Enhancing the Production of ε-Poly-l-Lysine by Engineering the Sucrose Metabolism Pathway in PD-1 Using Cane Molasses.

Cane molasses, a sugar-rich agro-industrial byproduct, was used to enhance the production of ε-poly-l-lysine (ε-PL) in PD-1 as a cost-effective carbon source. The sucrose metabolism pathway was engineered by heterologously expressing sucrose-6-phosphate hydrolase from W. The optimization of various promoters identified the SP44 promoter, increasing the total sugar utilization rate by 2.76-fold compared with the promoter. Additionally, adaptive laboratory evolution improved the total sugar utilization rate. The evolved strain achieved an ε-PL titer of 2.65 ± 0.15 g/L in flask experiments, increasing the ε-PL titer by 7.16-fold compared with the unevolved strain. Comparative transcriptomic analyses revealed that the enhanced tolerance of the evolved strain to high concentrations of cane molasses was primarily due to modifications in the sucrose metabolism pathways, microbial metabolism in heavy metals and phenols, and the amino acids transport and metabolic pathways. These changes enabled more efficient ε-PL production. During fed-batch fermentation in a 5-L fermentor using a concentration of 50 g/L cane molasses, the ε-PL titer reached 36.88 ± 0.62 g/L, and dry cell weight was 41.1 ± 1.0 g/L. This study illustrates that cane molasses is an economical carbon source for producing ε-PL on an industrial scale.