Changes of shrimp myofibrillar proteins hydrolyzed by Virgibacillus proteases: Structural characterization, mechanism visualization, and flavor compound formation.

To explore the mechanism of Virgibacillus proteases on hydrolysis of shrimp myofibrillar protein (SMP) and formation of volatile compounds, the fermented broth of Virgibacillus halodenitrificans was purified and the protease was identified as peptidase S8. The enzyme had optimum activity at pH 7.0-8.5 and 40-50 °C, and showed good stability at pH 6.5-8.5 and 20-50 °C. The enzyme showed certain salt and metal ion tolerance. Inhibitor results indicated that the enzyme might belong to the serine protease family. V. halodenitrificans proteases (BP) had a stronger ability to degrade SMP compared to Bacillus subtilis proteases (BS). After 60 min of hydrolysis, the hydrolysis index and surface hydrophobicity value of the BP sample were 36.7 % and 177.5 higher than those of the BS sample, respectively. Various spectral measurement results showed that the structural conformation of the BP-treated SMP was significantly changed, with a smaller particle size (510.4 nm) and a lower zeta potential (-27.7 mV). Molecular docking results showed that the enzyme had the highest degradation capacity for myofibrillar heavy chains, followed by actin, and the lowest for myofibrillar light chains, with the interaction forces being hydrogen bonding and hydrophobic interactions. In addition, BP-treated SMP had higher levels of peptides, small molecular weight peptides (<1 kDa), and umami amino acids compared to the BS sample. Solid-phase microextraction-gas chromatography-mass spectrometry (SPME-GC-MS) results showed that BP hydrolysates contained more volatile compounds and key volatile compounds than BS hydrolysates. Pyrazines and alcohols were the main volatile flavor compounds in BS and BP hydrolysates, respectively.