Journal
ACS SYNTHETIC BIOLOGY
Volume 9, Issue 1, Pages 53-62Publisher
AMER CHEMICAL SOC
DOI: 10.1021/acssynbio.9b00292
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Funding
- U.S. Department of Energy, Office of Science, Office of Biological and Environmental Research
- U.S. Department of Energy, Energy Efficiency and Renewable Energy, Bioenergy Technologies Office [DE-AC02-0SCH11231]
- Koret Research Scholars Program
- Basque Government through the BERC 2018-2021 program
- Spanish Ministry of Economy and Competitiveness MINECO: BCAM Severo Ochoa excellence accreditation [SEV-2017-0718]
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Caprolactam is an important polymer precursor to nylon traditionally derived from petroleum and produced on a scale of 5 million tons per year. Current biological pathways for the production of caprolactam are inefficient with titers not exceeding 2 mg/L, necessitating novel pathways for its production. As development of novel metabolic routes often require thousands of designs and result in low product titers, a highly sensitive biosensor for the final product has the potential to rapidly speed up development times. Here we report a highly sensitive biosensor for valerolactam and caprolactam from Pseudomonas putida KT2440 which is >1000X more sensitive to an exogenous ligand than previously reported sensors. Manipulating the expression of the sensor oplR (PP_3516) substantially altered the sensing parameters, with various vectors showing K-d values ranging from 700 nM (79.1 mu g/L) to 1.2 mM (135.6 mg/L). Our most sensitive construct was able to detect in vivo production of caprolactam above background at similar to 6 mu g/L. The high sensitivity and range of OplR is a powerful tool toward the development of novel routes to the biological synthesis of caprolactam.
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