Engineering Corynebacterium glutamicum for 1,5-pentanediamine production via enhanced product tolerance and efflux...

1,5-Pentanediamine (PDA) is a vital monomer with broad applications in textiles, medicine, and agriculture. Nevertheless, its microbial biosynthesis is severely restricted by cytotoxicity, primarily resulting from rapid intracellular accumulation during fermentation. To address this challenge, a dual tolerance engineering strategy was employed in Corynebacterium glutamicum to alleviate both intracellular and extracellular stress. Membrane-associated tolerance targets were first identified through rational screening, and adaptive laboratory evolution (ALE) uncovered a novel determinant, cgl1283, which markedly improved extracellular PDA tolerance. In parallel, a high-activity CgmA efflux transporter mutant was engineered to minimize intracellular PDA accumulation, thereby mitigating cytotoxic effects. Subsequent process optimization in a 5-L bioreactor enabled strain LF6.1 to produce 130 g/L PDA from glucose, with a yield of 0.40 g/g and a productivity of 3.94 g/L/h. Collectively, these results establish a comprehensive framework that integrates intracellular and extracellular tolerance engineering, offering an effective strategy for the microbial production of toxic yet high-value chemicals.