Periodic Reporting for period 2 - e-MICROBe (Energizing microbes with redox mediators for new bioproductions)
Okres sprawozdawczy: 2022-03-01 do 2023-08-31
In many bioprocesses a broad bioproduct portfolio can currently only be obtained when microorganisms can access oxygen as an electron acceptor. For numerous target substances, however, oxygen is detrimental to product stability and the bioprocess operation. The central aim of e-MICROBe is to innately couple microbial metabolism and electrochemistry via a self-secreted soluble electron mediator to achieve efficient oxygen independent energy metabolism and to directly steer and control metabolism and product formation. This will require creating entirely new physiological traits for production and utilization of redox mediators to generate cellular energy. Thereby, mediators can either act as electron discharge shuttle to enable electro-respiration at an anode or they are employed as inorganic energy donor to deliver electrons from a cathode into the metabolism. We will clarify the underlying reaction pathways in known environmental microorganisms and re-engineer the energy metabolism of common biotech hosts. Thereby, we will switch cellular energy generation from aerobic respiration to anaerobic anodic electro-respiration or from hydrogen consumption as autotrophic electron donor to cathodic electron consumption. The latter process will provide a mechanism to store electrical energy in microbial products. For a new level of in situ insight into microbial energy metabolism, a novel micro-scale bioelectrochemical reactor coupled to microscopic observation and high performance analysis will be developed. With this technique two fundamental concepts for future mediator-based bioprocesses will be evaluated: An all-in-one strategy where one cell is generating the mediators and the targeted product as well as a co-culture system, whereby one cell produces the mediators and a partner cell utilizes them for electro-respiration and product formation. This concept will lay the foundation for a plug-and-play exchange of biotech strains in a mediator-producing co-culture system.
WPs 1-4 are progressing well, WPs 2/3 are a bit behind schedule.
WP1: exploration of the physiology of phenazine usage and screening for new, unknown phenazine or redox mediator producers was completed; no direct energy conservation could be deciphered with phenazines in Pseudomonas-like strains. 1 Manuscript published, 1 submitted. Metabolic engineering aprroaches were redirected to synthetic biology concepts and new strains are currently being tested.
WP2: microBES reactor development, involves in-kind contribution not on premises by Fraunhofer IOF; We experienced slight delays for WP2: The collaboration with Fraunhofer IOF for the development of the microBES reactor has been delayed by ~ 6 month due to Institute closures (Corona-related), quarantines of involved people and delivery delays for parts. However, overall the development is progressing well and functional microBES chips are now ready to be used in WP3.
WP3: With function microBES from WP2 experiments with online cultivation of microbial cultures have started. The growth can be observed and the electrochemical measurements are functional. Currently, the tuning of the right operating conditions in the focus before defined hypotheses will be tested.
WP4: biological synthesis and evaluation of a redox negative soluble redox mediator: manuscript in preparation. A proof of concent for cathodic electron uptake with this mediator was achieved. No strains that include a sink for these cathodic electrons are established to create an outlet and a true application example. Bioelectrochemical characterization of this compound is ongoing.
WP1: exploration of the physiology of phenazine usage and screening for new, unknown phenazine or redox mediator producers was completed; no direct energy conservation could be deciphered with phenazines in Pseudomonas-like strains. 1 Manuscript published, 1 submitted. Metabolic engineering aprroaches were redirected to synthetic biology concepts and new strains are currently being tested.
WP2: microBES reactor development, involves in-kind contribution not on premises by Fraunhofer IOF; We experienced slight delays for WP2: The collaboration with Fraunhofer IOF for the development of the microBES reactor has been delayed by ~ 6 month due to Institute closures (Corona-related), quarantines of involved people and delivery delays for parts. However, overall the development is progressing well and functional microBES chips are now ready to be used in WP3.
WP3: With function microBES from WP2 experiments with online cultivation of microbial cultures have started. The growth can be observed and the electrochemical measurements are functional. Currently, the tuning of the right operating conditions in the focus before defined hypotheses will be tested.
WP4: biological synthesis and evaluation of a redox negative soluble redox mediator: manuscript in preparation. A proof of concent for cathodic electron uptake with this mediator was achieved. No strains that include a sink for these cathodic electrons are established to create an outlet and a true application example. Bioelectrochemical characterization of this compound is ongoing.
All project parts perform research beyond the state of the art and are in wide parts high risk - high gain. We already obtained new insights in the physiology of phenazine redox mediators and reviewed different strategies for integrating bioelectrochemistry with biotechnology (see publications). A first publication on the identification of new redox mediator producing microorganisms was published, a second on phenazine interaction with the energy metabolism is currently submitted. New synthetic biology routes are explored to link phenazines better into energy metabolism. Our new cathodic mediator is the first of its kind and will open a completely new strategy in bioelectrochemistry.