Soy protein isolate -(-)-epigallocatechin gallate conjugate: Covalent binding sites identification and IgE binding ability evaluation

•The SPI-EGCG conjugate was prepared under alkaline conditions.•The specific amino acids in SPI serving as covalent binding sites were identified.•There are reactive sites on both phenolic rings of the EGCG molecule.•The composition and sequence of amino acid affect the binding site of EGCG in SPI.•The reaction activity of amino acid residues affects the binding site in SPI. The conjugate prepared from (-)-epigallocatechin gallate (EGCG) and soy protein isolate (SPI) under alkaline and aerobic conditions was analyzed using a Nano-LC-Q-Orbitrap-MS/MS technique. The sulfhydryl and free amino groups of SPI were involved in covalent binding. Fifty-one peptides were conjugated with EGCG. Fifty-nine modified sites were identified, located on Cys, His, Arg, and Lys, respectively. It is the first time to confirm that each of the two phenolic rings of EGCG contained a reactive site that bound to an amino acid residue. The amino acid residue reactivity, amino acid sequence and composition affected the EGCG binding site in SPI. Lys and Arg residues are the most likely sites for modification, and modification appears to reduce IgE binding. This study is helpful to elucidate the pattern of covalent binding of polyphenols to proteins in food systems and provides a theoretical basis for the directional modification of soy proteins with polyphenols.