The probiotic strain Escherichia coli Nissle 1917 (EcN) with high biocompatibility and susceptibility to genetic modification is often applied in bacterial therapies for cancer. However, most studies have used plasmids as vectors to construct engineering strains from EcN. Plasmid-based expression systems suffer from genetic instability, and they need antibiotic selective pressure to maintain high copy number. This study aimed to employ EcN for synthesizing the photosensitizer 5-aminolevulinic acid (5-ALA). Firstly, the key genes of 5-ALA synthesis, hemAM and hemL, were integrated into the EcN genome by the phage integration technique. Then, chemically inducible chromosomal evolution (CIChE) was adopted to increase the copy number of hemAM and hemL and thus improved the stable synthesis of 5-ALA. The in vitro cell experiments verified that the constructed engineering strain can deliver stably synthesized 5-ALA to tumor cells and inhibit their growth. This study provided a basis for applying the engineering strains of EcN in the photodynamic therapy for tumors.
Keywords: 5-aminolevulinic acid; Escherichia coli Nissle 1917; chemically inducible chromosomal evolution (CIChE); phage integration technique.