Valerolactam is a monomer used to manufacture high-value nylon-5 and nylon-6,5. However, the biological production of valerolactam has been limited by the inadequate efficiency of enzymes to cyclize 5-aminovaleric acid to produce valerolactam. In this study, we engineered Corynebacterium glutamicum with a valerolactam biosynthetic pathway consisting of DavAB from Pseudomonas putida to convert L-lysine to 5-aminovaleric acid and β-alanine CoA transferase (Act) from Clostridium propionicum to produce valerolactam from 5-aminovaleric acid. Most of the L-lysine was converted into 5-aminovaleric acid, but promoter optimization and increasing the copy number of Act were insufficient to significantly improve the titer of valerolactam. To eliminate the bottleneck at Act, we designed a dynamic upregulation system (a positive feedback loop based on the valerolactam biosensor ChnR/Pb). We used laboratory evolution to engineer ChnR/Pb to have higher sensitivity and a higher dynamic output range, and the engineered ChnR-B1/Pb-E1 system was used to overexpress the rate-limiting enzymes (Act/ORF26/CaiC) that cyclize 5-aminovaleric acid into valerolactam. In glucose fed-batch culture, we obtained 12.33 g/L valerolactam from the dynamic upregulation of Act, 11.88 g/L using ORF26, and 12.15 g/L using CaiC. Our engineered biosensor (ChnR-B1/Pb-E1 system) was also sensitive to 0.01-100 mM caprolactam, which suggests that this dynamic upregulation system can be used to enhance caprolactam biosynthesis in the future.

The increase in population-related and environmental issues has emphasized the need for more efficient and sustainable production strategies for foods and chemicals. Carbohydrates are macronutrients sourced from crops and undergone transformation into various products ranging from foods to chemicals. Continuous efforts have led to the identification of a promising hybrid system that couples the electrochemical reduction of CO2 to intermediates containing one to three carbons (C1–3) with the transformation of the intermediates using engineered microorganisms into valuable products. Here we use yeast to transform C1–3 substrates into glucose and structurally tailored glucose derivatives, such as the sugar alcohol myo-inositol, the amino monosaccharide glucosamine, the disaccharide sucrose and the polysaccharide starch. By metabolic rewiring and mitigation of glucose repression, the titre of glucose and sucrose reached dozens of grams per litre. These results provide directions for microbial sugar-derived foods and chemicals production from renewable reduced CO2-based feedstocks. [Figure not available: see fulltext.].