Bacteria’s inner and outer membrane permeabilization by “de novo” antimicrobial peptide P1 through spectroscopic methods

ESPECHE, JC; MATURANA, P; MARTINEZ, M; MAFFIA, P; HOLLMANN, A

Congreso; XLVIII Reunión Anual de la Sociedad Argentina de Biofísica; 2019

In the present work, we evaluate the interaction of the antimicrobial peptide 1 (P1) withbacterial membranes as well as its ability to permeabilize the inner and outer membraneof Gram positive and Gram negative bacteria. P1 is a cationic peptide with 21 aminoacids (WPKWWKWKRRWGRKKAKKRRG), designed identifying short putative active regionsfrom AMP databases. Its antimicrobial activity was previously tested in Staphylococcusaureus and Escherichia coli strains having gotten a MIC value of 32 and 128 respectively.Inicially, we evaluated the interaction of P1 with bacterial envelope by using ZetaPotential in suspensions containing the mentioned strains. In both cases, zeta potentialbecame less negative after peptide incubation confirming the ability of the peptides tobound into the cell envelope. However, the effect becomes more noticeable in S. aureus.in a good agridemnt with MIC dataNext, in order to determine the ability of the peptide to permeabilize the outermembrane of these bacteria, fluorescence methods were used. Results have shown thatP1 is able to disrupt both strains’ outer membranes reaching higher values and fasterkinetics in the case of E. coli. In the case of S. aureus kinetics are slightly slower.Inner membrane permeabilization was assessed only for E. coli using absorbance. It wasfound that the kinetics for the disruption of this membrane is reached successfully by thepeptide. In this case, it required longer time lapses for the permeabilization to happen.From the results we got, we can conclude that the peptide is able to permeabilize bothouter and inner membrane of E. coli, reaching the first effect in much shorter times (fromseconds to a pair of minutes) than the second (1 hour approximately). This differencecould be due to the fact that the peptide needs to permeabilize the outer membrane firstin order to reach the inner. Differences observed in disruption speed for the membrane inS. aureus (Gram positive bacteria) could be explained since this bacteria have a thickercell wall, that might hinder the reach of the peptide to the membrane. Furthermore, arelation between concentration of peptide and membrane damage was stablished forboth membranes.