Activity enhancement of Trametes versicolor aflatoxin B1-degrading enzyme (TV-AFB1D) by molecular docking and site-directed mutagenesis

Aflatoxin B-1(AFB(1)) is highly carcinogenic and teratogenic as a secondary metabolite of Aspergillus flavus and Aspergillus parasiticus. Trametes versicolor aflatoxin B-1-degrading enzyme (TV-AFB(1)D) has capability of AFB(1)degradation. In this study, in order to improve the catalytic performance of TV-AFB(1)D, the molecular docking and site-directed mutagenesis techniques were applied to modify the key amino acid sites. Three mutation sites of Y305A, E436A, and H554A were selected for the directed mutation of TV-AFB(1)D. Four TV-AFB(1)D combinations of mutant H554A, Y305A/H554A, E436A/H554A, Y305A/E436A/H554A had potential to modify the catalytic performance according to the value of de-folding free energy. The TV-AFB(1)D mutations were expressed in engineering E. coli BL21 (DE3) with the size of approximately 77 kDa. The optimal temperature and pH value of TV-AFB(1)D were 32 degrees C and pH 7, respectively. E436A/H554A mutant represented the highest enzyme activities among these four mutants. The specific activity of E436A/H554A mutant (36 U/mg) was 1.84-fold in comparison with wild-type TV-AFB(1)D (19.6 U/mg) under the conditions of pH 7 and 32 degrees C. This study would contribute to the activity increase of TV-AFB(1)D by the performance improvement of AFB(1)degradation by site-directed mutagenesis approach. (C) 2021 Institution of Chemical Engineers. Published by Elsevier B.V. All rights reserved.

Automated summary

In this study, the catalytic performance of TV-AFB(1)D was improved through molecular docking and site-directed mutagenesis techniques. Among the mutants, E436A/H554A showed the highest enzyme activity, indicating a potential for enhancing AFB(1) degradation.