Improved enzymatic accessibility of peanut protein isolate pre-treated using thermosonication

Abstract Thermosonication pre-treatment was used to enhance the pancreatin-induced proteolysis of peanut protein isolate (PPI). Response surface methodology was applied to optimize the thermosonication conditions (including power-output and temperature), and the highest degree of hydrolysis (7.16%) was obtained at 475.0 W, 72 °C. SDS-PAGE analysis showed that at this optimized condition, the enzymatic accessibility of the major constitutive protein arachin in thermosonicated PPI (TS-PPI) was substantially improved compared to that in untreated PPI or sonicated PPI (475 W, 30 °C; S-PPI), resulting in a remarkable increase in protein solubility for the hydrolysates. Protein denaturation and conformation profiles of untreated PPI, S-PPI and TS-PPI were investigated using differential scanning calorimetry, intrinsic fluorescence emission spectroscopy, Fourier transform infra-red spectroscopy and thioflavin-T (ThT) fluorescence assay. It was found that heat could present a markedly additive effect to ultrasound on denaturing peanut proteins, leading to significant changes in protein conformation. TS-PPI was characterized by the appearance of high proportion of parallel intermolecular β -sheets and a strong fluorescence enhancement upon binding to ThT, suggesting that the protein unfolding and aggregation induced by thermosonication probably resulted in the formation of fibril protein aggregates in TS-PPI rather than spherical protein aggregates formed in S-PPI. As a result, the protein conformation of TS-PPI appeared to be more unfolded and flexible than that of untreated PPI or S-PPI, and therefore was more easily accessible to protease. This study shows that thermosonication pre-treatment could be a highly effective and feasible technique to improve the enzymatic accessibility of globular proteins, producing prominent functional benefits for the protein hydrolysates.