抄録
Bitterness-masking agents are compounds that reduce the unfavorable bitter taste of food ingredients or drugs. Here we propose a novel screening method employing peptide array analysis for their identification bitterness-masking agents. Daily intake of green tea is considered beneficial because of the health benefits of catechin, an important component. Epigallocatechin gallate (EGCG), a major green tea polyphenol, has a broad range of physiologically beneficial functions, but its bitterness is unappealing to humans. In the present study, EGCG-binding peptides, which are promising bitterness-masking agents of EGCG, were screened using an amino acid sequence of ribulose 1,5-bisphosphate carboxylase/oxygenase (RuBisCO), a protein obtained from the industrial waste of green tea production. Five hundred and ninety-eight types of pentadecapeptide contained in the RuBisCO sequence were chemically synthesized in parallel using SPOT technology. Seven peptides in the peptide library were characterized as EGCG-binding using redox-cycle staining. Deletion analysis of the amino acid residues from the N- or C-termini of these peptides highlighted the central regions (from 2 to 15 amino acids in length) as contributing to this binding. Subsequent alanine scanning revealed a critical contribution of Lys or Arg residues for all peptides obtained in the present study. Moreover, roles of these basic amino acid residues in the binding to EGCG were clearly indicated by analysis using a dipeptide library comprising 361 types of dipeptide. The EGCG-binding peptides MHFRVLAKALR and FTGLKSTSAFPVTRK identified using peptide array analysis were evaluated for their potential bitterness-masking effects using a cell-based assay employing hTAS2R39, a human bitter taste receptor for EGCG. The EGCG-binding peptides suppressed activation of hTAS2R39, indicating a bitterness-masking effect. These results revealed that bitterness-masking effects can be predicted for bitter substance-binding peptides using peptide array analysis.
