RESUMO
Grazing incidence x-ray scattering techniques and Monte Carlo (MC) simulations are combined to reveal the influence of molecular structure (genetic mutation) and divalent cations on the survival of gram negative bacteria against cationic peptides such as protamine. The former yields detailed structures of bacterial lipopolysaccharide (LPS) membranes with minimized radiation damages, while the minimal computer model based on the linearized Poisson-Boltzmann theory allows for the simulation of conformational changes of macromolecules (LPSs and peptides) that occur in the time scale of ms. The complementary combination of the structural characterizations and MC simulation demonstrates that the condensations of divalent ions (Ca2+ or Mg2+) in the negatively charged core saccharides are crucial for bacterial survival.
Assuntos
Escherichia coli/efeitos dos fármacos , Escherichia coli/metabolismo , Lipopolissacarídeos/química , Lipopolissacarídeos/metabolismo , Viabilidade Microbiana/efeitos dos fármacos , Método de Monte Carlo , Protaminas/farmacologia , Animais , Cálcio/farmacologia , Cristalografia por Raios X , Escherichia coli/genética , Escherichia coli/fisiologia , Lipídeo A/química , Lipopolissacarídeos/genética , Mutação , Pressão , Protaminas/metabolismoRESUMO
A model of the outer membrane of Gram-negative bacteria was created by the deposition of a monolayer of purified rough mutant lipopolysaccharides at an air/water interface. The density profiles of monovalent (K(+)) and divalent (Ca(2+)) cations normal to the lipopolysaccharides (LPS) monolayers were investigated using grazing-incidence X-ray fluorescence. In the absence of Ca(2+), a K(+) concentration peak was found in the negatively charged LPS headgroup region. With the addition of CaCl(2), Ca(2+) ions almost completely displaced K(+) ions from the headgroup region. By integrating the experimentally reconstructed excess ion density profiles, we obtained an accurate measurement of the effective charge density of LPS monolayers. The experimental findings were compared to the results of Monte Carlo simulations based on a coarse-grained minimal model of LPS molecules and showed excellent agreement.