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ARS Home » Pacific West Area » Albany, California » Western Regional Research Center » Healthy Processed Foods Research » Research » Publications at this Location » Publication #398734
Research Project: Prevention of Obesity Related Metabolic Diseases by Bioactive Components of Food Processing Waste Byproducts and Mitigation of Food Allergies

Location: Healthy Processed Foods Research

Title: Expression, purification, characterization, and patient IgE reactivity of new macadamia nut iso-allergen

Author
item Zhang, Yuzhu
item Bhardwaj, Shilpa
item LYU, SHU-CHEN - Stanford School Of Medicine
item CHINTHRAJAH, SHARON - Stanford School Of Medicine
item NADEAU, KARI - Stanford School Of Medicine
item LI, CAIMING - Jiangnan University

Submitted to: Protein Expression and Purification
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 11/28/2022
Publication Date: 11/30/2022
Citation: Zhang, Y., Bhardwaj, S.R., Lyu, S., Chinthrajah, S., Nadeau, K., Li, C. 2022. Expression, purification, characterization, and patient IgE reactivity of new macadamia nut iso-allergen. Protein Expression and Purification. 203. Article 106211. https://doi.org/10.1016/j.pep.2022.106211.
DOI: https://doi.org/10.1016/j.pep.2022.106211

Interpretive Summary: The commercial production of macadamia nut, considered a healthy food, is mainly in the United States. Macadamia nut is also known to be a very potent allergenic food for more than ten years. However, little information is available about macadamia nut allergens. Macadamia nut legumin was recently identified as a food allergen named Mac i 2. Purified allergens are required to study their properties and better understand their allergenicity. Recombinant production for food allergens is advantageous as the purification of allergens from foods is often complicated due to the level of their presence or the existence of multiple isoforms. Thus, the protocols for recombinant production of many allergens have been reported. However, legumins are hexameric proteins that require post-translational modification. The recombinant production of an allergen in this family with native structures has not been accomplished. This study reported the production of a macadamia nut protein based on its mRNA sequence available in the databases. Its expression in bacteria was successful and was confirmed to be a novel isoform of macadamia nut allergen Mac i 2. The information can be used to obtain valuable reagents for studying the properties of the food allergens in the legumin family and developing methods for allergen mitigation.

Technical Abstract: Structural and functional information about food allergens is essential for understanding the allergenicity of food proteins. All allergens belong to a small number of protein families. Various allergens from different families have been successfully produced recombinantly in E. coli for their characterization and applications in allergy diagnosis and treatment. However, recombinant hexameric 11S seed storage protein has not been reported, although numerous 11S legumins are known to be food allergens, including the recently identified macadamia nut allergen Mac i 2. Here we report the production of a macadamia nut legumin by expressing it in E. coli with a substrate site of HRV 3C protease and cleaving the purified protein with HRV 3C protease. The protease divided the protein into two chains and left a native terminus for the C-terminal chain, resulting in a recombinant hexameric 11S allergen for the first time after the residues upstream to the cleavage site flipped out of the way of the trimer-trimer interaction. The 11S allergens are known to have multiple isoforms in many species. The present study removed an obstacle in obtaining homogeneous allergens needed for studying allergens and mitigating allergenicity. Immunoreactivity of the protein with serum IgE confirmed it to be a new isoform of Mac i 2.