Current research

Current research projects are running under the roof of the funding initiative "Biotechnologie 2020+" initiated by the German Federal Ministry for Education and Research (BMBF)
http://biooekonomie.de/initiative-biotechnologie-2020
 

1. CASCOO – Bacterial Pathways and Enzymes for Conversion of CO2

The utilization of the climate gas CO2 as a feedstock for chemical and biotechnological processes has become a major focus in the strategies for fighting climate change and global warming.
In bacteria and archeae, six pathways are now known which allow autotrophic growth using CO2 as sole carbon source. In addition, CO2 fixation is also involved in numerous carboxylating enzymatic reactions during heterotrophic growth with organic carbon sources.
However, these enzymes have so far not been characterized in detail for their capacity and suitability for technical applications.
In this project, we isolate enzymes from different bacterial species which might be useful for conversion of CO2 into organic compounds.
In addition, we apply a systems biology approach using a combination of computational modeling and wet-lab experiments (in vitro enzyme activities, knock-out mutants, bioreactor cultivations, metabolimics, 13C metabolic flux analysis) for exploring CO2-fixing pathways in the facultative photosynthetic bacterium Rhodospirillum rubrum (Fig. 1) during heterotrophic growth with acetate.
 
Collaboration partner: Max-Planck-Insititute for Dynamics of Complex Technical Systems, Magdeburg

2. EnzCaps – Selection of Organic Solvent-tolerant Bacterial Strains and Enzymes

Fig 1. Fluorescence micrograph of Rhodospirillum rubrum cells (1000 x).

Bacterial cells show amazing capabilities to rapidly adapt to environmental stress factors and unfavourable growth conditions. An important example is the dramatic raise of antibiotic resistant variants during antibiotic treatment of pathogenic bacteria. In industrial biotechnology, however, these evolutionary mechanisms can be exploited to rapidly evolve microbial strains with desired properties. The underlying mechanisms of adaptive evolution are still only poorly understood and not predictable, thus preventing metabolic engineering approaches to develop the respective strains on a rational basis.
Specifically, we here target mutants of the facultative photosynthetic bacterium Rhodospirillum rubrum with increased tolerance towards organic solvents. For rapid selection of improved strains, cultivation under selective pressure is performed in continuous bioreactor cultivations (Cytostat) and by serial dilution in batch cultures. From the isolated mutants, transcriptomic data of sequence changes and gene expression levels are obtained by RNA-Seq using NexGenerationSequencing. The results should be helpful for obtaining a better rationale of the genetic changes that give rise to adaptive mutants and for identification of modified enzymes for biocatalysis in the presence of organic solvents.
 
Collaboration with University Stuttgart, Institute of Biomaterials and biomolecular Systems, Department of Bioenergetics and the Natural and Medical Sciences Institute (NMI) at the University of Tübingen
 
 

3. NXP-SynPath – Development of a novel N-controlled Expression System for Large-Scale Production of Recombinant Proteins

For heterologous expression of recombinant proteins in large scale bioreactors, most of the commonly employed expression systems such as IPTG induction of the lac promotor suffer from limited scale-up and tunability capacities.
In bacteria capable of nitrogen fixation, the expression of nitrogenase by the nif promotor is controlled by the available nitrogen source. Diazotrophic growth with N2 leads to maximal expression whereas the presence of ammonia strongly represses the expression of nitrogenase. The nif promotor thus provides an attractive possibility as a novel expression system where the expression level of recombinant proteins could be adjusted by varying the nitrogen source in large-scale bioreactors.
In NXP-SynPath a novel expression system is developed using the nif promotor of Rhodospirillum rubrum as a novel tool for the controlled and tunable expression of recombinant proteins in response to the ambient N-source.
R. rubrum is a facultative photosynthetic non-sulfur purple bacterium with exceptional high capacities for various applications in biotechnology (Fig. 2). In particular, the formation of intracytoplasmic membranes under microaerobic conditions allows the production of compounds which require a lipophilic compartment, at levels that are not attainable with currently established production organisms (Fig. 2).
Current experiments employ a plasmid-borne fluorescent protein mCherry, controlled by the nif promotor as a model system which allows online-monitoring of the expression level by spectrofluorometry.
 
Collaboration with University Stuttgart, Institute of Biomaterials and biomolecular Systems, Department of Bioenergetics
 

Fig. 2. Biotechnological potential of R. rubrum cultivated in a bioreactor under microaerobic conditions. Insert shows the capacity for producing membrane-located carotenoids in comparison to the limited membrane compartment of E. coli.

Team

Dr. Andreas Witt
Stephan Diesch
Prof. Dr. Hartmut Grammel
 
We currently offer opportunities for conducting B.Sc. or Master theses.

Selected Publications

Hädicke, O., Pozzi, R., Witt, A., Oldiges, M., Klamt, S. and H.Grammel (2016) Systembiologische Untersuchung bakterieller Stoffwechselnetzwerke und Enzyme für die technische Verwertung von CO2. Tagungsband zum 18. Heiligenstädter Kolloquium „Technische Systeme für die Lebenswissenschaften". ISBN: 978-3-00-054165-0

Ghosh, R., Autenrieth, C., Sawodny, O., Pozzi, R., Grammel, H., Xiong, X., and R. Krastev (2016) Entwicklung neuer Technologien für Multischritt-Reaktionen in Anwesenheit von organischen Lösungsmitteln. Tagungsband zum 18. Heiligenstädter Kolloquium „Technische Systeme für die Lebenswissenschaften". ISBN: 978-3-00-054165-0

Hädicke, O., H. Grammel, and S. Klamt (2011) Metabolic network modeling of redox balancing and biohydrogen production in purple nonsulfur bacteria. BMC Syst. Biol. 5:150.

Wang, G-S., H. Grammel, K. Abou-Aisha, R. Saegesser, and R. Ghosh (2012) High-level production of the industrial product, lycopene, using the photosynthetic bacterium Rhodospirillum rubrum. Appl. Env. Microbiol.78(20):7205-15.

Grammel, H., S. Klamt, R. Ghosh, and E.D. Gilles (2000) Redox-dependent control phenomena in photosynthetic bacteria. Proc. of the First International Conference on Systems Biology (Editors T.-M. Yi, M. Hucka, M. Morohashi und H. Kitano). Tokyo 2000, pp. 148-153.