A fungal biofilm reactor based on metal structured packing improves the quality of a Gla::GFP fusion protein produced by Aspergillus oryzae
article
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.
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.
Topics
Fungal biofilmBioreactorScale-upRecombinant proteinAntigen-antibody reactionsAspergillusBiofilmsBiomassBioreactorsElectrophoresisEnzyme activityFermentationFungiMetabolitesPhysiologyRecombinant proteinsShear stressComparative analysisExtracellular mediumGreen fluorescent proteinPhysiological mechanismsSecondary metabolitesSolid-state fermentationSubmerged fermentationProteinsAmylaseGlucan 1,4 alpha glucosidaseGlucoamylase green fluorescent protein fusion proteinGreen fluorescent proteinHybrid proteinUnclassified drugFermentationFungusMetalPhysiological responseProteinQualitative analysisRecirculating systemRecombinationSecondary metaboliteAlkalinizationAmino acid sequenceAspergillus nigerAspergillus oryzaeAutolysisBiofilm reactorBiotransformationCarbon sourceControlled studyEnzyme activityExtracellular spaceFungal biomassFungal colonizationFungus cultureFungus growthGlucose intakeHydrolysisImmersionImmunodetectionNonhumanPH measurementProtein analysisShear stressSolid state fermentationSubmerged fermentationWestern blotting
TNO Identifier
525956
Source
Applied Microbiology and Biotechnology, 99(15), pp. 6241-6254.
Pages
6241-6254
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