Temporin L and aurein 2.5 have identical conformations but subtly distinct membrane and antibacterial activities

Abstract

Frogs such as Rana temporaria and Litoria aurea secrete numerous closely related antimicrobial peptides (AMPs) as an effective chemical dermal defence. Damage or penetration of the bacterial plasma membrane is considered essential for AMP activity and such properties are commonly ascribed to their ability to form secondary amphipathic, α-helix conformations in membrane mimicking milieu. Nevertheless, despite the high similarity in physical properties and preference for adopting such conformations, the spectrum of activity and potency of AMPs often varies considerably. Hence distinguishing apparently similar AMPs according to their behaviour in, and effects on, model membranes will inform understanding of primary-sequence-specific antimicrobial mechanisms. Here we use a combination of molecular dynamics simulations, circular dichroism and patch-clamp to investigate the basis for differing anti-bacterial activities in representative AMPs from each species; temporin L and aurein 2.5. Despite adopting near identical, α-helix conformations in the steady-state in a variety of membrane models, these two AMPs can be distinguished both in vitro and in silico based on their dynamic interactions with model membranes, notably their differing conformational flexibility at the N-terminus, ability to form higher order aggregates and the characteristics of induced ion conductance. Taken together, these differences provide an explanation of the greater potency and broader antibacterial spectrum of activity of temporin L over aurein 2.5. Consequently, while the secondary amphipathic, α-helix conformation is a key determinant of the ability of a cationic AMP to penetrate and disrupt the bacterial plasma membrane, the exact mechanism, potency and spectrum of activity is determined by precise structural and dynamic contributions from specific residues in each AMP sequence.

A pleurocidin analogue with greater conformational flexibility, enhanced antimicrobial potency and in vivo therapeutic efficacy

“…4 and Supplementary Figs. [14][15][16][17][18][19][20][21][22][23][24][25][26][27][28][29]. By culturing either EMRSA-15 or P. aeruginosa RP73 in the presence or absence of each D-pleurocidin analogue, we aim to infer differences in their bactericidal strategy from the measures the bacteria take to overcome the challenge.…”

Novel temporin L antimicrobial peptides: promoting self-assembling by lipidic tags to tackle superbugs

“…The isoform L (temporin L, TL), H-Phe-Val-Gln-Trp-Phe-Ser-Lys-Phe-Leu-Gly-Arg-Ile-Leu-NH 2 , is a highly potent AMP with activity against both Gram-positive and Gram-negative bacteria 14 . Recent studies focussed on the mechanism of action of TL showed that hydrophobic residues, in particular the phenylalanine zipper motif, are involved in the initial peptide binding to the Gram-positive cytoplasmic membrane and the stabilisation of peptide aggregates inside the membrane, leading to the formation of large pores and consequential cell death 15 . In the Gram-negative bacterial membranes, the mechanism of permeabilisation appears to be different instead and may not involve membranolytic processes 15 .…”

“…Recent studies focussed on the mechanism of action of TL showed that hydrophobic residues, in particular the phenylalanine zipper motif, are involved in the initial peptide binding to the Gram-positive cytoplasmic membrane and the stabilisation of peptide aggregates inside the membrane, leading to the formation of large pores and consequential cell death 15 . In the Gram-negative bacterial membranes, the mechanism of permeabilisation appears to be different instead and may not involve membranolytic processes 15 . Unfortunately, TL also kills human erythrocytes at antimicrobial concentration, and earlier structure–activity relationship studies (SAR) showed that this drawback is correlated to its α-helical content 16–19 .…”

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

“…Interestingly, a recent study comparing the structure and function of temporin L and aurein 2.5 [124] found that, although both AMPs adopt α-helical structure, they disrupt membranes differently. The results presented for aurein 2.5 [124] mirror earlier studies [121,123], which showed that, as long as the residue at position 13 in aurein 2.2 is hydrophobic (i.e., L, I, V, A or F), the resulting peptides have similar structures and modes of action (i.e., as aurein 2.2, 2.3, and 2.5 do). The mechanism of action and structure of aurein 2.2 and aurein 2.3 have been extensively studied in recent years.…”