Genetically engineered bacteria and microalgae expressing a mutant of cytochrome P450 BM3 for efficient Diuron degradation in wastewater treatment
ABSTRACT The diverse members of the cytochrome P450 superfamily conduct the metabolism of a wide variety of compounds, both endogenous and xenobiotic, thus offering potential for degrading emerging contaminants such as prescription drugs and pesticides in wastewater treatment and aquaculture. This s...
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American Society for Microbiology
2025-06-01
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| Series: | Microbiology Spectrum |
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| Online Access: | https://journals.asm.org/doi/10.1128/spectrum.02905-24 |
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| author | Christian Helvig Thamali Kariyawasam Bas Vriens Martin Petkovich |
| author_facet | Christian Helvig Thamali Kariyawasam Bas Vriens Martin Petkovich |
| author_sort | Christian Helvig |
| collection | DOAJ |
| description | ABSTRACT The diverse members of the cytochrome P450 superfamily conduct the metabolism of a wide variety of compounds, both endogenous and xenobiotic, thus offering potential for degrading emerging contaminants such as prescription drugs and pesticides in wastewater treatment and aquaculture. This study demonstrates the expression and activity of the mutant P450 enzyme of CYP102A1 (BM3 MT35) engineered in Bacillus megaterium and Chlamydomonas reinhardtii, harnessed for the degradation of the herbicide Diuron. The P450 BM3 MT35 enzyme was transgenically expressed in Escherichia coli and purified via a His6-tag system, with its activity confirmed by spectral and in vitro metabolism assays. During in vivo experiments, transgenic Bacillus megaterium degraded 23% of Diuron after 24 h, reaching 65% by day 5 in Terrific Broth (TB) media. In synthetic and municipal wastewater, Diuron degradation reached 45% and 15%, respectively. No Diuron metabolism occurred in wild-type (WT) Bacillus megaterium in all the above tested conditions. Additionally, transgenic Chlamydomonas reinhardtii expressing P450 BM3 MT35 in the chloroplast also exhibited enhanced Diuron degradation (52%) compared to WT strains (6%). Overall, our results demonstrate the potential of genetically engineered bacteria and microalgae with cytochrome P450 as a viable strategy for the enhanced treatment of emerging contaminants in wastewater, potentially offering a new, sustainable alternative approach to pollution mitigation.IMPORTANCEWith a growing number and variety of prescription drugs, pesticides, food additives, and chemicals produced, wastewater is being contaminated with an increasing number of emerging pollutants that cannot be eliminated through classical wastewater treatment. New methods should therefore be developed to remove or deactivate these contaminants. Here, we demonstrate that by using genetically engineered bacteria and microalgae expressing a mutated enzyme, it is possible to efficiently metabolize a targeted pesticide, in this case, Diuron. These new findings should open the door to new ways to treat wastewater by developing low-cost and efficient modified microorganisms that will be able to specifically detoxify past and new emerging water contaminants that cannot be eliminated through classical wastewater treatment. |
| format | Article |
| id | doaj-art-2ca5ee96f63c4a2398455e8fa19897bf |
| institution | OA Journals |
| issn | 2165-0497 |
| language | English |
| publishDate | 2025-06-01 |
| publisher | American Society for Microbiology |
| record_format | Article |
| series | Microbiology Spectrum |
| spelling | doaj-art-2ca5ee96f63c4a2398455e8fa19897bf2025-08-20T02:24:25ZengAmerican Society for MicrobiologyMicrobiology Spectrum2165-04972025-06-0113610.1128/spectrum.02905-24Genetically engineered bacteria and microalgae expressing a mutant of cytochrome P450 BM3 for efficient Diuron degradation in wastewater treatmentChristian Helvig0Thamali Kariyawasam1Bas Vriens2Martin Petkovich3Department of Biomedical and Molecular Sciences, Queen's University, Kingston, Ontario, CanadaDepartment of Biomedical and Molecular Sciences, Queen's University, Kingston, Ontario, CanadaDepartment of Geological Sciences and Engineering, Queen's University, Kingston, Ontario, CanadaDepartment of Biomedical and Molecular Sciences, Queen's University, Kingston, Ontario, CanadaABSTRACT The diverse members of the cytochrome P450 superfamily conduct the metabolism of a wide variety of compounds, both endogenous and xenobiotic, thus offering potential for degrading emerging contaminants such as prescription drugs and pesticides in wastewater treatment and aquaculture. This study demonstrates the expression and activity of the mutant P450 enzyme of CYP102A1 (BM3 MT35) engineered in Bacillus megaterium and Chlamydomonas reinhardtii, harnessed for the degradation of the herbicide Diuron. The P450 BM3 MT35 enzyme was transgenically expressed in Escherichia coli and purified via a His6-tag system, with its activity confirmed by spectral and in vitro metabolism assays. During in vivo experiments, transgenic Bacillus megaterium degraded 23% of Diuron after 24 h, reaching 65% by day 5 in Terrific Broth (TB) media. In synthetic and municipal wastewater, Diuron degradation reached 45% and 15%, respectively. No Diuron metabolism occurred in wild-type (WT) Bacillus megaterium in all the above tested conditions. Additionally, transgenic Chlamydomonas reinhardtii expressing P450 BM3 MT35 in the chloroplast also exhibited enhanced Diuron degradation (52%) compared to WT strains (6%). Overall, our results demonstrate the potential of genetically engineered bacteria and microalgae with cytochrome P450 as a viable strategy for the enhanced treatment of emerging contaminants in wastewater, potentially offering a new, sustainable alternative approach to pollution mitigation.IMPORTANCEWith a growing number and variety of prescription drugs, pesticides, food additives, and chemicals produced, wastewater is being contaminated with an increasing number of emerging pollutants that cannot be eliminated through classical wastewater treatment. New methods should therefore be developed to remove or deactivate these contaminants. Here, we demonstrate that by using genetically engineered bacteria and microalgae expressing a mutated enzyme, it is possible to efficiently metabolize a targeted pesticide, in this case, Diuron. These new findings should open the door to new ways to treat wastewater by developing low-cost and efficient modified microorganisms that will be able to specifically detoxify past and new emerging water contaminants that cannot be eliminated through classical wastewater treatment.https://journals.asm.org/doi/10.1128/spectrum.02905-24emerging contaminantswastewater treatmentmetabolismgenetically engineeredcytochrome P450algae |
| spellingShingle | Christian Helvig Thamali Kariyawasam Bas Vriens Martin Petkovich Genetically engineered bacteria and microalgae expressing a mutant of cytochrome P450 BM3 for efficient Diuron degradation in wastewater treatment Microbiology Spectrum emerging contaminants wastewater treatment metabolism genetically engineered cytochrome P450 algae |
| title | Genetically engineered bacteria and microalgae expressing a mutant of cytochrome P450 BM3 for efficient Diuron degradation in wastewater treatment |
| title_full | Genetically engineered bacteria and microalgae expressing a mutant of cytochrome P450 BM3 for efficient Diuron degradation in wastewater treatment |
| title_fullStr | Genetically engineered bacteria and microalgae expressing a mutant of cytochrome P450 BM3 for efficient Diuron degradation in wastewater treatment |
| title_full_unstemmed | Genetically engineered bacteria and microalgae expressing a mutant of cytochrome P450 BM3 for efficient Diuron degradation in wastewater treatment |
| title_short | Genetically engineered bacteria and microalgae expressing a mutant of cytochrome P450 BM3 for efficient Diuron degradation in wastewater treatment |
| title_sort | genetically engineered bacteria and microalgae expressing a mutant of cytochrome p450 bm3 for efficient diuron degradation in wastewater treatment |
| topic | emerging contaminants wastewater treatment metabolism genetically engineered cytochrome P450 algae |
| url | https://journals.asm.org/doi/10.1128/spectrum.02905-24 |
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