Selective Interaction of Colistin with Lipid Model Membranes

Although colistin's clinical use is limited due to its nephrotoxicity, colistin is considered to be an antibiotic of last resort because it is used to treat patients infected with multidrug-resistant bacteria. In an effort to provide molecular details about colistin's ability to kill Gram-negative (G(−)) but not Gram-positive (G(+)) bacteria, we investigated the biophysics of the interaction between colistin and lipid mixtures mimicking the cytoplasmic membrane of G(+), G(−) bacteria as well as eukaryotic cells. Two different models of the G(−) outer membrane (OM) were assayed: lipid A with two deoxy-manno-octulosonyl sugar residues, and Escherichia coli lipopolysaccharide mixed with dilaurylphosphatidylglycerol. We used circular dichroism and x-ray diffuse scattering at low and wide angle in stacked multilayered samples, and neutron reflectivity of single, tethered bilayers mixed with colistin. We found no differences in secondary structure when colistin was bound to G(−) versus G(+) membrane mimics, ruling out a protein conformational change as the cause of this difference. However, bending modulus KC perturbation was quite irregular for the G(−) inner membrane, where colistin produced a softening of the membranes at an intermediate lipid/peptide molar ratio but stiffening at lower and higher peptide concentrations, whereas in G(+) and eukaryotic mimics there was only a slight softening. Acyl chain order in G(−) was perturbed similarly to KC. In G(+), there was only a slight softening and disordering effect, whereas in OM mimics, there was a slight stiffening and ordering of both membranes with increasing colistin. X-ray and neutron reflectivity structural results reveal colistin partitions deepest to reach the hydrocarbon interior in G(−) membranes, but remains in the headgroup region in G(+), OM, and eukaryotic mimics. It is possible that domain formation is responsible for the erratic response of G(−) inner membranes to colistin and for its deeper penetration, which could increase membrane permeability.Fil: Dupuy, Fernando Gabriel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Instituto Superior de Investigaciones Biológicas. Universidad Nacional de Tucumán. Instituto Superior de Investigaciones Biológicas; Argentina. University of Carnegie Mellon; Estados UnidosFil: Pagano, Isabella. University of Carnegie Mellon; Estados UnidosFil: Andenoro, Kathryn. University of Carnegie Mellon; Estados UnidosFil: Peralta, Maria Florencia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones en Ciencias de la Salud. Universidad Nacional de Córdoba. Instituto de Investigaciones en Ciencias de la Salud; Argentina. University of Carnegie Mellon; Estados UnidosFil: Elhady, Yasmene. University of Carnegie Mellon; Estados UnidosFil: Heinrich, Frank. University of Carnegie Mellon; Estados UnidosFil: Tristram-nagle, Stephanie. University of Carnegie Mellon; Estados Unido

Elastic behavior of model membranes with antimicrobial peptides depends on lipid specificity and d-enantiomers

In an effort to provide new treatments for the global crisis of bacterial resistance to current antibiotics, we have used a rational approach to design several new antimicrobial peptides (AMPs). The present study focuses on 24-mer WLBU2 and its derivative, D8, with the amino acid sequence, RRWVRRVRRWVRRVVRVVRRWVRR. In D8, all of the valines are the Denantiomer. We use X-ray low- and wide-angle diffuse scattering data to measure elasticity and lipid chain order. We show a good correlation between in vitro bacterial killing efficiency and both bending and chain order behavior in bacterial lipid membrane mimics; our results suggest that AMP-triggered domain formation could be the mechanism of bacterial killing in both Grampositive and Gram-negative bacteria. In red blood cell lipid mimics, D8 stiffens and orders the membrane, while WLBU2 softens and disorders it, which correlate with D8’s harmless vs. WLBU2’s toxic behavior in hemolysis tests. These results suggest that elasticity and chain order behavior can be used to predict mechanisms of bactericidal action and toxicity of new AMPs.Fil: Kumagai, Akari. University of Carnegie Mellon; Estados UnidosFil: Dupuy, Fernando Gabriel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet Noa Sur. Instituto Superior de Investigaciones Biológicas. Grupo de Investigación y Desarrollo del Noroeste Argentino | Universidad Nacional de Tucumán. Instituto Superior de Investigaciones Biológicas. Grupo de Investigación y Desarrollo del Noroeste Argentino; ArgentinaFil: Arsov, Zoran. Jožef Stefan Institute; EsloveniaFil: Elhady, Yasmene. University of Carnegie Mellon; Estados UnidosFil: Moody, Diamond. University of Carnegie Mellon; Estados UnidosFil: Erns, Robert. University of Maryland; Estados UnidosFil: Deslouches, Berthony. University of Pittsburgh; Estados UnidosFil: Montelaro, Ronald. University of Pittsburgh; Estados UnidosFil: Di, Yuanpu Peter. University of Pittsburgh; Estados UnidosFil: Tristram-Nagle, Stephanie. University of Carnegie Mellon; Estados Unido