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Multi-enzyme Screening Using a High-throughput Genetic Enzyme Screening System
Published on: August 8, 2016
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Rapid screening system to identify unspecific peroxygenase activity.
Marina Schramm1, Carlos Renato Carrillo Avilés1, Johannes Kalmbach1
1Institute of Biotechnology, Brandenburg University of Technology Cottbus-Senftenberg, Senftenberg, Germany.
Clinical Hemorheology and Microcirculation
|April 28, 2025
Summary
We developed a simple screening method for identifying active unspecific peroxygenases (UPOs) in yeast. This high-throughput approach quickly found two new UPOs capable of producing human drug metabolites.
Area of Science:
- Biocatalysis
- Enzymology
- Synthetic Chemistry
Background:
- Unspecific peroxygenases (UPOs) are crucial biocatalysts for synthesizing pharmaceutical metabolites.
- Identifying new, expressible UPOs is challenging due to costly and time-consuming screening methods.
Purpose of the Study:
- To develop a simple, high-throughput agar plate-based method for screening active, secreted UPOs.
- To identify novel UPOs from a gene library expressed in Saccharomyces cerevisiae.
Main Methods:
- Heterologous expression of a UPO gene library in Saccharomyces cerevisiae.
- Agar plate-based screening for secreted UPO activity.
- Partial purification and characterization of identified UPOs.
Main Results:
- A novel, high-throughput screening method for UPOs was successfully established.
- Two short UPOs, DbiUPO from Dendrothele bispora and AniUPO from Aspergillus niger, were identified.
- Both UPOs demonstrated catalytic activity in producing human drug metabolites, including lipid mediators and clopidogrel's active metabolite.
Conclusions:
- The developed screening method enables rapid identification of active UPOs.
- The newly identified DbiUPO and AniUPO are valuable biocatalysts for producing specific human drug metabolites.
