Construction of an autolytic system in Escherichia coli Nissle 1917 for efficient release of intracellular recombinant proteins.

• The cell autolytic system constructed in E. coli Nissle 1917 strain using the autolytic gene 2PaT4L can cause cell autolysis. • The production of sfGFP and TaTS after 24 h of autolysis induction was higher than that after 3 h. • The 2Pa-T4L based autolysis system of E. coli Nissle 1917 efficiently releases recombinant proteins into the culture medium, simplifying downstream protein recovery.". Escherichia coli has emerged as the preferred option in the field of recombinant protein production. However, this necessitates a cell lysis process to extract substantial quantities of recombinant protein. The E. coli Nissle 1917 (EcN) autolysis system was engineered to enhance convenience in the recovery process. This concept would simplify the procedure for retrieving recombinant proteins, streamlining the overall production process. Previous studies showed that the fusion of cell-penetrating peptides (Pa) with T4 lysozyme (T4L) to generate 2PaT4L enables E. coli to undergo autolysis, releasing intracellular proteins into the cultivation medium. In this study, the expression plasmid of 2PaT4L was constructed and transformed into ENL6P, the ECN T7 expression system host cell. The induction of this autolysis process is achieved through the induction of IPTG. After IPTG induction, the autolysis of cells gradually reached a dynamic equilibrium with the growth at a low cell concentration. After 24 h of cell autolysis induction, the yields of green fluorescent protein and trehalose synthase could reach 177.27 ± 7.40 and 84.59 ± 4.39 mg/L, respectively. The results show that the ECN autolytic expression system can overexpress recombinant proteins and simplify the recovery process. [Display omitted] [ABSTRACT FROM AUTHOR]