04 Fakultät Energie-, Verfahrens- und Biotechnik
Permanent URI for this collectionhttps://elib.uni-stuttgart.de/handle/11682/5
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Item Open Access Methylthioadenosine (MTA) boosts cell‐specific productivities of Chinese hamster ovary cultures : dosage effects on proliferation, cell cycle and gene expression(2020) Verhagen, Natascha; Zieringer, Julia; Takors, RalfA major goal for process and cell engineering in the biopharmaceutical industry is enhancing production through increasing volumetric and cellspecific productivities (CSP). Here, we present 50-deoxy-50-(methylthio)adenosine (MTA), the degradation product of S-(50-adenosyl)-L-methionine (SAM), as a highly attractive native additive which can boost CSP by 79% when added to exponentially growing cells at a concentration of 250-300 lM. Notably, cell viability and cell size remain higher than in non-treated cultures. In addition, cell cycle arrests first in S-, then in G2-phase before levelling out compared to non-treated cultivations. Intensive differential gene analysis reveals that expression of genes for cytoskeleton mediated proteins and vesicle transport is amplified by treatment. Furthermore, the interaction of MTA with cell proliferation additionally stimulated recombinant protein formation. The results may serve as a promising starting point for further developments in process and cell engineering to boost productivity.Item Open Access Investigation of tracer gas transport in a new numerical model of lung acini(2022) Schmidt, Christoph; Joppek, Christoph; Trinkmann, Frederik; Takors, Ralf; Cattaneo, Giorgio; Port, JohannesObstructive pulmonary diseases are associated with considerable morbidity. For an early diagnosis of these diseases, inert gas washouts can potentially be used. However, the complex interaction between lung anatomy and gas transport mechanisms complicates data analysis. In order to investigate this interaction, a numerical model, based on the finite difference method, consisting of two lung units connected in parallel, was developed to simulate the tracer gas transport within the human acinus. Firstly, the geometries of the units were varied and the diffusion coefficients ( D ) were kept constant. Secondly, D was changed and the geometry was kept constant. Furthermore, simple monoexponential growth functions were applied to evaluate the simulated data. In 109 of the 112 analyzed curves, monoexponential function matched simulated data with an accuracy of over 90%, potentially representing a suitable numerical tool to predict transport processes in further model extensions. For total flows greater than 5 × 10 -4 ml/s, the exponential growth constants increased linearly with linear increasing flow to an accuracy of over 95%. The slopes of these linear trend lines of 1.23 µl -1 ( D = 0.6 cm 2 /s), 1.69 µl -1 ( D = 0.3 cm 2 /s), and 2.25 µl -1 ( D = 0.1 cm 2 /s) indicated that gases with low D are more sensitive to changes in flows than gases with high D .Item Open Access Balancing glucose and oxygen uptake rates to enable high amorpha‐4,11‐diene production in Escherichia coli via the methylerythritol phosphate pathway(2021) Patil, Vikas; Santos, Christine N. S.; Ajikumar, Parayil K.; Sarria, Stephen; Takors, RalfAmorpha‐4,11‐diene (AMD4,11) is a precursor to artemisinin, a potent antimalarial drug that is traditionally extracted from the shrubs of Artemisia annua. Despite significant prior efforts to produce artemisinin and its precursors through biotechnology, there remains a dire need for more efficient biosynthetic routes for its production. Here, we describe the optimization of key process conditions for an Escherichia coli strain producing AMD4,11 via the native methylerythritol phosphate (MEP) pathway. By studying the interplay between glucose uptake rates and oxygen demand, we were able to identify optimal conditions for increasing carbon flux through the MEP pathway by manipulating the availability of NADPH required for terpenoid production. Installation of an optimal qO2/qglucose led to a 6.7‐fold increase in product titers and a 6.5‐fold increase in carbon yield.Item Open Access Optimum blue light exposure : a means to increase cell-specific productivity in Chinese hamster ovary cells(2024) Föller, Stefanie; Regett, Niklas; Lataster, Levin; Radziwill, Gerald; Takors, RalfResearch for biopharmaceutical production processes with mammalian cells steadily aims to enhance the cell-specific productivity as a means for optimizing total productivities of bioreactors. Whereas current technologies such as pH, temperature, and osmolality shift require modifications of the cultivation medium, the use of optogenetic switches in recombinant producer cells might be a promising contact-free alternative. However, the proper application of optogenetically engineered cells requires a detailed understanding of basic cellular responses of cells that do not yet contain the optogenetic switches. The knowhow of ideal light exposure to enable the optimum use of related approaches is missing so far. Consequently, the current study set out to find optimum conditions for IgG1 producing Chinese hamster ovary (CHO) cells which were exposed to blue LED light. Growth characteristics, cell-specific productivity using enzyme-linked immunosorbent assay, as well as cell cycle distribution using flow cytometry were analyzed. Whereas too harsh light exposure causes detrimental growth effects that could be compensated with antioxidants, a surprising boost of cell-specific productivity by 57% occurred at optimum high light doses. The increase coincided with an increased number of cells in the G1 phase of the cell cycle after 72 h of illumination. The results present a promising new approach to boost biopharmaceutical productivity of mammalian cells simply by proper light exposure without any further optogenetic engineering.