The pET system is commonly used for producing foreign proteins in Escherichia coli, but its reliance on the costly and metabolically demanding inducer IPTG limits its industrial use. This study engineered a low-temperature inducible system (LTIS) in E. coli Nissle 1917 (EcN) by combining the T7 expression system with the thermal inducible mechanism CI857-λPRPL to generate the new LTIS strain, ENL7P. The strain ENL7P-sfGFP-Km underwent overnight culture at 37 °C for 14-16 h, followed by subculturing at 30 °C for 24 h. This resulted in a notable 5.53-fold increase in the sfGFP induction rate when the strain was cultivated under 37-30 °C conditions. Moreover, gene expression was induced using a two-stage strategy. Initially, the strain was cultured overnight at 39 °C for 14-16 h, followed by a subculture at 30 °C for 6 h, and finally, another subculture at 30 °C for 24 h. This cultivation strategy led to an impressive 158.37-fold induction rate for sfGFP. Similar effects could be achieved through utilization of the LTIS system for inducing the production of thermophilic trehalose synthase from Thermus antranikianii (TaTS). The results of this study proved that the LTIS system has the potential for industrial applications.
Keywords: CI857-λPRPL system; Escherichia coli; LacI; Nissle 1917; T7 expression system; low-temperature inducible system.