Integrating compartmentalized catalysis with low-energy recovery via a magneto-responsive water-in-water Pickering emulsion platform for sustainable biomanufacturing of 4-methyl-5-thiazoleethanol

Abstract

Simultaneously achieving compartmentalized catalysis and low-energy recovery remains a significant challenge in conventional liquid fermentation. To address this issue, citrate-functionalized Fe3O4 nanoparticles were utilized to construct the water/water (W/W) Pickering emulsion system. A recombinant Bacillus amyloliquefaciens strain was engineered for 4-methyl-5-thiazoleethanol (MTE) biosynthesis through competing pathway knockout (ΔylmB) and site-directed mutagenesis of TenA (Y113A/F169A). In liquid fermentation, the engineered strain (ΔylmB / pHY-BaTenAY113A/F169A) was achieved a 6.80-fold increase in MTE production (from 273.67 mg/L to 1862.05 mg/L). When cultivated in the magneto-responsive W/W Pickering emulsion under optimized conditions, the MTE yield was further increased to 3061.14 mg/L, representing a 20.47% improvement over the Fe3O4 nanoparticle-free control. This enhancement was attributed to the stable compartmentalized micro-environment of the aqueous two-phase system, which promoted bacterial proliferation and sustained catalytic activity. Notably, the interfacial Fe3O4 nanoparticles enabled gentle magnetic separation, achieving supernatant clarification (OD₆₀₀ = 0.27 ± 0.04) comparable to that of high-speed centrifugation (0.20 ± 0.02). Magnetically harvested bacteria readily resumed normal growth in fresh media, and the nanoparticles could be reused for two cycles with retaining 47.80% of their saturation magnetization and supporting a sustained MTE yield of 2968.88 mg/L. A magneto-responsive aqueous two-phase system was developed to couple compartmentalized biosynthesis with low-energy bacterial recovery. This integrated platform would provide a circular and sustainable strategy for biomanufacturing high-value compounds.