Genetically encoded sensors enable real-time observation of metabolite production
Here, engineering cells to produce valuable metabolic products is hindered by the slow and laborious methods available for evaluating product concentration. Consequently, many designs go unevaluated, and the dynamics of product formation over time go unobserved. In this work, we develop a framework for observing product formation in real time without the need for sample preparation or laborious analytical methods. We use genetically encoded biosensors derived from small-molecule responsive transcription factors to provide a fluorescent readout that is proportional to the intracellular concentration of a target metabolite. Combining an appropriate biosensor with cells designed to produce a metabolic product allows us to track product formation by observing fluorescence. With individual cells exhibiting fluorescent intensities proportional to the amount of metabolite they produce, high-throughput methods can be used to rank the quality of genetic variants or production conditions. We observe production of several renewable plastic precursors with fluorescent readouts and demonstrate that higher fluorescence is indeed an indicator of higher product titer. Using fluorescence as a guide, we identify process parameters that produce 3-hydroxypropionate at 4.2 g/L, 23-fold higher than previously reported. We also report, to our knowledge, the first engineered route from glucose to acrylate, a plastic precursor with globalmore »
- Publication Date:
- Grant/Contract Number:
- FG02-02ER63445
- Type:
- Published Article
- Journal Name:
- Proceedings of the National Academy of Sciences of the United States of America
- Additional Journal Information:
- Journal Volume: 113; Journal Issue: 9; Journal ID: ISSN 0027-8424
- Publisher:
- National Academy of Sciences, Washington, DC (United States)
- Research Org:
- Harvard Univ., Boston, MA (United States)
- Sponsoring Org:
- USDOE Office of Science (SC), Biological and Environmental Research (BER) (SC-23)
- Country of Publication:
- United States
- Language:
- English
- Subject:
- 59 BASIC BIOLOGICAL SCIENCES; 60 APPLIED LIFE SCIENCES; biotechnology; directed evolution; biosensor; metabolic engineering; synthetic biology
- OSTI Identifier:
- 1237386
- Alternate Identifier(s):
- OSTI ID: 1348838