Engineering energy metabolism boosts therapeutic protein production via the E. coli hemolysin A system

Escherichia coli currently serves as a widely used tool for therapeutic protein production; however, the high accumulation of heterologous proteins in the cytoplasm poses a risk of degradation by proteases and the formation of inactive inclusion bodies. Although it provides a direct secretion pathway, the application of the ATP-driven hemolysin A (HlyA) secretion system is severely hindered by its low efficiency.

In this work, we first developed a novel Hly62 secretion system based on Vitreoscilla hemoglobin gene expression, which significantly improved the energy metabolism of the strain. The novel system facilitates the efficient secretion of different kinds of therapeutic proteins and enables rapid detection of bioactivities using crude supernatant.

Moreover, the αTnb and αPnb-IL15 purified by one-step affinity chromatography exhibited not only potent anti-inflammatory and anti-tumor activities, but also low immunogenicity in vivo. High density cell fermentation and metabolomics analysis further validated the promotion effect of energy metabolism upregulation on the secretion efficiency of the HlyA secretion system. Therefore, our study established a rapid, simple, and efficient Hly62 secretion system based on energy metabolism engineering, revealing its significant application potential in biopharmaceutical manufacturing.