Crosslinked Recombinant-Ara h 1 Catalyzed by Microbial Transglutaminase: Preparation, Structural Characterization and Allergic Assessment

Abstract

As the one of the major allergens in peanut, the allergenicity of Ara h 1 is influenced by its intrinsic structure, which can be modified by different processing. However, molecular information in this modification has not been clarified to date. Here, we detected the influence of microbial transglutaminase (MTG) catalyzed cross-linking on the recombinant peanut protein Ara h 1 (rAra h 1). Electrophoresis and spectroscopic methods were used to analysis the structural changes. The immunoreactivity alterations were characterized by enzyme linked immunosorbent assay (ELISA), immunoblotting and degranulation test. Structural features of cross-linked rAra h 1 varied at different reaction stages. Hydrogen bonds and disulfide bonds were the main molecular forces in polymers induced by heating and reducing. In MTG-catalyzed cross-linking, ε-(γ-glutamyl) lysine isopeptide bonds were formed, thus inducing a relatively stable structure in polymers. MTG catalyzed cross-linking could modestly but significantly reduce the immunoreactivity of rAra h 1. Decreased content of conserved secondary structures led to a loss of protection of linear epitopes. Besides, the reduced surface hydrophobic index and increased steric hindrance of rAra h 1 made it more difficult to bind with antibodies, thus hindering the subsequent allergic reaction.

Identification of the Immunoglobulin E Epitope of Arginine Kinase, an Important Allergen from Crassostrea angulata

“…The structure of allergen is closely related to its immunoreactivity. The allergenicity of Ara h 1 is affected by its tertiary structure . It has been reported that the integrity structure of crab AK is essential for its immunobinding activity .…”

“…The allergenicity of Ara h 1 is affected by its tertiary structure. 10 It has been reported that the integrity structure of crab AK is essential for its immunobinding activity. 15 The tertiary structure of an allergen is usually maintained by chemical bonds, which mainly include hydrogen bonds, hydrophobic bonds, disulfide bonds, etc.…”

“…The structure of many allergens is related to their immunoreactivity. − At present, chemical denaturants can be used to investigate the relationships between the structure and immunoreactivity of the allergen . As a denaturing agent, sodium dodecyl sulfate (SDS), can destroy hydrogen bonds and hydrophobicity of proteins, thus destroying the secondary and tertiary structures of protein molecules .…”

Analysis of Immunoreactivity of α/α2-Tropomyosin from Haliotis discus hannai, Based on IgE Epitopes and Structural Characteristics

“…Meanwhile, the secondary structure regions were predicted by Discovery Studio software. The analysis of chemical bonds was performed by Wang et al 30 The contents of ionic bond, hydrogen bond, hydrophobic force, and disulfide bond were measured. The surface hydrophobicity of α/ α 2 -TM was analyzed using ANS as a probe, as described by Han et al 8 Statistical Analysis.…”

Peanut protein – an underutilised by‐product with great potential: a review

“…Microbial transglutaminase formed a compact structure and reduced surface hydrophobic index and increased steric hindrance of rAra h 1 (Hu et al ., 2019). As a result, the formed complex did not bind with antibodies, and consequently had reduced allergic reaction (Tian et al ., 2020).…”