Hybrid metabolic engineering enables xylose-driven co-production of polyhydroxybutyrate and violacein in Escherichia coli

Within lignocellulosic biomass, xylose is the second most abundant sugar after glucose. As a renewable and sustainable substrate, it is gaining attention as a feedstock for microbial bioprocesses. In this study, we demonstrated the co-production of polyhydroxybutyrate (PHB) and violacein from xylose. We initially confirmed the feasibility of co-production through genome-scale metabolic simulations, followed by optimization using a hybrid expression system that combines a conventional tac promoter and synthetic promoter-ribosome-binding site-terminator (semi-endo PRT) elements in dual plasmids. Additionally, we assessed the antimicrobial activity of violacein against type I methanotrophs. Recombinant Escherichia coli DH5α harboring a hybrid system produced 111.3 ± 19.7 and 0.88 ± 0.23 mg/L of PHB and violacein, respectively, in M9 using xylose as the sole carbon source, without tryptophan supplementation. Using the synthetic PRT system, 528.9 ± 104 mg/g dry cell weight (DCW) of PHB was obtained. Additionally, violacein inhibits the growth of Methylomicrobium alcaliphilum 20Z at 9 µg/mL in nitrate mineral salt medium containing methanol as the sole carbon source. The use of lignocellulose-derived sugars for co-production offers an environmentally sustainable bio-manufacturing approach that contributes to greenhouse gas mitigation and supports the transition toward a circular bioeconomy.