Mode of action of the antimicrobial peptide indolicidin.
Indolicidin is a cationic antimicrobial peptide isolated from bovine neutrophils. It consists of only 13 amino acids, has the highest tryptophan content of any known protein, and is amidated at the carboxyl terminus in nature. By circular dichroism spectroscopy a weak poly-L-proline II extended helix structure was observed that became substantially more pronounced upon interaction with liposomes. Indolicidin bound purified surface lipopolysaccharide with high affinity and permeabilized the outer membrane of Escherichia coli to the small hydrophobic molecule 1-N-phenylnapthylamine (Mr 200), results consistent with indolicidin crossing the outer membrane via the self-promoted uptake pathway. The methyl esterification of indolicidin's carboxyl terminus increased its activity for Gram-negative and Gram-positive bacteria. In Gram-negative bacteria this was associated with an increased binding to lipopolysaccharide and increased permeabilization of the outer membrane. The cytoplasmic membrane was the site of action of indolicidin as assayed in E. coli by the unmasking of cytoplasmic beta-galactosidase due to membrane permeabilization. The mechanism for this activity was shown to be the ability of the peptide to cause an increase in the transmembrane current of planar lipid bilayers. This current increase was activated by transmembrane potentials in excess of -70 to -80 mV. Consistent with this, there was a substantial decrease in indolicidin-mediated bacterial killing and permeabilization of the cytoplasmic membrane of E. coli that had been pretreated with the uncoupler carbonyl cyanide-m-chlorophenyl hydrazone. In planar bilayers, indolicidin induced the formation of discrete channels, which ranged in conductance from 0.05-0.15 nS. Thus despite the small size and unique composition of indolicidin, it was capable of killing Gram-negative bacteria by crossing the outer membrane and causing disruption of the cytoplasmic membrane by channel formation.
Falla
T J
TJ
Department of Microbiology and Immunology, University of British Columbia, Vancouver, British Columbia, V6T 1Z3 Canada.
Karunaratne
D N
DN
Hancock
R E
RE
D000900:Anti-Bacterial Agents / Q000378:metabolism / Q000494:pharmacology*; D023181:Antimicrobial Cationic Peptides; D002462:Cell Membrane / Q000187:drug effects / Q000502:physiology; D006090:Gram-Negative Bacteria / Q000187:drug effects*; D008051:Lipid Bilayers; D008070:Lipopolysaccharides / Q000378:metabolism; D008564:Membrane Potentials / Q000187:drug effects; D008826:Microbial Sensitivity Tests; D010455:Peptides / Q000378:metabolism / Q000494:pharmacology*
D000900:Anti-Bacterial Agents; D023181:Antimicrobial Cationic Peptides; D008051:Lipid Bilayers; D008070:Lipopolysaccharides; D010455:Peptides; C073143:indolicidin