Antibacterial activity of an anti-lipopolysaccharide factor (MjALF-D) identified from kuruma prawn (Marsupenaeus japonicus)

Machine learning-guided directed evolution for the development of small-molecule antibiotics originating from antimicrobial peptides

“…LBD B showed higher antimicrobial activity than LBD A, C-D , LBD X and LBD Y , and the highest activity (< 2.5 μM) from LBD B was observed for the gram-negative bacterium E. coli, Shigella flexnerib, Aeromonas hydrophilac and the gram-positive bacteria B. subtilis and C. glutamicum. Again, in our previous study( 36 ), 40 μM MIC was chosen as a threshold, and any values higher than that are considered not active; thus, no further experiments regarding bioassays were continued. Overall, the antibacterial spectrum of LBD A-D was broadened compared to that of LBD Mj , and more interestingly, LBD A-D did not develop any resistance to all strains tested in this study, regardless of whether they were tested at either sub-MIC or sur-MIC levels, at least under our laboratory conditions.…”

“…The antibacterial spectrum detection assay was performed as described previously( 36, 67, 68 ). We selected 6 Gram-negative bacteria ( E. coli, Pseudomonas fluorescens, P. aeruginosa, S. flexneri, A. hydrophila and V. alginolyticus) and 3 Gram-positive bacteria ( B. subtilis, S. aureus, C. glutamicum) for the liquid growth inhibition assay.…”

“…Antibacterial test. The antibacterial spectrum detection assay was performed as described previously (36,67,68). We selected 6 Gram-negative bacteria (E. coli, Pseudomonas fluorescens, P. aeruginosa, S. Logarithmic phase cultures of E. coli and S. aureus were incubated with LBDB as described above, and LBDB was replaced with sterile Milli-Q water as a control.…”

“…Recent studies have shown that structurally modified LBDs can improve bioactivities and thus provide a promising reservoir for new antimicrobial agent design or optimization ( 33 ),( 34 ),( 35 ) . We have recently reported the ALF family AMP (ALF Mj ) isolated from the kuruma prawn Marsupenaeus japonicus , which shows activity against Photobacterium damselae and Micrococcus luteus ( 36 ) however, its corresponding LBD Mj did not generally exhibit activities when the experiments were performed in parallel, leaving a chance for further structural modification.…”

“…Then, a 2-fold serial dilution of LBDs with final concentrations ranging from 1.25 to 40 μM was incubated with the microbes at 28°C for 24 h. Dose-dependent antibacterial activity and time-killing curves. Kinetic studies were performed to further determine the bactericidal rate of LBDB according to previous procedures(36,69,70), and we chose E. coli and S. aureus as representative gram-negative and gram-positive bacteria, respectively.…”