Fungal biofilm reactor improves the quality of a fusion protein GLA::GFP produced by Aspergillus oryzae

Fungal biofilm is known to promote the excretion of secondary metabolites, in accordance with solid-state related physiological mechanisms. In this work, the potentialities of fungal biofilm will be investigated in the context of the production of a Gla::GFP fusion protein by Aspergillus oryzae. Since the production of this protein is under the control of the promoter glaB, specifically induced in solid-state fermentation, biofilm mode of culture is expected to enhance the global productivity. However, we found that the glaB promoter is also activated in submerged bioreactor and the fusion protein production is higher in this mode of culture. This result is related to the high shear stress leading to biomass autolysis and leakage of intracellular fusion protein into the extracellular medium. Moreover, 2D-gel electrophoresis highlights preservation of the fusion protein integrity produced in biofilm conditions whereas proteolysis strongly affects fusion protein recovery in the submerged cultures performed at high stirring rate. Fungal biofilm reactor design was then further investigated and the scale-up potentialities were evaluated. Indeed, the specific design investigated in this work involves the use of metal structured packing exhibiting a high specific area and that can be easily expanded to large-scale bioprocessing conditions.