Browsing by Subject "Corynebacterium glutamicum"

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Design and evaluation of a 3D‐printed, lab‐scale perfusion bioreactor for novel biotechnological applications
(2023) Merkel, Manuel; Noll, Philipp; Lilge, Lars; Hausmann, Rudolf; Henkel, Marius
3D‐printing increased in significance for biotechnological research as new applications like lab‐on‐a‐chip systems, cell culture devices or 3D‐printed foods were uncovered. Besides mammalian cell culture, only few of those applications focus on the cultivation of microorganisms and none of these make use of the advantages of perfusion systems. One example for applying 3D‐printing for bioreactor development is the microbial utilization of alternative substrates derived from lignocellulose, where dilute carbon concentrations and harmful substances present a major challenge. Furthermore, quickly manufactured and affordable 3D‐printed bioreactors can accelerate early development phases through parallelization. In this work, a novel perfusion bioreactor system consisting of parts manufactured by fused filament fabrication (FFF) is presented and evaluated. Hydrophilic membranes are used for cell retention to allow the application of dilute substrates. Oxygen supply is provided by membrane diffusion via hydrophobic polytetrafluoroethylene membranes. An exemplary cultivation of Corynebacterium glutamicum ATCC 13032 supports the theoretical design by achieving competitive biomass concentrations of 18.4 g L−1 after 52 h. As a proof‐of‐concept for cultivation of microorganisms in perfusion mode, the described bioreactor system has application potential for bioconversion of multi‐component substrate‐streams in a lignocellulose‐based bioeconomy, for in‐situ product removal or design considerations of future applications for tissue cultures. Furthermore, this work provides a template‐based toolbox with instructions for creating reference systems in different application scenarios or tailor‐made bioreactor systems.
High‐level recombinant protein production with Corynebacterium glutamicum using acetate as carbon source
(2022) Kiefer, Dirk; Tadele, Lea Rahel; Lilge, Lars; Henkel, Marius; Hausmann, Rudolf
In recent years, biotechnological conversion of the alternative carbon source acetate has attracted much attention. So far, acetate has been mainly used for microbial production of bioproducts with bulk applications. In this study, we aimed to investigate the potential of acetate as carbon source for heterologous protein production using the acetate‐utilizing platform organism Corynebacterium glutamicum. For this purpose, expression of model protein eYFP with the promoter systems T7lac and tac was characterized during growth of C. glutamicum on acetate as sole carbon source. The results indicated a 3.3‐fold higher fluorescence level for acetate‐based eYFP production with T7 expression strain MB001(DE3) pMKEx2‐eyfp compared to MB001 pEKEx2‐eyfp. Interestingly, comparative eyfp expression studies on acetate or glucose revealed an up to 83% higher biomass‐specific production for T7 RNAP‐dependent eYFP production using acetate as carbon source. Furthermore, high‐level protein accumulation on acetate was demonstrated for the first time in a high cell density cultivation process with pH‐coupled online feeding control, resulting in a final protein titer of 2.7 g/L and product yield of 4 g per 100 g cell dry weight. This study presents a first proof of concept for efficient microbial upgrading of potentially low‐cost acetate into high‐value bioproducts, such as recombinant proteins.