Deoxynivalenol (DON), a mycotoxin with global distribution, has caused serious economic losses and poses food and feed safety issues. Recently, a quinone-dependent dehydrogenase (DADH) was identified to degrade DON into the less toxic 3-keto-DON by oxidation of the critical toxic hydroxyl group, which was then reduced by aldo-keto reductase AKR13B3 into the relatively non-toxic 3-epi-DON. Nevertheless, the poor thermostability of DADH limits its practical application. Here, a combinatorial mutant M3 with significantly improved thermostability was screened using computation-aided design combining ancestral sequence reconstruction and structural analysis. The mutants M3 exhibited high thermostability with an 8.93-fold longer half-life at 55℃ than that of wild-type. Moreover, structural analyses and molecular dynamics simulations revealed that the reduced flexibility, enhanced structural rigidity, favorable electrostatic potential, and increased number of hydrogen bonds in mutant M3 were the primary factors underlying its improved thermostability. This work provides a facile and efficient strategy to improving the thermostability of DON detoxification enzymes for agricultural and food industry applications.
Ma et al. (Sun,) studied this question.