Enhancing proteolytic activity of Lysobacter enzymogenes using cold atmospheric plasma

Area of Science:

• Microbiology
• Biotechnology
• Plasma Science

Background:

• L. enzymogenes produces valuable proteolytic enzymes like lysyl endopeptidase (Lys C).
• Strain improvement is crucial for optimizing industrial enzyme production.
• Cold atmospheric plasma (CAP) is an emerging tool for microbial mutagenesis.

Purpose of the Study:

• To investigate the efficacy of CAP for inducing random mutagenesis in L. enzymogenes.
• To identify and characterize L. enzymogenes mutants with enhanced proteolytic activity.
• To explore the molecular basis of CAP-induced mutagenesis in the lys C gene.

Main Methods:

• L. enzymogenes (ATCC 29487) was exposed to CAP for varying durations (30-150 s).
• Mutant strains were screened for enhanced proteolytic activity using radial caseinolytic and Anson's assays.
• The lys C gene sequences of selected mutants were determined.
• Lysyl endopeptidase purification and SDS-PAGE analysis were performed.

Main Results:

• CAP treatment reduced bacterial survival with increased exposure time.
• 38 out of 100 mutants exhibited higher proteolytic activity than the wild type.
• The M1-30s mutant showed a 1.94-fold increase in proteolytic activity.
• Point mutations in the lys C gene of the M14-150s mutant correlated with enhanced protease activity.

Conclusions:

• CAP is a viable technique for random mutagenesis in L. enzymogenes.
• CAP mutagenesis can significantly enhance proteolytic enzyme production.
• This approach facilitates the development of industrial microbial strains for enzyme production.