Enzymatic properties of a bacterial microcystinase A produced in Saccharomyces cerevisiae

Authors: DE GODOI SILVA, FERNANDO - University North Of Parana; GREINER, RALF - Max Rubner-Institut (MRI); DIAS LOPES, DAIANE - Orise Fellow; Hector, Ronald; HASHIMOTO, ELISABETE - Federal University Of Technology - Parana; HIROOKA, ELISA - University North Of Parana

Submitted to: Biotechnology Progress
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 9/27/2025
Publication Date: 10/16/2025
Citation: De Godoi Silva, F., Greiner, R., Dias Lopes, D., Hector, R.E., Hashimoto, E.H., Hirooka, E.Y. 2025. Enzymatic properties of a bacterial microcystinase A produced in Saccharomyces cerevisiae. Biotechnology Progress. https://doi.org/10.1002/btpr.70083.
DOI: https://doi.org/10.1002/btpr.70083

Interpretive Summary: Microcystins are dangerous toxins produced by certain types of blue-green algae, called Cyanobacteria, that pose a growing risk to public health, especially in water sources. To help combat this threat, this research is identifying ways to remove these toxins more effectively. One promising solution involves using a natural enzyme called microcystinase A, which can break down the microcystins. This enzyme was originally discovered in a type of bacteria, but we have now successfully produced it using brewer’s yeast, making it easier to access, characterize, and use. This study identified the best conditions for maximizing the enzyme’s activity for degrading microcystins. These findings suggest that the enzyme could be an effective tool for cleaning up microcystin contaminated water.

Technical Abstract: Microcystins (MCs) are toxins produced by Cyanobacteria, posing a significant emerging threat to human and public health. Thus, control strategies combining frequent toxin monitoring with removal techniques are urgently needed. In this context, microcystin degradation using the bacterial enzyme microcystinase A, originally derived from Sphingosinicella microcystinivorans B9, has been identified as a sustainable and effective approach. To facilitate access to the enzyme, the gene encoding microcystinase A was successfully expressed in the Saccharomyces cerevisiae PE-2 strain. The recombinant microcystinase A was produced as an intracellular enzyme and applied in MC degradation assays. Optimal conditions for enzymatic activity were identified at 42.2°C and pH 6.3. The maximum degradation rate of microcystin was determined to be 3.09 mg/L/h, and a K' of 2.81 µM was obtained when assays were performed at 37°C and pH 7.4. The recombinant microcystinase A remained fully active for 2 hours at 20°C. Exposure to 50°C for one hour resulted in 60% residual activity, while 30 minutes at 65°C led to complete inactivation. The enzyme was also denatured when exposed to alkaline pH conditions. Therefore, the results of this study indicate that recombinant microcystinase A shows potential for MC degradation under mildly acidic conditions and temperatures up to 45°C.