Interaction of polycationic antibiotics with Pseudomonas aeruginosa lipopolysaccharide and lipid A studied by using dansyl-polymyxin

Abstract

A fluorescent derivative of polymyxin B (dansyl-polymyxin) was used to study the interaction of polycations with lipopolysaccharide (LPS) and lipid A from Pseudomonas aeruginosa. Dansyl-polymyxin Pseudomonas aeruginosa is presently recognized as one of the leading causes of death from gram-negative septicemia in North America (4). A major factor contributing to the success of this organism as an opportunistic pathogen is its intrinsic resistance to antibiotics. A proposed basis for antibiotic resistance in P. aeruginosa is the low rate of antibiotic permeation across the outer membrane due to the intrinsic barrier provided by the highly negatively charged lipopolysaccharide (LPS) and the poor functioning of the major porin protein F (16). Despite the major barrier presented by the outer membrane of P. aeruginosa, polycationic antibiotics like aminoglycosides are able to cross the membrane to some extent and have been used successfully in therapy (4).While porin proteins provide one pathway for antibiotics to cross the outer membrane, workers in our laboratory have proposed the self-promoted pathway for antibiotic uptake as an alternative means for polycationic antibiotics (e.g., aminoglycosides and polymyxins) to pass through the outer membrane of P. aeruginosa (5,6,15). The results of previous studies of the interaction of polymyxin B with the outer membranes of other bacteria (20) were consistent with the proposal that polymyxin B uptake occurs via the selfpromoted uptake pathway, as described for P. aeruginosa (5). The self-promoted uptake model requires that the polycationic antibiotic must first bind to LPS, resulting in the displacement of Mg2' and disruption of the stabilizing effect afforded by the Mg2+ cross-bridging of adjacent LPS molecules in the outer membrane. The resultant permeabilized outer membrane then allows uptake of the interacting polycationic antibiotic; hence, the polycations promote their own uptake across the outer membrane. This model is supported by the facts that polymyxin B is known to interact with the LPS from Salmonella typhimurium (9)

Inhibition of Lipopolysaccharide Activity by a Bacterial Cyclic Lipopeptide Surfactin

“…1). It is an amphiphilic compound and known to be one of the most powerful biosurfactants [22,26]. Surfactin was also reported to inhibit phospholipase A 2 (PLA 2 ) [27], and to enhance plasminogen activation [28].…”

“…Surfactin is an amphiphilic compound and has surfaceactivating potential [22]. So, we studied whether the cell adhesion could be inhibited by general surface-activating compounds.…”

Dissociation of Antimicrobial and Hemolytic Activities in Cyclic Peptide Diastereomers by Systematic Alterations in Amphipathicity

“…We investigated the interaction between GS14 diastereomers and the bacterial outer membranes by monitoring the displacement of LPS-bound dansyl-polymyxin B by the peptides (19,39). The LPS binding affinity of GS14 was extremely strong, approaching that of polymyxin B itself (19), whereas all the diastereomers had lower affinity (Table I).…”

Capsule polysaccharide is a bacterial decoy for antimicrobial peptides

“…Secondly, we evaluated the binding of dansyl-polymyxin B to CPSs. This compound has been extensively used to study the binding of APs to polysaccharides, proteins and intact bacteria (Bengoechea et al, 1998;Campos et al, 2006;Moore et al, 1986). The dansylated peptide exhibits low fluorescent yield when diluted into aqueous buffer; however, the fluorescent yield increases, with the maximum fluorescence shifting to a lower wavelength, in a hydrophobic environment (Moore et al, 1986;Newton, 1955).…”