Identification and in silico prediction of metabolites of the model compound, tebufenozide by human CYP3A4 and CYP2C19.

The metabolites of tebufenozide, a model compound, formed by the yeast-expressed human CYP3A4 and CYP2C19 were identified to clarify the substrate recognition mechanism of the human cytochrome P450 (CYP) isozymes. We then determined whether tebufenozide metabolites may be predicted in silico. Hydrogen abstraction energies were calculated with the density functional theory method B3LYP/6-31G(∗). A docking simulation was performed using FRED software. Several alkyl sites of tebufenozide were hydroxylated by CYP3A4 whereas only one site was modified by CYP2C19. The accessibility of each site of tebufenozide to the reaction center of CYP enzymes and the susceptibility of each hydrogen atom for metabolism by CYP enzymes were evaluated by a docking simulation and hydrogen abstraction energy estimation, respectively.

Proteolytic activity of selected commercial Lactobacillus helveticus strains on soy protein isolates.

Soy protein isolates were fermented by three commercial Lactobacillus helveticus strains for a maximum of seven days to investigate the resulting proteolysis. The proteolytic activity of the most active strain (LH88) was further analyzed (LC-MS/MS and GC-MS) and it was shown that the ß-conglycinin α subunit 1, ß-conglycinin α' subunit, glycinin G1, and 2S albumin were specifically degraded. Peptigram analysis and visualization of the crystal structure showed that the hydrolysis sites of ß-conglycinin α subunit, α' subunit, and the glycinin G1 were located on the surface of the molecule or at the mobile disordered region, hence being highly accessible for the proteinase of LH88. The proteins were partially further degraded to free amino acids, and subsequently catabolized to volatile compounds. However, most of the proteins remained native, even after seven days of fermentation, thus additional modification of protein structure or adjustment of fermentation conditions are required for effective generation of flavor compounds.