The development and application of a single-cell biosensor for the detection of L-methionine and branched-chain amino acids

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 Delta aceE. 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. (C) 2012 Elsevier Inc. All rights reserved.