Biophysical investigations into the interaction of lipopolysaccharide with polymyxins

It is reported on the interaction of bacterial lipopolysaccharide (LPS, endo-toxin), having different sugar chain length covalently bound to its hydrophobic moiety lipid A, with the polycationic antibiotics polymyxin B (PMB) and PMB-nonapeptide (PMBN). The binding enthalpies and the lipid:peptide binding stoichiometries were determined by isothermal titration calorimetry (ITC). For LPS with a long sugar chain (S: smooth form LPS from Yersinia enterocolitica), the titration curves exhibit a strong exo-therm which can be interpreted to result from the electrostatic interaction of the negative charges of the LPS with the positive charges of the peptides. In contrast, the titration curves of LPS with a short sugar chain (LPS Re from Escherichia coli) and of free lipid A yield complex patterns of endo- and exo-therms, which result from the superposition of electrostatic binding, fluidization of the acyl chains and a transition between different three-dimensional aggregate structures of the endo-toxins due to peptide binding. Infrared spectroscopy indicates that the fluidizing effect of the polymyxins is similar for both types of LPS and for lipid A. Small-angle X-ray diffraction reveals, however, that the Re-type LPS and lipid A are converted from a cubic into a multilamellar structure, whereas, the S-form LPS transforms from a unilamellar into a multilamellar structure with a small number of lamellae. The presented data allow a better understanding of the interaction of peptides with endo-toxin molecules.