Genetic regulatory element based whole-cell biosensors for the detection of metabolic disorders

Clinicians require simple, and cost-effective diagnostic tools for the quantitative determination of amino acids in physiological fluids for the detection of metabolic disorder diseases. Besides, amino acids also act as biological markers for different types of cancers and cardiovascular diseases. Herein, we applied an in-silico based approach to identify potential amino acid-responsive genetic regulatory elements for the detection of metabolic disorders in humans. Identified sequences were further transcriptionally fused with GFP, thus generating an optical readout in response to their cognate targets. Screening of genetic regulatory elements led us to discover two promoter elements (pmetE::GFP and ptrpL::GFP) that showed a significant change in the fluorescence response to homocysteine and tryptophan, respectively. The developed biosensors respond specifically and sensitively with a limit of detection of 3.8 mu M and 3 mu M for homocysteine and tryptophan, respectively. Furthermore, the clinical utility of this assay was demonstrated by employing it to identify homocystinuria and tryptophanuria diseases through the quantification of homocysteine and tryptophan in plasma and urine samples within 5 h. The precision and accuracy of the biosensors for disease diagnosis were well within an acceptable range. The general strategy used in this system can be expanded to screen different genetic regulatory elements present in other gram-negative and gram-positive bacteria for the detection of metabolic disorders.

Automated summary

In this study, potential amino acid-responsive genetic regulatory elements were identified using in-silico approach, and fused with GFP to generate an optical readout. The developed biosensors showed specific and sensitive detection of homocysteine and tryptophan, allowing for the diagnosis of homocystinuria and tryptophanuria diseases.