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Aflatoxin detoxification by manganese peroxidase purified from Pleurotus ostreatus
1Department of Botany Faculty of Science Cairo University Giza Egypt.
Abstract:
Manganese peroxidase (MnP) was produced from white rot edible mushroom Pleurotus ostreatus on the culture filtrate. The enzyme was purified to homogeneity using (NH4)2SO4 precipitation, DEAE-Sepharose and Sephadex G-100 column chromatography. The final enzyme activity achieved 81 U mL(-1), specific activity 78 U mg(-1) with purification fold of 130 and recovery 1.2% of the crude enzyme. SDS-PAGE indicated that the pure enzyme have a molecular mass of approximately 42 kDa. The optimum pH was between 4-5 and the optimum temperature was 25 °C. The pure MnP activity was enhanced by Mn(2+), Cu(2+), Ca(2+) and K(+) and inhibited by Hg(+2) and Cd(+2). H2O2 at 5 mM enhanced MnP activity while at 10 mM inhibited it significantly. The MnP-cDNA encoding gene was sequenced and determined (GenBank accession no. AB698450.1). The MnP-cDNA was found to consist of 497 bp in an Open Reading Frame (ORF) encoding 165 amino acids. MnP from P. ostreatus could detoxify aflatoxin B1 (AFB1) depending on enzyme concentration and incubation period. The highest detoxification power (90%) was observed after 48 h incubation at 1.5 U mL(-1) enzyme activities.
Insights
This study details the purification and characterization of manganese peroxidase (MnP) from Pleurotus ostreatus. The purified enzyme effectively detoxified aflatoxin B1 (AFB1), showing potential for mycotoxin bioremediation.
Area of Science:
- Biochemistry
- Enzymology
- Mycology
Background:
- White rot fungi like Pleurotus ostreatus produce extracellular enzymes.
- Manganese peroxidase (MnP) is a key enzyme involved in lignin degradation.
- Bioremediation of mycotoxins is an important application for fungal enzymes.
Purpose of the Study:
- To purify and characterize manganese peroxidase (MnP) from Pleurotus ostreatus.
- To investigate the enzyme's potential for detoxifying aflatoxin B1 (AFB1).
Main Methods:
- Enzyme purification using ammonium sulfate precipitation and column chromatography (DEAE-Sepharose, Sephadex G-100).
- Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) for molecular mass determination.
- Enzyme activity assays under varying pH, temperature, metal ion, and H2O2 concentrations.
- DNA sequencing of the MnP-encoding gene and aflatoxin B1 detoxification assays.
Main Results:
- Purified MnP exhibited a specific activity of 78 U mg⁻¹ with a purification fold of 130.
- The enzyme has a molecular mass of approximately 42 kDa, optimal activity at pH 4-5 and 25 °C.
- MnP activity was influenced by various metal ions and H2O2 concentrations.
- The MnP-cDNA sequence (497 bp) encodes 165 amino acids.
- 90% detoxification of aflatoxin B1 was achieved with 1.5 U mL⁻¹ MnP after 48 hours incubation.
Conclusions:
- Pleurotus ostreatus is a source of purified manganese peroxidase.
- The characterized MnP shows significant potential for the bioremediation of aflatoxin B1.
