide tagged mass probe provides chemical mapping including quantitative and visual information of HER2.

Cited by 12 publications

References 40 publications

Construction of a Mass-Tagged Oligo Probe Set for Revealing Protein Ratiometric Relationship Associated with EGFR–HER2 Heterodimerization in Living Cells

Construction of a Mass-Tagged Oligo Probe Set for Revealing Protein Ratiometric Relationship Associated with EGFR–HER2 Heterodimerization in Living Cells

Ultrasensitive detection of β-lactamase-associated drug-resistant bacteria using a novel mass-tagged probe-mediated cascaded signal amplification strategy

Steric Hindrance On–Off Mass-Tagged Probe Set Enables Detection of Protein Homodimer in Living Cells

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