Genome mining reveals the genes of carboxypeptidase for OTA-detoxification in Bacillus subtilis CW14

Bacillus subtilis CW14, isolated from fresh elk droppings in Beijing Zoo, is a Gram-positive, conferred Generally Recognized as Safe (GRAS) bacterium with the capacity of ochratoxin A (OTA) detoxification. The genome sequence of the CW14 strain showed a size of 4,287,522 bp with 44.06% GC content. It was predicted many putative enzymes involved in degrading mycotoxin by analyzing the signal peptides and the transmembrane regions. Nine extracellular enzymes were predicted relating to OTA detoxification, including four D-Ala-D-Ala carboxypeptidases, two hydrolases, two amidases, and one lactamase. Indeed, two of the carboxypeptidase genes dacA and dacB, expressed in Escherichia coli, were verified contributing to OTA detoxification. DacA and OTA were mixed incubated for 24 h, and the degradation rate reached 71.3%. After purification, the concentration of recombinant DacA protein was 0.5 mg/mL. Bacillus subtilis CW14 and its carboxypeptidases may be used as OTA detoxification agents in food and feed industry production.