Genome mining of Bacillus licheniformis MCC2514 for the identification of lasso peptide biosynthetic gene cluster and its characterization

The probiotic strain Bacillus licheniformis MCC2514 has been shown to produce a strong antibacterial peptide and the whole genome sequence of this strain is also reported in our previous study. The present study is focused on the genome level investigation of this peptide antibiotic and its characterization. Genome mining of the culture revealed the presence of three putative bacteriocin clusters, viz. lichenicidin, sonorensin and lasso peptide. Hence, the mode of action of the peptide was investigated by reporter assay, scanning electron microscopy, and Fourier Transform Infrared spectroscopy. Additionally, the peptide treated groups of Kocuria rhizophila showed a reduction in the fold expression for transcription-related genes. The gene expression studies, quantitative beta-galactosidase induction assay using the RNA stress reporter strain, yvgS along with the homology studies concluded that lasso peptide is responsible for the antibacterial activity of the peptide which acts as an inhibitor of RNA biosynthesis. Gene expression analysis showed a considerable increase in fold expression of lasso peptide genes at various fermentation hours. Also, the peptide was isolated, and its time-kill kinetics and minimum inhibitory concentration against the indicator pathogen K. rhizophila were examined. The peptide was also purified and the molecular weight was determined to be similar to 2 kDa. Our study suggests that this bacteriocin can function as an effective antibacterial agent in food products as well as in therapeutics as it contains lasso peptide, which inhibits the RNA biosynthesis.