Interactions of de novo designed peptides with bacterial membranes: Implications in antimicrobial activity, Maturana

MATURANA P; MARTINEZ, M; ESPECHE, JC; MAFFÍA, PAULO C.; HOLLMANN A.

La Plata

Congreso; XLVII Reunión Anual de la Sociedad Argentina de Biofisica; 2018

Sociedad Argentina de Biofísica

Antimicrobial peptides (AMPs) are small molecules, usually cationic, that display antimicrobial activity against a wide range of bacteria, fungi, and viruses. In previous work by using model membranes we studied two related novel peptides, P6 that show no antimicrobial activity and P6.2 witch exhibited good antibacterial activity. Besides biophysical techniques on model membranes can provide very detailed information on the interaction of AMPs with membranes, until recently, it was unknown if any of the information predicted was relevant for AMPs activity in real bacteria. In this context, in the present work, we aimed to unravel the way of action of this peptide by studying its interaction with whole bacteria using Escherichia coli and Staphylococcus aureus as a bacterial model. Zeta Potential experiments confirming that both peptides were able to interact with bacterial envelope of E. coli and S. aureus. However, the effects on P6.2 were much more noticeable in both bacteria. Interesting, besides on both kinds of bacteria the CIM obtained for P6.2 were similar (12 M) the affinity seems be higher in the case of E. coliIn order to get an insight into the effect that both peptides induce in the bacterial membrane, the disruption membrane assays were carried out. In the case of E. coli, the outer (OM) and inner (IM) membrane permeabilization were analyzed. For OM permeabilization study the probe 1-Nphenylnaphthylamine (NPN), only able to insert in previously damage OM, was used. Beside both peptides were able to damage the OM, 5 times concentration of P6 was needed in order to obtain comparable results than those obtained with P6.2. Also, P6.2 exhibited much faster damage kinetic. IM disruption was assessed following the colored product of cytosolic -galactosidase. Anew, P6.2 not only exhibits more product formation, implying a greater disruption of the membrane but a faster kinetic.In the case o S. aureus, NPN assays were also carried out to evaluate the ability of the peptides to disrupt the bacterial membrane. In the same way that for E. Coli, both peptides were able to permeabilize the plasmatic membrane of S. aureus, but the effects were much more noticeable with the peptide 6.2. When NPN uptake data were compared in both bacteria, E. coli showed faster kinetics and a lower amount of peptide to obtain the full permeabilization. Those results could be explained due to the OM in E. coli its higher exposure to the media whereas in S. aureus peptides requires pass trough peptidoglycan to reach the plasmatic membrane. All data put together allows postulating, in a physiologic model, that the lower affinity of P6 for bacterial envelope results in a minor final concentration of the peptide in the bacterial membrane unable to trigger the antimicrobial activity.