Abstract
The bacteriocin microcin J25 (MccJ25) inhibits the growth of Gram-negative pathogens including Salmonella and Shigella species, and Escherichia coli. This 21-amino acid peptide has remarkable stability to heat and extreme pH values and resistance to many proteases, thanks to a characteristic lasso structure. In this study, we used the dynamic simulator TIM-1 as gastro-intestinal tract model to evaluate the stability and antibacterial activity of MccJ25 during passage through the proximal portion of the human gastrointestinal tract. MccJ25 concentration was measured in the different simulator sections by HPLC, and inhibition of Salmonella enterica serotype Enteritidis was evaluated using qualitative and quantitative assays. LC-MS/MS analysis and subsequent molecular networking analysis on the Global Natural Product Social Molecular Networking platform (GNPS) and analysis of the peptide degradation in the presence of proteolytic enzymes mimicking the gastro-intestinal conditions permitted to delineate the fate of MccJ25 through identification of the main degradation products. MccJ25 was relatively stable under gastric conditions, but degraded rapidly in the compartment mimicking the duodenum, notably in the presence of pancreatin. Among pancreatin components, elastase I appeared primarily responsible for MccJ25 breakdown, while α-chymotrypsin was less efficient.