Rational Screening of Four Peroxidases with High Aflatoxin B1 Degradation Efficiency via Integrated Computational Simulations

Aflatoxin B1 (AFB1) degradation remains a critical challenge due to limitations of conventional methods. This study proposed a computational screening strategy to screen the AFB1-degrading enzymes, and four peroxidases derived from <i>Agrocybe pediades</i> (ApeLip), <i>Klebsiella pneumoniae</i> (KpDyp), <i>Trametopsis cervina</i> (TrcLip), and <i>Pleurotus eryngii</i> (VPL2) were selected. The enzymes heterologously expressed and purified in <i>Escherichia coli</i> Shuffle T7-B possessed the ability to efficiently degrade AFB1 and a certain degree of alkaline tolerance. The enzymes ApeLip, KpDyp, TrcLip, and VPL2 degraded AFB1 by 90.06, 91.55, 91.93, and 92.91%, respectively, under optimal conditions. Molecular dynamics simulations indicated that van der Waals interactions are the main binding mechanism, especially acting through hydrophobic residues. UPLC-QTOF-MS analysis confirmed the hydroxylation of the C₈═C₉ bond of AFB1 to produce AFB1-8,9-dihydrodiol with lower toxicity. The combined computer simulation and practical framework establishes an effective platform for rational enzyme discovery and effectively promotes the development of mycotoxin mitigation strategies.