Purification, Structural Characterization, and Genomic Insights of an Anti-MRSA Peptide from Mangrove-Derived Brevibacillus sp. A34

The widespread prevalence of methicillin-resistant Staphylococcus aureus (MRSA) in community and healthcare settings has driven the urgent search for novel antibacterial agents. Mangrove sediments are an underexplored source of antimicrobial-producing bacteria. This study characterized an antimicrobial fraction and the genome of the mangrove-derived isolate Brevibacillus sp. A34. The active fraction was purified via ammonium sulfate precipitation, cation-exchange chromatography, and reversed-phase chromatography. De novo MS/MS analysis assigned the octapeptide LLLLLLAH (905.19 Da), designated as A34 AMP, which was further confirmed by chemical synthesis. This active peptide inhibited all four tested S. aureus strains, including three MRSA strains, with a minimum inhibitory concentration (MIC) of 62 µg/mL. At 2× and 4× MIC, A34 AMP exhibited a concentration-dependent bactericidal effect within 20 h. Increased SYTOX Green uptake and morphological alterations supported a membrane permeabilization mechanism, and antimicrobial activity was retained following thermal, pH, protease, and surfactant treatments. Genome mining identified 23 putative biosynthetic gene clusters, including an 11-module nonribosomal peptide synthetase (NRPS) cluster linked to A34 AMP through three-dimensional adenylation-domain modeling. Collectively, these findings highlight A34 AMP from Brevibacillus sp. A34 as a promising bioactive anti-staphylococcal peptide scaffold.