UV-He-Ne Laser Sequential Mutagenesis Improves Protease Production of Bacillus velezensis: Comparison with Traditional UV Mutagenesis.

This study combined the damage effect of UV and the repair effect of He-Ne laser, employing UV-He-Ne laser (UV-LA) sequential mutagenesis to improve the protease-producing ability of Bacillus velezensis. Compared with UV mutagenesis, UV-LA treatment elevated the positive mutation rate from 12.50% to 16.67%, and the enzyme activity of the mutant strain increased from 20.81 U/mL to 34.04 U/mL. Following 20 passages, the degree of enzyme production decline of the UV-LA strain was reduced from 44% to 23% compared with that of the UV strain, showing better genetic stability. Subsequent investigation revealed that the physiological activity (L-LDH, ATP content) and the activities of various key metabolic enzymes (PFK, IDH, ATPase) in the UV-LA strain exceeded those in the UV strain. Genome sequencing results showed that the UV-LA strain had fewer mutations than the UV strain, and the mutation sites were more concentrated in the regulatory region. Proteomic analysis identified 108 differentially expressed proteins (56 upregulated and 52 downregulated) between the UV strain and UV-LA strain, primarily associated with bacterial energy metabolism, translation, and secretion systems. Research indicates that UV-LA treatment may leverage the laser's light effects to reduce UV damage, thereby achieving regulation of metabolic and secretory processes. This study is expected to develop a novel and efficient mutagenesis method for microbial breeding and to offer new ideas for expanding the application of UV mutagenesis technology.