generic-protocol-for-optimization-heterologous-protein-production

Generic Protocol for Optimization of Heterologous Protein Production Using Automated Microbioreactor Technology

A core business in industrial biotechnology using microbial production cell factories is the iterative process of strain engineering and optimization of ...

Forschungszentrum Jülich

Bioeconomy Science Center (BioSC)

Rhamnolipids are potent biosurfactants with high potential for industrial applications. However, rhamnolipids are currently produced with the opportunistic pathogen Pseudomonas aeruginosa during growth on hydrophobic substrates such as plant oils. The heterologous production of rhamnolipids entails two essential advantages: Disconnecting the rhamnolipid biosynthesis from the complex quorum sensing regulation and the opportunity of avoiding pathogenic production strains, in particular P. aeruginosa. In addition, separation of rhamnolipids from fatty acids is difficult and hence costly.

Growth independent rhamnolipid production from glucose using the non-pathogenic Pseudomonas putida KT2440.

Technical bulk enzymes represent a huge market, and the extracellular production of such enzymes is favorable due to lowered cost for product recovery. Protein secretion can be achieved via general secretion (Sec) pathway. Specific sequences, signal peptides (SPs), are necessary to direct the target protein into the translocation machinery. For example, >150 Sec-specific SPs have been identified for Bacillus subtilis alone. As the best SP for a target protein of choice cannot be predicted a priori, screening of homologous SPs has been shown to be a powerful tool for different expression organisms. While SP libraries between closely related species were successfully applied to optimize recombinant protein secretion, this was not investigated for distantly related species. Therefore, in this study a Sec SP library from low-GC firmicutes B. subtilis is investigated to optimize protein secretion in high-GC actinobacterium Corynebacterium glutamicum using cutinase from Fusarium solani pisi as model protein.

Use of a Sec signal peptide library from Bacillus subtilis for the optimization of cutinase secretion in Corynebacterium glutamicum.

The calculation of growth rates provides basic metric for biological fitness and is standard task when using microbioreactors (MBRs) in microbial phenotyping. MBRs easily produce huge data at high frequency from parallelized high-throughput cultivations with online monitoring of biomass formation at high temporal resolution. Resulting high-density data need to be processed efficiently to accelerate experimental throughput.

Article	Year
Growth independent rhamnolipid production from glucose using the non-pathogenic Pseudomonas putida KT2440. Microbial cell factories\| PubMed ID: 21999513	2011
Use of a Sec signal peptide library from Bacillus subtilis for the optimization of cutinase secretion in Corynebacterium glutamicum. Microbial cell factories\| PubMed ID: 27927208	2016
Automated growth rate determination in high-throughput microbioreactor systems. BMC research notes\| PubMed ID: 29178966	2017

Johannes Hemmerich

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