AUTHOR=Yang Peizhou , Xiao Wei , Lu Shuhua , Jiang Shuying , Jiang Suwei , Chen Jianchao , Wu Wenjing , Zheng Zhi , Jiang Shaotong 

TITLE=Characterization of a Trametes versicolor aflatoxin B1-degrading enzyme (TV-AFB1D) and its application in the AFB1 degradation of contaminated rice in situ

JOURNAL=Frontiers in Microbiology

VOLUME=13

YEAR=2022

URL=https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2022.960882

DOI=10.3389/fmicb.2022.960882

ISSN=1664-302X

ABSTRACT=<p>Aflatoxin B<sub>1</sub> (AFB<sub>1</sub>) contaminates rice during harvest or storage and causes a considerable risk to human and animal health. In this study, <italic>Trametes versicolor</italic> AFB<sub>1</sub>–degrading enzyme (TV–AFB<sub>1</sub>D) gene recombinantly expressed in engineered <italic>E. coli</italic> BL21 (DE3) and <italic>Saccharomyces cerevisiae</italic>. The TV–AFB<sub>1</sub>D enzymatic characteristics and AFB<sub>1</sub> degradation efficiency in contaminated rice were investigated. Results showed that the size of recombinant TV-AFB<sub>1</sub>D expressing in <italic>E. coli</italic> BL21 (DE3) and <italic>S. cerevisiae</italic> was appropriately 77 KDa. The kinetic equation of TV-AFB<sub>1</sub>D was <italic>y</italic> = 0.01671<italic>x</italic> + 1.80756 (<italic>R</italic><sup>2</sup> = 0.994, <italic>K<sub>m</sub></italic> = 9.24 mM, and <italic>V<sub>max</sub></italic> = 553.23 mM/min). The <italic>Kcat</italic> and <italic>Kcat</italic>/<italic>K<sub>m</sub></italic> values of TV-AFB<sub>1</sub>D were 0.07392 (s<sup>−1</sup>) and 8 M<sup>−1</sup> s<sup>−1</sup>, respectively. The AFB<sub>1</sub> concentration of contaminated rice decreased from 100 μg/ml to 32.6 μg/ml after treatment at 32°C for 5 h under the catabolism of TV-AFB<sub>1</sub>D. <italic>S. cerevisiae</italic> engineered strains carrying aldehyde oxidase 1 (<italic>AOX1</italic>) and <italic>Cauliflower mosaic virus</italic> 35 S (<italic>CaMV</italic> 35 S) promoters caused the residual AFB<sub>1</sub> contents, respectively, decreased to 3.4 and 2.9 μg/g from the initial AFB<sub>1</sub> content of 7.4 μg/g after 24 h of fermentation using AFB<sub>1</sub>-contaminated rice as substrate. The AFB<sub>1</sub> degradation rates of <italic>S. cerevisiae</italic> engineered strains carrying <italic>AOX1</italic> and <italic>CaMV</italic> promoters were 54 and 61%, respectively. Engineered <italic>S. cerevisiae</italic> strains integrated with TV-AFB<sub>1</sub>D expression cassettes were developed to simultaneously degrade AFB<sub>1</sub> and produce ethanol using AFB<sub>1</sub>-contaminated rice as substrate. Thus, TV-AFB<sub>1</sub>D has significant application potential in the AFB<sub>1</sub> decomposition from contaminated agricultural products.</p>