The filamentous fungus Aspergillus niger is known for its high secretion efficiency and capacity of homologous proteins and is therefore an important production strain in biotechnological processes, especially in the field of nutrition additives. Furthermore, its ability to carry out post-translational modifications results in an increasing usage of A. niger as production host for the formation of therapeutic agents. However, the obtainable yields of recombinant proteins are considerably lower than of homologous proteins. Hence, current research is focussed on genetically engineered strain improvement and the optimization of cultivation processes to increase the production of desired recombinant products. Depending on the environmental parameters A. niger growth occurs either as distinct pellets or as freely dispersed mycelia in submerged cultivations, whereas the optimal type of morphology varies due to specific product properties. Protein secretion is mainly observed in the region of growing hyphal tips. In this study, the production of a recombinant protein is analysed in submerged cultivations of a genetically modified A. niger strain. Its genome comprises extra copies of the homologous beta-fructofuranosidase gene regulated by a constitutive promoter. The influences of different environmental parameters on morphology as well as the resulting productivity in regard to the recombinant product are investigated. The transcription activity of genes involved in hyphal branching, septation and polarized growth and its alteration in course of the cultivation process are quantified by real-time PCR. Results are related to microscopic images. Furthermore, the corresponding gene expression of beta-fructofuranosidase as model product is monitored via real-time PCR. The resulting enzyme activities within the bulk phase of the cultivation and the biomass-linked product activity are determined. As a result, the presented work intends to elucidate the influence of different environmental parameters on the morphological development even on the transcriptional level. This should allow a thorough characterization of protein formation as well as product secretion in dependency of morphological structures.
Full conference title:
9th EUROPEAN CONFERENCE ON FUNGAL GENETICS
- ECFG 9th (2008)