Team:DTU-Denmark/Proof Of Concept
The overall goal of the project was to optimize Aspergillus niger as a cell factory through the engineering of morphology. While filamentous fungi are great producers of both proteins and secondary metabolites, they are often disregarded due to their filamentous structure being difficult to work with. We hoped the altered morphology would lead to ease of production as well as increased production.
Top: Plate pictures of the different strains grown on Czepek Yeast Extract Agar. Bottom: Microscopy pictures of the different strains. The size bar shows 75μm.
Three strains with high potential as improved cell factories were identified. The knockout strains Δgul-1, ΔchsC_Δgul-1 and ΔspaA_Δgul-1 all showed increased specific glucoamylase activity compared to the reference strain. Our project used glucoamylase as a reference product due to it being an industrially relevant protein as well as being easily measurable. From the figure below it can be seen that all three mutants achieved a more than two-fold increase in specific glucoamylase activity. Δgul-1 achieved the greatest increase of 290%. Furthermore, the growth rate remained close to that of the reference strain with Δgul-1 and ΔspaA_Δgul-1 having a reduction of 2-14% and 1-13% respectively and ΔchsC_Δgul-1 having an increase in growth rate of 14-24%. (See measurements pagefor details on how these experiments were conducted.)
Protein secretion and glucoamylase activity for Δgul-1, ΔchsC_Δgul-1 and ΔspaA_Δgul-1 compared to the reference strain ATCC 1015 strains run in the bioreactor for the last time-point samples. Green: Glucoamylase activity in UA/mL. Blue: Specific activity in UA/mg calculated from the activity and the protein concentration. Purple: Protein concentration in mg/mL.
Bar chart of the growth rate for both duplicates for Δgul-1, ΔchsC_Δgul-1, ΔspaA_Δgul-1 strains compared to the reference strain ATCC 1015 obtained from the bioreactor.
These results confirmed our original hypothesis that morphology has an impact on protein production. This is significant as increasing protein production while maintaining a similar or increased growth rate would lead to decreased production costs compared to existing systems. We hypothesize that similar results might be obtained in strains engineered to produce other proteins greatly benefiting industrial players. Thoughts on how this could be implemented in industry can be read on our implementation page.Through alterations in morphology, we have reached our goal of making a strain that increases protein production without compromising the growth rate.
