Interaction between antimicrobial peptides and microorganisms: interdisciplinary study of mechanisms

HUGO, A. A.; TYMCZYSZYN, E. E.; SZYMANOWSKI, F.; MOBILI, P.; BALATTI, G. E.; MARTINI, M. F.; PICKHOLZ, M.; GÓMEZ-ZAVAGLIA, A. AND PÉREZ, P. F.

Simposio; V Simposio Internacional de Bacterias Lácticas (SIBAL 2016).; 2016

Differentialsusceptibility to defensins (cationic antimicrobial peptides) could be arelevant property of potentially probiotic microorganisms. Thepresent work aimed at gaining further insight on the interaction betweendefensins and lactobacilli through a multidisciplinary approach that includesbiological, spectroscopic and molecular dynamics methods.Strains:lactobacilli with different susceptibility to defensins (Lact. delbrueckii subsp.lactis   CIDCA 133 and Lact. delbrueckii subsp. bulgaricus CIDCA 331). Liposomesprepared with bacterial lipids. Antimicrobial peptides: Human beta defensins 1and 2 (HBD1 and HBD2) or surrogate molecules (polimixin and nisin). Effect onwhole bacteria was assessed by flow cytometry after propidium iodide (PI)staining and carboxyfluorescein release was evaluated in liposomes.Spectroscopic methods: lecithin liposomes with nisin or polymyxin were analysedby FTIR. Molecular dynamics: Coarse Grain (CG) model and MARTINI force field incombination with the elastic springs network (Elnedyn) for HBD1 and modelmembranes of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphatidyl-choline (POPC) and2-oleoyl-1-pamlitoyl-sn-glyecro-3-glycerol (POPG).  Exposureof lactobacilli to HBD-2 lead to an increase of the membrane permeability beingstrain CIDCA 331 the most susceptible strain. Liposomes prepared with lipidsfrom strain CIDCA 133 were destabilized neither by HBD-1 nor by HBD-2, whereasliposomes derived from strain CIDCA 331 were susceptible to HBD-2 but not toHBD-1. Effect of defensins was strongly inhibited in the presence of NaCl, andthe activity increased in water. FTIR studies showed significant spectralmodifications in the presence of the surrogates molecules, nisin and polymyxin.The most significant changes were observed at the lipid hydrocarbon chain CH2antisymmetrical stretching band (~ 2928 cm-1), the phosphate antisymmetricalstretching band (~ 1230 cm-1) and the ester CO-O-C symmetrical stretching band(~ 1065 cm-1) Computational simulations show that HBD1 is essentially found atthe lipid water interphase for both, POPC and POPG, lipid bilayers. However,the interaction is most specific with POPG membranes. Ourresults demonstrate the relevance of the multidisciplinary approaches in orderto gain further insight on the interaction of potentially probioticmicroorganisms and key effectors of the host´s immune system.