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A fungal biofilm reactor based on metal structured packing improves the quality of a Gla::GFP fusion protein produced by Aspergillus oryzae

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Author: Zune, Q. · Delepierre, A. · Gofflot, S. · Bauwens, J. · Twizere, J.C. · Punt, P.J. · Francis, F. · Toye, D. · Bawin, T. · Delvigne, F.
Type:article
Date:2015
Source:Applied Microbiology and Biotechnology, 15, 99, 6241-6254
Identifier: 525956
doi: doi:10.1007/s00253-015-6608-z
Keywords: Nutrition · Fungal biofilm · Bioreactor · Scale-up · Recombinant protein · Antigen-antibody reactions · Aspergillus · Biofilms · Biomass · Bioreactors · Electrophoresis · Enzyme activity · Fermentation · Fungi · Metabolites · Physiology · Recombinant proteins · Shear stress · Comparative analysis · Extracellular medium · Green fluorescent protein · Physiological mechanisms · Secondary metabolites · Solid-state fermentation · Submerged fermentation · Proteins · Amylase · Glucan 1,4 alpha glucosidase · Glucoamylase green fluorescent protein fusion protein · Green fluorescent protein · Hybrid protein · Unclassified drug · Fermentation · Fungus · Metal · Physiological response · Protein · Qualitative analysis · Recirculating system · Recombination · Secondary metabolite · Alkalinization · Amino acid sequence · Aspergillus niger · Aspergillus oryzae · Autolysis · Biofilm reactor · Biotransformation · Carbon source · Controlled study · Enzyme activity · Extracellular space · Fungal biomass · Fungal colonization · Fungus culture · Fungus growth · Glucose intake · Hydrolysis · Immersion · Immunodetection · Nonhuman · PH measurement · Protein analysis · Shear stress · Solid state fermentation · Submerged fermentation · Western blotting · Food and Nutrition · Healthy Living · Life · MSB - Microbiology and Systems Biology · ELSS - Earth, Life and Social Sciences

Abstract

Fungal biofilm is known to promote the excretion of secondary metabolites in accordance with solid-staterelated physiological mechanisms. This work is based on the comparative analysis of classical submerged fermentation with a fungal biofilmreactor for the production of a Gla::green fluorescent protein (GFP) fusion protein by Aspergillus oryzae. The biofilmreactor comprises a metal structured packing allowing the attachment of the fungal biomass. Since the production of the target protein is under the control of the promoter glaB, specifically induced in solid-state fermentation, the biofilm mode of culture is expected to enhance the global productivity. Although production of the target protein was enhanced by using the biofilm mode of culture, we also found that fusion protein production is also significant when the submerged mode of culture is used. This result is related to high shear stress leading to biomass autolysis and leakage of intracellular fusion protein into the extracellular medium. Moreover, 2-D gel electrophoresis highlights the preservation of fusion protein integrity produced in biofilm conditions. Two fungal biofilm reactor designs were then investigated further, i.e.with full immersion of the packing or with medium recirculation on the packing, and the scale-up potentialities were evaluated. In this context, it has been shown that full immersion of the metal packing in the liquid medium during cultivation allows for a uniformcolonization of the packing by the fungal biomass and leads to a better quality of the fusion protein.