Minor sequence modifications in temporin B cause drastic changes in antibacterial potency and selectivity by fundamentally altering membrane activity

Abstract

Antimicrobial peptides (AMPs) are a potential source of new molecules to counter the increase in antimicrobial resistant infections but a better understanding of their properties is required to understand their native function and for effective translation as therapeutics. Details of the mechanism of their interaction with the bacterial plasma membrane are desired since damage or penetration of this structure is considered essential for AMPs activity. Relatively modest modifications to AMPs primary sequence can induce substantial changes in potency and/or spectrum of activity but, hitherto, have not been predicted to substantially alter the mechanism of interaction with the bacterial plasma membrane. Here we use a combination of molecular dynamics simulations, circular dichroism, solid-state NMR and patch clamp to investigate the extent to which temporin B and its analogues can be distinguished both in vitro and in silico on the basis of their interactions with model membranes. Enhancing the hydrophobicity of the N-terminus and cationicity of the C-terminus in temporin B improves its membrane activity and potency against both Gram-negative and Gram-positive bacteria. In contrast, enhancing the cationicity of the N-terminus abrogates its ability to trigger channel conductance and renders it ineffective against Gram-positive bacteria while nevertheless enhancing its potency against Escherichia coli. Our findings suggest even closely related AMPs may target the same bacterium with fundamentally differing mechanisms of action.

The Antimicrobial Peptide Temporin G: Anti-Biofilm, Anti-Persister Activities, and Potentiator Effect of Tobramycin Efficacy Against Staphylococcus aureus

“…Temporins are a family of frog-skin AMPs whose various isoforms have already shown activity against Gram-positive bacteria [23,41,42]. For instance, TA has a MIC ranging from 4 to 16 µg/mL towards human methicillin-resistant S. aureus (MRSA) clinical isolates [18]; TB and different synthetic analogs inhibit the growth of various S. aureus strains alone or in combination with TA [22,43]. The cytotoxic TL is noticeably more potent than other temporin peptides in killing S. aureus [44,45] and can act in synergy with TA or TB against Gram-negative bacteria by inducing changes in the biophysical properties of the peptides (e.g., prevention of their oligomeric state) when bound to the lipopolysaccharide [46].…”

“…Temporins were initially isolated from the European red frog Rana temporaria, and nowadays they represent one of the largest groups of amphibian AMPs (more than 100 members) [15,16] and are among the shortest AMPs (only 10-13 residues) present in nature to date. While some isoforms (i.e., temporin A, (TA [FLPLIGRVLSGIL-NH 2 ]); temporin B (TB [LLPIVGNLLKSLL-NH 2 ]); temporin L (TL [FVQWFSKFLGRIL-NH 2 ]) of this family have already been characterized for their antimicrobial and cytotoxic properties [16][17][18][19][20][21][22][23][24], very little is known about the antimicrobial potential of temporin G (TG [FFPVIGRILNGIL-NH 2 ]) [25], which was isolated from the same frog specimen, especially against the bacterial biofilm phenotype. To the best of our knowledge, this is the first manuscript describing TG activity against preformed S. aureus biofilm as well as against persister cells.…”

Design, Engineering and Discovery of Novel α-Helical and β-Boomerang Antimicrobial Peptides against Drug Resistant Bacteria

“…TB_L1FK (Table 1) is a computationally designed analog of TB that contains a replacement of Leu1 to Phe, the deletion of residue Asn at position 7, and an addition of Lys13 at the C-terminus. Studies comparing the antibacterial activity of TB_KKG6A and TB_L1KF [109,110] showed that both analogs showed higher MIC values of 32 to 64 μM, against most of the Gram negative strains tested; however, Figure 1. Structures of (A) the TL dimer; (B) TA; and, (C) TB in LPS.…”

Therapeutic Potential of Antimicrobial Peptides in Polymicrobial Biofilm-Associated Infections

“…Unlike most antibiotics that may lose activity if their basic structure is modified, AMPs are particularly prone to molecular alteration. They can be optimized by altering their primary sequence through the incorporation or deletion of hydrophobic or charged amino acids, which has been shown to affect their selectivity for Gram-positive, Gram-negative or fungal membranes [ 113 , 114 , 115 , 116 ]. The killing of specific pathogens within a mixed bacterial population has been achieved in vitro by AMPs, referred to as specifically targeted antimicrobial peptides (STAMPs).…”