Design, NMR characterization and activity of a 21-residue peptide fragment of bacteriocin AS-48 containing its putative membrane interacting region.
Bacteriocin AS-48 is a 70-residue cyclic polypeptide from Enterococcus faecalis that shows a broad antimicrobial spectrum against both Gram-positive and Gram-negative bacteria. The structure of bacteriocin AS-48 consists of a globular arrangement of five helices with a high positive electrostatic potential in the region comprising helix 4, the turn linking helix 4 and 5, and the N-terminus of helix 5. This region has been considered to participate in its biological activity and in particular in membrane permeation. To understand the mechanism of the antibacterial activity of AS-48 and to discriminate the several mechanisms proposed, a simplified bacteriocin was designed consisting of 21 residues and containing the high positively charged region. A disulfide bridge was introduced at an appropriate position to stabilize the peptide and to conserve the helix-turn-helix arrangement in the parent molecule. According to (1)H and (13)C NMR data, the designed simplified bacteriocin fragment adopts a significant population of a native-like helical hairpin conformation in aqueous solution, which is further stabilized in 30% TFE. The designed peptide does not show any antibacterial activity, though it is shown to compete with the intact native bacteriocin AS-48. These results suggest that the mechanism of membrane disruption by bacteriocin is not as simple as being driven by a deposition of positively charged molecules on the plane of the bacterial membrane. Some other regions of the protein must be present such as, for instance, hydrophobic regions so as to enhance the accumulation of the peptide and favour membrane permeation.
Jiménez
M Angeles
MA
Instituto de Química-Física Rocasolano, Consejo Superior de Investigaciones Científicas, Serrano 119, 28006-Madrid, Spain. majimenez@iqfr.csic.es
Barrachi-Saccilotto
Ana C
AC
Valdivia
Eva
E
Maqueda
Mercedes
M
Rico
Manuel
M
D001430:Bacteriocins / Q000737:chemistry* / Q000276:immunology / Q000494:pharmacology; D002462:Cell Membrane / Q000187:drug effects; D019077:Helix-Turn-Helix Motifs; D008089:Listeria monocytogenes / Q000187:drug effects; D009682:Magnetic Resonance Spectroscopy; D010446:Peptide Fragments / Q000138:chemical synthesis* / Q000276:immunology / Q000494:pharmacology
D001430:Bacteriocins; D010446:Peptide Fragments; C440289:bacteriocin AS-48, Enterococcus faecalis