Simultaneous degradation of two mycotoxins enabled by a fusion enzyme in food-grade recombinant Kluyveromyces lactis

Abstract

Aflatoxin B₁ (AFB₁) and zearalenone (ZEN) are two predominant mycotoxins ubiquitously found in corn, peanuts, and other grains, which pose a great threat to human health. Therefore, safe and effective methods for detoxification of these mycotoxins are urgently needed. To achieve simultaneous degradation of multiple mycotoxins, a fusion enzyme ZPF1 was constructed by linking zearalenone hydrolase and manganese peroxidase with a linker peptide GGGGS. This fusion enzyme was secretory expressed successfully in the newly constructed food-grade recombinant strain Kluyveromyces lactis GG799(pKLAC1-ZPF1), and was investigated with the mycotoxins degradation efficiency in two reaction systems. Results showed that both AFB₁ and ZEN can be degraded by ZPF1 in reaction system 1 (70.0 mmol/L malonic buffer with 1.0 mmol/L MnSO₄, 0.1 mmol/L H₂O₂, 5.0 µg/mL AFB₁ and ZEN, respectively) with the ratios of 46.46% and 38.76%, respectively. In reaction system 2 (50.0 mmol/L Tris–HCl, with 5.0 µg/mL AFB₁ and ZEN, respectively), AFB₁ cannot be degraded while ZEN can be degraded with the ratio of 35.38%. To improve the degradation efficiency of these mycotoxins, optimization of the induction and degradation conditions were fulfilled subsequently. The degradation ratios of AFB₁ and ZEN by ZPF1 in reaction system 1 reached 64.11% ± 2.93% and 46.21% ± 3.17%, respectively. While in reaction system 2, ZEN was degraded by ZPF1 at a ratio of 41.45% ± 3.34%. The increases of degradation ratios for AFB₁ and ZEN in reaction system 1 were 17.65% and 7.45%, respectively, while that for ZEN in reaction system 2 was 6.07%, compared with the unoptimized results.

Keywords

Mycotoxins / Aflatoxin B₁ / Zearalenone / Fusion enzyme / Degradation / Kluyveromyces lactis / Food-grade

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Funding

Jiangsu Agriculture Science and Technology Innovation Fund (JASTIF)(CX(19)3109)

Key Research and Development Program of Jiangsu Province(BE2018306)

Jiangsu Overseas Research & Training Program for University Prominent Young & Middle-aged Teachers and Presidents