Performance of immobilized recombinant Escherichia coli on β‐cyclodextrin glucanotransferase secretion and cell viability in a membrane bioreactor

BACKGROUND The efficiency of recombinant enzyme secretion into the medium is essential for simplifying purification and increasing yield. However, Escherichia coli , a commonly used expression host, faces challenges such as cell lysis and plasmid instability during enzyme secretion. Cell immobilization is proposed to address this problem. This study aimed to investigate the performance of immobilized recombinant E. coli focusing on β ‐cyclodextrin glucanotransferase ( β ‐CGTase) secretion, cell lysis and plasmid stability in a membrane bioreactor. The study compared two immobilization configurations, IC‐9 (9 hollow‐fiber membranes) and IC‐19 (19 hollow‐fiber membranes), as well as a shake flask system. RESULTS IC‐9 exhibited superior performance, achieving 1.5‐fold higher β ‐CGTase activity than IC‐19. Additionally, IC‐9 showed 87% lower cell lysis and 12% higher plasmid stability compared to IC‐19, indicating improved process reliability. The immobilized cells in the membrane bioreactor exhibited 2.5‐fold higher β ‐CGTase secretion, 75% lower cell lysis and 29% higher plasmid stability compared to the shake flask system. Moreover, β ‐CGTase secretion by immobilized cells was twice as high as that of free cells and remained stable over seven cycles, achieving a cumulative β ‐CGTase activity of 1120.21 U mL −1 . Kinetic analysis showed that immobilized cells exhibited a lower maximum specific growth rate (0.012 h − 1 ) compared to free cells (0.149 h − 1 ), but significantly higher β ‐CGTase secretion, with an α (growth‐associated production coefficient) of 1.317 U mg −1 mL −1 compared to 0.004 U mg −1 mL −1 for free cells. CONCLUSION These results highlight the potential of membrane bioreactors for large‐scale recombinant enzyme production with improved efficiency and operational stability.