Interaction between antimicrobial peptides and microorganisms: interdisciplinary study of mechanisms

SZYMANOWSKI F; PICKHOLZ M; TYMCZYSZYN E; BALATTI G E; PEREZ P; HUGO A; MOBILI P; GOMEZ-ZAVAGLIA A

San Miguel de Tucuman

Simposio; V Simposio Internacional de Bacterias Lácticas (SIBAL). Alimentos, Salud y Aplicaciones; 2016

Differential susceptibility to defensins (cationic antimicrobial peptides) could be a relevant property of potentially probiotic microorganisms. The present work aimed at gaining further insight on the interaction between defensins and lactobacilli through a multidisciplinary approach that includes biological, 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). Liposomes prepared with bacterial lipids. Antimicrobial peptides: Human beta defensins 1 and 2 (HBD1 and HBD2) or surrogate molecules (polimixin and nisin). Effect on whole 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 analysed by FTIR. Molecular dynamics: Coarse Grain (CG) model and MARTINI force field in combination with the elastic springs network (Elnedyn) for HBD1 and model membranes of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphatidyl-choline (POPC) and 2-oleoyl-1-pamlitoyl-sn-glyecro-3-glycerol (POPG). Exposure of lactobacilli to HBD-2 lead to an increase of the membrane permeability being strain CIDCA 331 the most susceptible strain. Liposomes prepared with lipids from strain CIDCA 133 were destabilized neither by HBD-1 nor by HBD-2, whereas liposomes derived from strain CIDCA 331 were susceptible to HBD-2 but not to HBD-1. Effect of defensins was strongly inhibited in the presence of NaCl, and the activity increased in water. FTIR studies showed significant spectral modifications in the presence of the surrogates molecules, nisin and polymyxin. The most significant changes were observed at the lipid hydrocarbon chain CH2 antisymmetrical stretching band (~ 2928 cm-1), the phosphate antisymmetrical stretching band (~ 1230 cm-1) and the ester CO-O-C symmetrical stretching band (~ 1065 cm-1) Computational simulations show that HBD1 is essentially found at the lipid water interphase for both, POPC and POPG, lipid bilayers. However, the interaction is most specific with POPG membranes. Our results demonstrate the relevance of the multidisciplinary approaches in order to gain further insight on the interaction of potentially probiotic microorganisms and key effectors of the host´s immune system.