uridine-diphosphate-n-acetylmuramic-acid and lanthionine

The combined activity of the constituents of two-component antibiotic systems is always significantly higher than the sum of the activities of the individual pieces. Understanding the principles behind this phenomenon might provide new ways to design new antibiotics. In this issue of Molecular Microbiology, Wiedemann and coworkers have made a big step towards understanding the mechanism of action of the two-component lanthionine-containing antibiotic lacticin 3147. It has now become clear that this two-component system specifically targets the bacterial cell wall precursor Lipid II. This makes this essential bacterial lipid one of the most sought-after targets in nature. Surprisingly, in view of its small size (MW 1875 Da), this is now the fifth different way that this key molecule is known to be targeted.

Topics: Alanine; Anti-Bacterial Agents; Bacteriocins; Nisin; Sulfides; Uridine Diphosphate N-Acetylmuramic Acid

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 beta-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.

Topics: Alanine; Anti-Bacterial Agents; Macromolecular Substances; Mass Spectrometry; Microbial Sensitivity Tests; Micrococcus luteus; Microscopy, Electron; Models, Molecular; Nisin; Oxidation-Reduction; Staphylococcus aureus; Streptococcus pneumoniae; Sulfides; Uridine Diphosphate N-Acetylmuramic Acid

The increasing resistance of human pathogens to conventional antibiotics presents a growing threat to the chemotherapeutic management of infectious diseases. The lanthionine antibiotics, still unused as therapeutic agents, have recently attracted significant scientific interest as models for targeting and management of bacterial infections. We investigated the action of one member of this class, subtilin, which permeabilizes lipid membranes in a lipid II-dependent manner and binds bactoprenyl pyrophosphate, akin to nisin. The role the C and N termini play in target recognition was investigated in vivo and in vitro by using the natural N-terminally succinylated subtilin as well as enzymatically truncated subtilin variants. Fluorescence dequenching experiments show that subtilin induces leakage in membranes in a lipid II-dependent manner and that N-succinylated subtilin is roughly 75-fold less active. Solid-state nuclear magnetic resonance was used to show that subtilin forms complexes with membrane isoprenyl pyrophosphates. Activity assays in vivo show that the N terminus of subtilin plays a critical role in its activity. Succinylation of the N terminus resulted in a 20-fold decrease in its activity, whereas deletion of N-terminal Trp abolished activity altogether.

Topics: Alanine; Anti-Bacterial Agents; Bacteriocins; Cell Membrane; Coated Vesicles; Diphosphates; Fluoresceins; Lactococcus lactis; Magnetic Resonance Spectroscopy; Microbial Sensitivity Tests; Peptides; Phosphatidylcholines; Phosphatidylglycerols; Succinic Acid; Sulfides; Tryptophan; Uridine Diphosphate N-Acetylmuramic Acid