The potency of the broad spectrum bacteriocin, bactofencin A, against staphylococci is highly dependent on primary structure, N-terminal charge and disulphide formation
O'Connor, Paula M.
O'Shea, Eileen F.
Cotter, Paul D.
Ross, R. Paul
Bactofencin A is a novel class IId bacteriocin, produced by the intestinal isolate Lactobacillus salivarius DPC6502, which has potent activity against medically significant pathogens including Staphylococcus aureus. This bacteriocin is unusual in that it has a highly cationic N terminus and a single disulfide bond between Cys7 and Cys22, resulting in a large C terminal loop. In this study, a library of synthetic bactofencin A variants were screened against the mastitis isolate, S. aureus DPC5246, to identify key residues responsible for activity. It was apparent that substituting either cysteine of the disulfide bond with either serine or alanine significantly reduced the activity of the bacteriocin, confirming the importance of the C terminal loop. Substituting N terminal amino acids with alanine had no effect on activity, whereas sequential removal of the N terminal positively charged residues resulted in an increasingly inactive peptide. A complete (synthetic) alanine scanning analysis revealed that the residues between Val9 and Gly17 were most affected by substitution suggesting that this area has a major influence on the potency of the bacteriocin. Substituting residues in the loop region between Cys7 and Cys22 for D-amino acid equivalents had a more detrimental effect on activity than L-alanine substitutions. Specifically Y10A, N11A, P15A and T16A are active at 4, 16, 1 and 16 μM respectively while their D equivalents were inactive at 1000 μM, the highest concentration tested. Ultimately, this study identifies the critical features in the primary structure of the bacteriocin which gives it such potent activity against pathogenic staphylococci.
Lactobacillus salivarius , Amino acid , Pathogenic staphylococci , N terminal
O’ Connor, P. M., O’ Shea, E. F., Cotter, P. D., Hill, C. and Ross, R. P. (2018) 'The potency of the broad spectrum bacteriocin, bactofencin A, against staphylococci is highly dependent on primary structure, N-terminal charge and disulphide formation', Scientific Reports, 8(1), 11833 (8pp). doi: 10.1038/s41598-018-30271-6
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