Automated screening system for rational selection and characterization of most efficient biocatalysts and process parameters.
Reactions catalyzed by enzymes obtain an increasing relevance in the field of chemical engineering. Parallel modern molecular biological methods like “Directed Evolution” generate a huge number of recombinant enzymes. Therefore, new high throughput methods for the screening of biocatalysts are required. Such techniques afford the development of an optimized process considering the number of potentially interesting enzymes as well as a wide range of process parameters.
As a consequence, a system was developed combining an on-line monitoring system for cultivations and reactions in microtiter plates (BioLector) with an automated liquid handling station. This combination allows automated high-throughput experimentation with extensive output of relevant process parameters (biomass, product formation, DOT, pH,…). The interaction of both systems offers lots of possibilities to realize different protocols:
- Fully automated sampling and sample storage
- Adding of compounds (e.g. inducers, substrates, pH controlling agents)
- Downstream processes (biomass separation, enzyme extraction, enzyme purification)
- Assays for enzyme characterization
In addition, great efforts were made to characterize different types of microtiter plates regarding their process relevant technical parameters so that they can finally be used as defined reaction systems. As a consequence, modifications were performed to improve particular properties, e.g. the oxygen transfer.
This project is part of the Cluster of Excellence "Tailor-Made Fuels from Biomass". The excellence cluster adopts an interdisciplinary approach through the application of optimized synthesis processes towards research of new, biomass-based, synthetic fuels. The goal is to verify their potential, with regard to modern combustion technologies, while simultaneously reducing the reliance on fossil fuels. The interdisciplinary nature of this approach is the key to a holistic and sustainable solution for future vehicular traffic, which is a basic necessity for a modern society.
This project is concerned with the development of an automated high-throughput system for characterizing efficient biocatalysts and process conditions. For the excellence cluster “Tailor-Made Fuels from Biomass” the project is focusing on the enzymatic degradation of cellulosic biomass to sugars for fermentation to biofuels. Therefore, both the optimal expression system for cellulolytic enzymes (microorganism, culture conditions) and the hydrolyzing step (enzyme, solvent) are under investigation. On-line measurement techniques, such as light scattering or fluorescence, are used for determining different parameters to evaluate growth and expression kinetics as well as activity and long term stability of the enzymes.
Samorski M, Muller-Newen G, Büchs J. 2005. Quasi-continuous combined scattered light and fluorescence measurements: A novel measurement technique for shaken microtiter plates. Biotechnology and Bioengineering 92(1):61-68.
Huber R, Ritter D, Hering T, Hillmer AK, Kensy F, Muller C, Wang L, Büchs J. 2009. Robo-Lector - a novel platform for automated high-throughput cultivations in microtiter plates with high information content. Microbial Cell Factories 8:42.
Hermann R, Lehmann M, Büchs J. 2003. Characterization of gas-liquid mass transfer phenomena in microtiter plates. Biotechnology and Bioengineering 81(2):178-186.
Funke M, Diederichs S, Kensy F, Müller C, Büchs J. 2009b. The Baffled Microtiter Plate: Increased Oxygen Transfer and Improved Online Monitoring in Small Scale Fermentations. Biotechnology and Bioengineering 103(6):1118-1128.
Kensy F, Engelbrecht C, Büchs J. 2009a. Scale-up from microtiter plate to laboratory fermenter: evaluation by online monitoring techniques of growth and protein expression in Escherichia coli and Hansenula polymorpha fermentations. Microbial Cell Factories 8:68.
Kensy F, Zang E, Faulhammer C, Tan RK, Büchs J. 2009b. Validation of a high-throughput fermentation system based on online monitoring of biomass and fluorescence in continuously shaken microtiter plates. Microbial Cell Factories 8:31
Rachinskiy K, Schultze H, Boy M, Bornscheuer U, Büchs J. 2009. "Enzyme Test Bench," a High-Throughput Enzyme Characterization Technique Including the Long-Term Stability. Biotechnology and Bioengineering 103(2):305-322.