Presentation Information
[4Ferm-10-KL]Improvement of fluid properties of culture medium in a stirred-tank bioreactor by cell-surface engineering of the industrial fungus Aspergillus oryzae
○Keietsu Abe1 (1. Graduate School of Agricultural Science, Tohoku University (Japan))
Keywords:
filamentous fungi,Aspergillus,bioreactor,cell wall,hydrophobin,fluid property
Filamentous fungi are industrially used for fermentative production of enzymes and chemicals. In such production processes, a large-scale stirred-tank bioreactor (STBR) is commonly used for culturing filamentous fungi due to the versatility of the culture equipment and the ease of adding nutrients and induction substrates to the culture tank and diffusing them within the tank. The scale of STBR ranges from tens of kL to hundreds of kL. Filamentous fungi exhibit various forms depending on the culture conditions, ranging from pellet-like structures where hyphae adhere and intertwine to form clumps, to pulp-like structures where hyphae are dispersed. This morphology affects the fluid properties of the culture medium, making it an unstable factor in fermentation production. The method that can completely control the morphology of filamentous fungi has not yet been established, posing a challenge in the fermentation industry. While it is expected that factors on the cell surface are involved in pellet formation of filamentous fungi under liquid culture conditions, the factors responsible for pellet formation remained unclear until recently.
In the present study, we developed a novel liquid culture technology that improves the fluid properties of culture media growing the industrial fungus Aspergillus oryzae. In the novel culture method, we bred A. oryzae hyphal dispersion mutants where the biosynthetic genes of cell wall polysaccharides functioning as hyphal adhesion factors were disrupted. We previously found that a gene-deficient strain of the model filamentous fungus Aspergillus nidulans lacking the α-1,3-glucan (AG) synthase gene did not form hyphal pellets but instead dispersed its hyphal structure, and thus identified AG as a hyphal adhesion factor. In A. oryzae, complete hyphae dispersal was achieved for the first time in an AG-GAG double knockout strain (AGΔ-GAGΔ), which lacked not only AG deficiency but also the ability to synthesize the extracellular matrix polysaccharide galactosaminogalactan (GAG), thus identifying GAG as a second hyphae adhesion factor.1) Through computational fluid dynamics (CFD) analysis of STBR culture, we revealed that the AGΔ-GAGΔ strain exhibits higher enzyme productivity compared to the wild-type strain, due to its hyphal dispersibility and low culture medium viscosity, which improve oxygen utilization efficiency in the liquid phase.2) We also found that the additional deletion of the rolA gene encoding a biosurfactant protein hydrophobin RolA in A. oryzae, reduces wall growth in the culture vessel and further improves the liquid properties. Controlling both the fungal morphology and fluid properties of culture broth through the displaying levels of cell surface molecules such as polysaccharides and surfactant proteins is a new technology for industrial fermentation of filamentous fungi.
1) Miyazawa K. et al., Front. Microbiol. 10:2090 (2019)
2) Susukida S. et al., Biotechnol. Bioeng. 122:2389-2399 (2025)
In the present study, we developed a novel liquid culture technology that improves the fluid properties of culture media growing the industrial fungus Aspergillus oryzae. In the novel culture method, we bred A. oryzae hyphal dispersion mutants where the biosynthetic genes of cell wall polysaccharides functioning as hyphal adhesion factors were disrupted. We previously found that a gene-deficient strain of the model filamentous fungus Aspergillus nidulans lacking the α-1,3-glucan (AG) synthase gene did not form hyphal pellets but instead dispersed its hyphal structure, and thus identified AG as a hyphal adhesion factor. In A. oryzae, complete hyphae dispersal was achieved for the first time in an AG-GAG double knockout strain (AGΔ-GAGΔ), which lacked not only AG deficiency but also the ability to synthesize the extracellular matrix polysaccharide galactosaminogalactan (GAG), thus identifying GAG as a second hyphae adhesion factor.1) Through computational fluid dynamics (CFD) analysis of STBR culture, we revealed that the AGΔ-GAGΔ strain exhibits higher enzyme productivity compared to the wild-type strain, due to its hyphal dispersibility and low culture medium viscosity, which improve oxygen utilization efficiency in the liquid phase.2) We also found that the additional deletion of the rolA gene encoding a biosurfactant protein hydrophobin RolA in A. oryzae, reduces wall growth in the culture vessel and further improves the liquid properties. Controlling both the fungal morphology and fluid properties of culture broth through the displaying levels of cell surface molecules such as polysaccharides and surfactant proteins is a new technology for industrial fermentation of filamentous fungi.
1) Miyazawa K. et al., Front. Microbiol. 10:2090 (2019)
2) Susukida S. et al., Biotechnol. Bioeng. 122:2389-2399 (2025)
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