Novel modifications of nonribosomal peptides from Brevibacillus laterosporus MG64 and investigation of their mode of action.

Nonribosomal peptides (NRPs) are a class of secondary metabolites usually produced by microorganisms. They are of paramount importance in different applications including biocontrol and pharmacy. Brevibacillus spp. is a rich source of NRPs yet has received little attention. In this study, we characterize four novel bogorol variants (bogorol I-L, cationic linear lipopeptides) and four succilins (succilin I-L, containing a succinyl group that is attached to the Orn3/Lys3 in bogorol I-L) from the biocontrol strain B. laterosporus MG64. Further investigation revealed that the bogorol family of peptides employ an adenylation pathway for lipoinitiation, which is different from the usual pattern that is based on an external ligase and coenzyme A. Moreover, the formation of valinol was proven to be mediated by a terminal reductase domain and a reductase encoded by gene bogI Furthermore, succinylation, which is a novel type of modification in the family of bogorols, was discovered. Its occurrence requires a high concentration of substrate (bogorols) but its responsible enzyme remains unknown. Bogorols display potent activity against both Gram-positive and Gram-negative bacteria. Investigation of their mode of action reveals that bogorols form pores in the cell membrane of both Gram-positive and Gram-negative bacteria. A combination of bogorols and relacidines, another class of NRPs produced by B. laterosporus MG64, displays a synergistic effect on different pathogens, suggesting great potential of both peptides as well as their producer B. laterosporus MG64 for broad applications. Our study provides a further understanding of the bogorol family of peptides as well as their applications.Importance NRPs form a class of secondary metabolites with biocontrol and pharmaceutical potential. This work describes the identification of novel bogorol variants and succinylated bogorols (namely succilins) and further investigate their biosynthetic pathway and mode of action. Adenylation domain-mediated lipoinitiation of bogorols represents a novel pathway of NRPs to incorporate fatty acid tail. This pathway provides the possibility to engineer the lipid tail of NRPs without identifying a fatty acid coenzyme ligase, which is usually not present in the biosynthetic gene cluster. The terminal reductase domain (TD) and BogI-mediated valinol formation and its effect on the biological activity of bogorols are revealed. Succinylation, which is barely reported in NRPs, is discovered in the bogorol family of peptides. We demonstrate that bogorols combat bacterial pathogens by forming pores in the cell membrane. We also report synergistic effect of two natural products (relacidine B and bogorol K) produced by the same strain, which is relevant for competition for a niche.