Oxidation of Lanthionines Renders the Lantibiotic Nisin Inactive

Abstract

The peptide antibiotic nisin A belongs to the group of antibiotics called lantibiotics. They are classified as lantibiotics because they contain the structural group lanthionine. Lanthionines are composed of a single sulfur atom that is linked to the ␤-carbons of two alanine moieties. These sulfur atoms are vulnerable to environmental oxidation. A mild oxidation reaction was performed on nisin A to determine the relative effects it would have on bioactivity. High-mass-accuracy Fourier transform ion cyclotron resonance mass spectrometry data revealed the addition of seven, eight, and nine oxygens. These additions correspond to the five lanthionines, two methionines, and two histidines that would be susceptible to oxidation. Subsequent bioassays revealed that the oxidized form of nisin A had a complete loss of bactericidal activity. In a competition study, the oxidized nisin did not appear to have an antagonistic affect on the bioactivity of nisin A, since the addition of an equal molar concentration of the oxidized variant did not have an influence on the bactericidal activity of the native antibiotic. Electron microscopy data revealed that the oxidized forms were still capable of assembling into large circular complexes, demonstrating that oxidation does not disrupt the lateral assembly mechanism of the antibiotic. Affinity thin-layer chromatography and fluorescence microscopy experiments suggested that the loss of activity is due to the inability of the oxidized form of nisin to bind to the cell wall precursor lipid II. Given the loss of bioactivity following oxidation, oxidation should be an important factor to consider in future production, purification, pharmacokinetic, and pharmacodynamic studies.

Fundamental functionality: recent developments in understanding the structure–activity relationships of lantibiotic peptides

“…Additionally, many lantibiotics are not stable over the full pH range, as under basic conditions, many peptides undergo oxidation of the lan/ melan thioethers, resulting in a loss of antimicrobial activity. 10 These structural disadvantages along with a desire for smaller/simpler peptides have driven much of the research into lantibiotic analogs. If clinically useful peptides are to be developed from lantibiotics, solutions need to be found for these inherent challenges.…”

Site-Directed Mutations in the Lanthipeptide Mutacin 1140

“…Resistance to hydrolysis may also be, in part, a reflection of the unusual, horseshoe-shaped three-dimensional structure of mutacin 1140 (20,21). The hinge region contains a potentially protease-susceptible arginine at residue 13. This residue appears to be protected, as evidenced by the relative difficulty of inactivating mutacin 1140 by trypsin or pronase treatment (20)(21)(22).…”

“…Nisin, one of the most studied lanthipeptides, is produced by Lactococcus lactis and has been used in the food industry for over 50 years. It was discovered that both nisin and mutacin 1140 abduct lipid II from the site of new cell wall synthesis, ultimately causing cell death (10)(11)(12)(13). Rings A and B provide the structural motif for binding to lipid II and are referred to as the lipid II binding domain (14).…”

Post-Translational Modifications of Natural Antimicrobial Peptides and Strategies for Peptide Engineering

“…membranes). Wilson-Stanford et al [30] found that oxidation of lantibiotics such as nisin A disrupted antibacterial activity. It was proposed that the oxidized form of sulfur is unable to bind to lipid II of the cell wall.…”

“…While terminal capping, selective D-amino acid incorporation, disulfide bond formation, and head-to-tail cyclization have been applied to rational design of therapeutic peptides, other modifications such as phosphorylation and glycosylation are probably created mainly for specific functional roles of polypeptides only (Table 1). Some type of chemical modifications such as oxidation during peptide storage, however, should be avoided, since it could reduce peptide activity [29,30].…”