Identification, characterization, and application of a highly sensitive lactam biosensor from Pseudomonas putida

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 requires thousands of designs and results in low product titers, a highly sensitive biosensor for the final product has the potential to reduce development time. Here we report a highly sensitive biosensor for valerolactam and caprolactam from Pseudomonas putida KT2440 which is >1000x more sensitive to exogenous ligand than previously reported sensors. Manipulating the expression of the sensor oplR (PP_3516) substantially altered the sensing parameters, with various vectors showing Kd values ranging from 700 nM to 1.2 mM. Our most sensitive construct was able to detect in vivo production of caprolactam above background at 11 μg/L. The high sensitivity and range of OplR is a powerful tool towards the development of novel routes to the biological synthesis of caprolactam.