Allosteric-Regulation-Based DNA Circuits in Saccharomyces cerevisiae to Detect Organic Acids and Monitor Hydrocarbon Metabolism In Vitro.
Michael Dare AsemoloyeMario Andrea MarchisioPublished in: Methods in molecular biology (Clifton, N.J.) (2024)
We show the engineering of prokaryotic-transcription-factor-based biosensing devices in Saccharomyces cerevisiae cells for an in vitro detection of common hydrocarbon intermediates/metabolites and potentially, for monitoring of the metabolism of carbon compounds. We employed the bacterial receptor proteins MarR (multiple antibiotic-resistant receptor) and PdhR (pyruvate dehydrogenase-complex regulator) to detect benzoate/salicylate and pyruvate, respectively. The yeast-enhanced green fluorescence protein (yEGFP) was adopted as an output signal. Indeed, the engineered yeast strains showed a strong and dynamic fluorescent output signal in the presence of the input chemicals ranging from 2 fM up to 5 mM. In addition, we describe how to make use of these strains to assess over time the metabolism of complex hydrocarbon compounds due to the hydrocarbon-degrading fungus Trichoderma harzianum (KY488463).
Keyphrases
- saccharomyces cerevisiae
- transcription factor
- escherichia coli
- label free
- single molecule
- induced apoptosis
- small molecule
- cell cycle arrest
- ms ms
- quantum dots
- protein protein
- cell death
- dna binding
- signaling pathway
- circulating tumor
- amino acid
- loop mediated isothermal amplification
- real time pcr
- circulating tumor cells
- energy transfer
- cell wall