Interaction of protamine with gram-negative bacteria membranes: possible alternative mechanisms of internalization in Escherichia coli, Salmonella typhimurium and Pseudomonas aeruginosa.
This study was concerned with the interaction between the cationic antimicrobial peptide, protamine (Ptm) and the cytoplasmic membranes of the gram-negative bacteria Escherichia coli, Salmonella typhimurium and Pseudomonas aeruginosa. The objective of the study was to explain the observed paradox of internalization without permanent disruption of the cell envelope. We carried out Monte Carlo computer simulation of Ptm in an aqueous environment in the presence of ~100 mM NaCl and model membranes consisting of either (65:35) or (75:25) PE:PG molar ratios. The (75:25) model, representative of the gram-negative cytoplasmic membrane, showed that the Ptm center of mass remained at least 7 nm from the membrane surface leading to the prediction that Ptm would not internalize via disruption of the inner membrane. By using immunoelectron microscopy of Ptm-treated cells, we showed that Ptm internalization to the cytoplasm took place rapidly in the presence or absence of the outer envelope. Ultrastructural examination revealed no obvious morphological changes to cells that were treated with subinhibitory or bactericidal levels of Ptm. Reconstituted phospholipid bilayers were constructed and were unperturbed by Ptm treatment over a wide range of concentrations and applied transmembrane voltages. We conclude that in the cases of the cell envelopes of E. coli, S. typhimurium and P. aeruginosa, Ptm internalized by means independent of the phospholipid bilayer, most likely mediated by one or more membrane proteins such as cation-selective barrel-like proteins. Work is currently underway to test this hypothesis.
Department of Physics, St. Francis Xavier University, Antigonish, NS, B2G 2W5, Canada.
David A
DA
Pink
Fida M
FM
Hasan
Bonnie E
BE
Quinn
Mathias
M
Winterhalter
Mukund
M
Mohan
Tom A
TA
Gill
D000595:Amino Acid Sequence; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D002462:Cell Membrane / Q000187:drug effects*; D003198:Computer Simulation; D004352:Drug Resistance, Microbial; D004926:Escherichia coli / Q000378:metabolism*; D006860:Hydrogen Bonding; D008051:Lipid Bilayers; D008826:Microbial Sensitivity Tests; D008969:Molecular Sequence Data; D011479:Protamines / Q000494:pharmacology*; D011485:Protein Binding; D011550:Pseudomonas aeruginosa / Q000378:metabolism*; D012486:Salmonella typhimurium / Q000378:metabolism*; D055672:Static Electricity
D000900:Anti-Bacterial Agents; D008051:Lipid Bilayers; D011479:Protamines
antimicrobial peptides; gram negative bacteria; membranes; protamine