Immunoproteomic analysis and identification of possible allergenic proteins in Artemisia annua pollen

Mei Wang; Jingui Ma; Zhigang Yang; Weibiao Wang; Yuping Sa; Fen Ma; Weiman Zhang; Xinmin He; Guoning Chen; Xueqin Ma

doi:10.1016/j.intimp.2024.113837

Immunoproteomic analysis and identification of possible allergenic proteins in Artemisia annua pollen

Int Immunopharmacol. 2024 Dec 16:146:113837. doi: 10.1016/j.intimp.2024.113837. Online ahead of print.

Authors

Mei Wang¹, Jingui Ma¹, Zhigang Yang¹, Weibiao Wang¹, Yuping Sa¹, Fen Ma¹, Weiman Zhang¹, Xinmin He¹, Guoning Chen², Xueqin Ma³

Affiliations

¹ School of Pharmacy, Ningxia Medical University, 1160 Shenli Street, Yinchuan 750001, China.
² School of Pharmacy, Ningxia Medical University, 1160 Shenli Street, Yinchuan 750001, China. Electronic address: chengn1992@stu.xjtu.edu.cn.
³ School of Pharmacy, Ningxia Medical University, 1160 Shenli Street, Yinchuan 750001, China. Electronic address: maxueqin217@126.com.

PMID: 39689605
DOI: 10.1016/j.intimp.2024.113837

Abstract

Background: Artemisia annua (A. annua) is a wind-pollinated weed and a major allergen responsible for allergic respiratory diseases in Northern China.

Methods: This study involved the separation of pollen proteins from A. annua utilizing SDS-PAGE and Coomassie Blue staining techniques. The effectiveness of extracting allergens from Artemisia annua pollen (AAP) were confirmed both in vivo and in vitro. A mouse model was established using A. annua pollen (AAP) extracts. In vitro, the interaction between allergenic proteins in the AAP extract and specific IgE antibodies present in patients' sera was analyzed, using IgE-immunoblotting and ELISA methods. Protein bands were subsequently analyzed by mass spectrometry. The recombinant Art an 3 (rArt an 3) protein was obtained via recombination, expression, and purification. Finally, the binding activity of rArt an 3 specific IgE was subsequently examined using Western blot and inhibition ELISA (iELISA).

Results: The electrophoretic profiles of AAP showed band patterns ranging from 10 to 70 kDa, with the most prominet IgE-binding pollen proteins detected at approximately 12 kDa. After stimulation of AAP, the sensitized group of mice exhibited significant allergic symptoms compared to the control group. Mass spectrometry analysis identified 7 proteins, including putative aldehyde oxidase Art an 7, Art v 1-like protein, Art an 3.0101 allergen precursor, Art an 3.0102 allergen precursor, Art an 3.0201 allergen precursor, lipid transfer protein 3, and non-specific lipid-transfer protein. The results of immunoblotting and iELISA showed that rArt an 3 could bind to IgE antibodies in the patient sera, and co-incubation of rArt an 3 with serum, the binding serum significantly inhibited IgE binding to the crude AAP.

Conclusions: The Art an 3 is a key allergenic protein in AAP with a high IgE sensitization rate in the studied population sample. These findings enhance our understanding of the sensitization components of AAP and its sensitization characteristics within the Chinese population, thereby promoting the development of precise molecular diagnostics and therapeutic interventions.

Keywords: Allergen; Artemisia annua; Immunoproteomics; Mass spectrometry; Recombinant protein.