PUB - Publikationen an der Universität Bielefeld

The detection and quantification of specific metabolites in single bacterial cells is a major goal for industrial biotechnology. We have developed a biosensor based on the transcriptional regulator Lrp that detects intracellular l-methionine and branched-chain amino acids in Corynebacterium glutamicum. In assays, fluorescence output showed a linear relationship with cytoplasmic concentrations of the effector amino acids. In increasing order, the affinity of Lrp for the amino acids is l-valine, l-isoleucine, l-leucine and l-methionine. The sensor was applied for online monitoring and analysis of cell-to-cell variability of l-valine production by the pyruvate dehydrogenase-deficient C. glutamicum strain ΔaceE. Finally, the sensor system was successfully used in a high-throughput (HT) FACS screen for the isolation of amino acid-producing mutants after random mutagenesis of a non-producing wild type strain. These applications illustrate how one of nature's sensor devices – transcriptional regulators – can be used for the analysis, directed evolution and HT screening for microbial strain development. Highlights ► Development of a single cell biosensor for the intracellular detection of amino acids. ► Biosensor-based online monitoring of valine production of a C. glutamicum strain. ► Visualization of cell-to-cell variability of microbial production strains. ► Establishment of a FACS high-throughput screening.