A New Laccase of Lac 2 from the White Rot Fungus Cerrena unicolor 6884 and Lac 2-Mediated Degradation of Aflatoxin B1
Abstract
:1. Introduction
2. Results
2.1. Species Identification
2.2. Preparation and Purification
2.3. Laccase Identification
2.4. AFB1 Degradation by Lac 2 from Cerrena Unicolor 6884
2.5. Analysis of Products of AFB1 Degradation Catalyzed by Lac 2
3. Discussion
4. Conclusions
5. Materials and Methods
5.1. Organism, Chemicals and Other Materials
5.2. Phylogenetic Analysis
5.3. Illumina Transcriptome Analysis of Cerrena Unicolor 6884
5.4. Production of Laccase
5.5. Enzyme Assay
5.6. Purification of Laccase
5.7. Identification of Laccase by UHPLC-MS/MS
5.8. Cloning of the Laccase cDNA from Cerrena Unicolor 6884
5.9. In Vitro Degradation of AFB1 with Lac 2
5.10. AFB1 Assay
5.11. UPLC-MS/MS Analysis of AFB1 Degradation Products
Supplementary Materials
Author Contributions
Funding
Conflicts of Interest
Abbreviations
ABTS | [2,2′ -azino-bis-(3-ethylbenzothiazoline-6-sulfonic acid)] |
ACN | Acetonitrile |
AFB1 | aflatoxin B1 |
AFG1 | aflatoxin G1 |
AFM1 | aflatoxin M1 |
AFQ1 | aflatoxin Q1 |
AS | Acetosyringone |
DAD | Diode array detector |
DEAE | diethylamino ethyl |
FLD | Fluorescent detector |
HBT | Hydroxy benzotriazole |
Lac 2 | laccase 2 |
LMS | laccase mediator system |
SA | Syrinagaldehyde |
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Purification Step | Total Activity (U) | Total Protein (mg) | Specific Activity (U/mg) | Purification Fold | Recovery (%) |
---|---|---|---|---|---|
Crude extract | 44,641.67 | 655.86 | 68.07 | 1.00 | 100 |
NH4(SO4)2 | 33,653.33 | 163.49 | 205.84 | 3.02 | 75.39 |
DEAE cellulose | 6592.00 | 4.01 | 1643.70 | 24.15 | 14.77 |
Assigned Protein | Coverage [%] | Peptides | Sequence | Confidence | Theo. MH+ [Da] |
---|---|---|---|---|---|
Lac 2 from Cerrena. unicolor 6884 | 48 | 13 | VVELVIPPLAVGGPHPFHLHGHNFWVVR | High | 3123.71556 |
TVGGPAQSPLNEADLRPLVPAPVPGNAVPGGADINHR | 3653.91467 | ||||
SQTGPADAELAVISVEHNKR | 2122.08872 | ||||
SQTGPADAELAVISVEHNK | 1965.98761 | ||||
SAGSDEYNFDDAILRDVVSIGAGTDEVTIR | 3185.52331 | ||||
SAGSDEYNFDDAILR | 1672.74492 | ||||
NASVEEPK | 873.43124 | ||||
NAAILR | 657.40423 | ||||
MLTPTSIHWHGFFQK | 1845.91049 | ||||
YSFVLNANQPDDNYWIR | 2114.99303 | ||||
MLTPTSIHWHGFFQK | 1829.91557 | ||||
GAFVVYDPNDPHK | 1458.70120 | ||||
DVVSIGAGTDEVTIR | 1531.79623 | ||||
DLYDVDDESTVITLADWYHVLAQTVVGAATPDSTLINGLGR | 4404.18816 |
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Zhou, Z.; Li, R.; Ng, T.B.; Lai, Y.; Yang, J.; Ye, X. A New Laccase of Lac 2 from the White Rot Fungus Cerrena unicolor 6884 and Lac 2-Mediated Degradation of Aflatoxin B1. Toxins 2020, 12, 476. https://doi.org/10.3390/toxins12080476
Zhou Z, Li R, Ng TB, Lai Y, Yang J, Ye X. A New Laccase of Lac 2 from the White Rot Fungus Cerrena unicolor 6884 and Lac 2-Mediated Degradation of Aflatoxin B1. Toxins. 2020; 12(8):476. https://doi.org/10.3390/toxins12080476
Chicago/Turabian StyleZhou, Zhimin, Renkuan Li, Tzi Bun Ng, Yunyun Lai, Jie Yang, and Xiuyun Ye. 2020. "A New Laccase of Lac 2 from the White Rot Fungus Cerrena unicolor 6884 and Lac 2-Mediated Degradation of Aflatoxin B1" Toxins 12, no. 8: 476. https://doi.org/10.3390/toxins12080476