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MrgX2-SNAP-tag/cell membrane chromatography model coupled with liquid chromatography-mass spectrometry for anti-pseudo-allergic compound screening in Arnebiae Radix

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Abstract

Pseudo-allergic reactions (PARs) are IgE-independent hypersensitivity reactions. Mas-related G protein-coupled receptor-X2 (MrgX2) was proved the key receptor of PAR. The anti-pseudo-allergic compound discovery based on MrgX2 was of great value. Cell membrane chromatography (CMC) based on MrgX2 provides a convenient and effective tool in anti-pseudo-allergic compound screening and discovery, and further improvements of this method are still needed. In this work, SNAP-tag was introduced at C-terminal of Mas-related G protein-coupled receptor (MrgX2-SNAP-tag), and an MrgX2-SNAP-tag/CMC model was then conducted using CMC technique. Comparative experiments showed that the new model not only satisfied the good selectivity and specificity of screening but also exhibited more stable and longer life span than traditional MrgX2/CMC model. By coupling with HPLC–MS, two compounds were screened out from Arnebiae Radix and identified as shikonin and acetylshikonin. Nonlinear chromatography was performed to study the interactions between two screened compounds and MrgX2, and binding constant (KA) of shikonin and acetylshikonin with MrgX2 were 2075.67 ± 0.34 M−1 and 32201.36 ± 0.35 M−1, respectively. Furthermore, β-hexosaminidase and histamine release assay in vitro demonstrated that shikonin (1–5 μM) and acetylshikonin (2.5–10 μM) could both antagonize C48/80-induced allergic reaction. In conclusion, the MrgX2-SNAP-tag/CMC could be a reliable model for screening pseudo-allergy-related components from complex systems.

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Funding

This work was supported by National Natural Science Foundation of China (Grant Numbers: 82104118, 81930096, and 81973278).

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Authors and Affiliations

  1. School of Pharmacy, Xi’an Jiaotong University, 76# Yanta West Road, Xi’an, 710061, China

    Qianqian Jia, Jia Fu, Chunlei Gao, Hong Wang, Saisai Wang, Peida Liang, Shengli Han, Yanni Lv & Langchong He

  2. Institute of Pharmaceutical Science and Technology, Western China Science & Technology Innovation Harbour, Xi’an, 710115, China

    Qianqian Jia, Jia Fu, Chunlei Gao, Hong Wang, Saisai Wang, Peida Liang, Shengli Han, Yanni Lv & Langchong He

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  1. Qianqian Jia
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Contributions

QJ: conceptualization, methodology, writing—original draft. JF: methodology, validation. CG and HW: data curation, investigation. SW and PL: data curation. SH: writing—reviewing. YL and LH: writing—reviewing and editing.

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Correspondence to Yanni Lv or Langchong He.

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Jia, Q., Fu, J., Gao, C. et al. MrgX2-SNAP-tag/cell membrane chromatography model coupled with liquid chromatography-mass spectrometry for anti-pseudo-allergic compound screening in Arnebiae Radix. Anal Bioanal Chem 414, 5741–5753 (2022). https://doi.org/10.1007/s00216-022-04167-4

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