Identificationand Characterization of Two MultimycotoxinDegrading Enzymes from Klebsiella pneumoniae HNGD-HS06

Co-contamination of foods with multiple mycotoxins highlights the need for broad-spectrum detoxification enzymes. In this study, Klebsiella pneumoniae HNGD-HS06 was isolated from contaminated peanuts and degraded aflatoxin B1 (AFB1), zearalenone (ZEN), and alternariol (AOH) by 72.52%, 63.42%, and 76.17%, respectively. Genome-guided screening and heterologous expression identified two candidate proteins, HSFtsp and HSCueO. HSCueO shared only 28.08% sequence identity with CotA laccase, while HSFtsp has not previously been associated with mycotoxin degradation. HSFtsp degraded AFB1, ZEN, and AOH by up to 99.16%, 95.12%, and 94.72%, respectively, whereas HSCueO achieved 98.97%, 99.23%, and 99.74% degradation. Product analysis and zebrafish assays suggested that both enzymes transformed these toxins into less toxic products, while molecular docking supported potential enzyme-substrate interactions. Both enzymes retained detoxification activity in contaminated peanut, corn, and wheat flour, expanding enzyme resources for multimycotoxin control in foods.