Abstract
Aflatoxin B1 (AFB1) is a known toxic human carcinogen and can be detoxified by laccases, which are multicopper oxidases that convert several environmental pollutants and toxins. In this study, a new laccase that could catalyze AFB1 degradation was purified and identified from the white-rot fungus Cerrena unicolor 6884. The laccase was purified using (NH4)2SO4 precipitation and anion exchange chromatography, and then identified as Lac 2 through zymogram and UHPLC-MS/MS based on the Illumina transcriptome analysis of C. unicolor 6884. Six putative laccase protein sequences were obtained via functional annotation. The lac 2 cDNA encoding a full-length protein of 512 amino acids was cloned and sequenced to expand the fungus laccase gene library for AFB1 detoxification. AFB1 degradation by Lac 2 was conducted in vitro at pH 7.0 and 45 °C for 24 h. The half-life of AFB1 degradation catalyzed by Lac 2 was 5.16 h. Acetosyringone (AS), Syrinagaldehyde (SA) and [2,2′ -azino-bis-(3-ethylbenzothiazoline-6-sulfonic acid)] (ABTS) at 1 mM concentration seemed to be similar mediators for strongly enhancing AFB1 degradation by Lac 2. The product of AFB1 degradation catalyzed by Lac 2 was traced and identified to be Aflatoxin Q1 (AFQ1) based on mass spectrometry data. These findings are promising for a possible application of Lac 2 as a new aflatoxin oxidase in degrading AFB1 present in food and feeds.
Highlights
Aflatoxins (AFs), which are commonly produced by Aspergillus fungi such as Aspergillus flavus and Aspergillus parasiticus, are toxoids leading to fluorescence, human carcinogenicity and mutagenicity.The toxicity of AFs is related to the C8 -C9 double bond of the difuran ring and the lactone ring within the coumarin ring [1,2]
Aflatoxin B1 (AFB1) and its metabolites are mainly distributed in corn, peanuts, rice, wheat, oil by-products, human carcinogen, but it can affect a wide range of commodities [4,5]
Our results showed that laccase from C. unicolor was a valuable enzyme in AFB1 degradation
Summary
The toxicity of AFs is related to the C8 -C9 double bond of the difuran ring and the lactone ring within the coumarin ring [1,2] Based on these data, AFs can be structurally classified into two main groups: the cyclopentenone group (e.g., aflatoxin B1 (AFB1 , Figure 1), aflatoxin M1 (AFM1 ), aflatoxin Q1 (AFQ1 ), and aflatoxicol), and the cyclo-lactone group (e.g., aflatoxin G1 (AFG1 ) and its derivatives); AFB1 , among the various aflatoxins, is commonly considered to be the most carcinogenic, inhibiting the synthesis of DNA, RNA and proteins [3]. Toxins 2020, 12, 476 derivative, is a known human carcinogen, but it can affect a wide range of commodities [4,5]. AFB1 and its metabolites are mainly distributed in corn, peanuts, rice, wheat, oil by-products, human carcinogen, but it can affect a wide range of commodities [4,5]. It is estimated that over 5 billion people worldwide are at risk mainly distributed in corn, peanuts, rice, wheat, oil by-products, dairy products and condiments [6,7]
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