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Strain Construction and Process Development for Efficient Recombinant Production of Mannuronan C-5 Epimerases in
Anne Tøndervik1, Randi Aune1, Adelheid Degelmann2
1Department of Biotechnology and Nanomedicine, SINTEF Industry, Trondheim, Norway.
Frontiers in Plant Science
|June 23, 2022
Summary
Hansenula polymorpha yeast can produce alginate epimerases for tailored alginate production. While larger enzymes showed degradation, it was manageable, indicating potential for industrial applications.
Area of Science:
- Biotechnology
- Biochemistry
- Microbial Engineering
Background:
- Alginates, polysaccharides from brown algae and bacteria, have industrial and medical applications.
- Mannuronan C-5 epimerases modify alginate structure, creating valuable G-rich blocks.
- Tailor-made alginates produced in vitro offer precise properties for various applications.
Purpose of the Study:
- To evaluate Hansenula polymorpha as a host for recombinant production of Azotobacter vinelandii alginate epimerases (AlgE1, AlgE4, AlgE6).
- To assess the suitability of H. polymorpha for large-scale industrial production of these enzymes.
Main Methods:
- Construction of H. polymorpha expression strains using optimized synthetic genes.
- High cell density cultivations to assess enzyme expression and stability.
- Investigated strategies to control proteolytic degradation of expressed epimerases.
Main Results:
- Larger epimerases (AlgE1, AlgE6) were susceptible to proteolytic degradation by yeast proteases.
- Degradation of AlgE1 and AlgE6 was significantly reduced by chaperone co-expression or optimized cultivation.
- The smaller AlgE4 epimerase demonstrated stability under all tested conditions.
Conclusions:
- Hansenula polymorpha shows potential for industrial production of alginate epimerases.
- Strategies exist to overcome challenges like proteolytic degradation for large-scale enzyme manufacturing.
- This yeast system facilitates the in vitro production of customized alginates with specific M-G patterns.
