Insight into the mechanism of action of temporin-SHa, a new broad-spectrum antiparasitic and antibacterial agent

Abstract

Antimicrobial peptides (AMPs) are promising drugs to kill resistant pathogens. In contrast to bacteria, protozoan parasites, such as Leishmania, were little studied. Therefore, the antiparasitic mechanism of AMPs is still unclear. In this study, we sought to get further insight into this mechanism by focusing our attention on temporin-SHa (SHa), a small broad-spectrum AMP previously shown to be active against Leishmania infantum. To improve activity, we designed analogs of SHa and compared the antibacterial and antiparasitic mechanisms. [K3]SHa emerged as a highly potent compound active against a wide range of bacteria, yeasts/fungi, and trypanosomatids (Leishmania and Trypanosoma), with leishmanicidal intramacrophagic activity and efficiency toward antibiotic-resistant strains of S. aureus and antimony-resistant L. infantum. Multipassage resistance selection demonstrated that temporins-SH, particularly [K3]SHa, are not prone to induce resistance in Escherichia coli. Analysis of the mode of action revealed that bacterial and parasite killing occur through a similar membranolytic mechanism involving rapid membrane permeabilization and depolarization. This was confirmed by high-resolution imaging (atomic force microscopy and field emission gun-scanning electron microscopy). Multiple combined techniques (nuclear magnetic resonance, surface plasmon resonance, differential scanning calorimetry) allowed us to detail peptide-membrane interactions. [K3]SHa was shown to interact selectively with anionic model membranes with a 4-fold higher affinity (KD = 3 x 10−8 M) than SHa. The amphipathic α-helical peptide inserts in-plane in the hydrophobic lipid bilayer and disrupts the acyl chain packing via a detergent-like effect. Interestingly, cellular events, such as mitochondrial membrane depolarization or DNA fragmentation, were observed in L. infantum promastigotes after exposure to SHa and [K3]SHa at concentrations above IC50. Our results indicate that these temporins exert leishmanicidal activity via a primary membranolytic mechanism but can also trigger apoptotis-like death. The many assets demonstrated for [K3]SHa make this small analog an attractive template to develop new antibacterial/antiparasitic drugs.

Synthesis of breast cancer targeting conjugate of temporin-SHa analog and its effect on pro- and anti-apoptotic protein expression in MCF-7 cells

“…substituted temporin-[A2,6,9]SHa were not found cytotoxic against mammalian cell lines such as THP-1 monocytes (IC50>60 μM) and HepG2 cells (IC50 >600 μM) [23]. We synthesized various D-Ala substituted analogs of temporin-SHa and screened them against several cancer cell lines as well as 3T3 cells to identify analogs with anticancer activity.…”

“…Many antimicrobial peptides were reported to possess anticancer activities. However, the antimicrobial peptide temporin-SHa was reported as non-cytotoxic against the HepG2 cell line [23]. We decided to perform anticancer screening of temporin-SHa and its analogs ( (Fig.…”

“…As a proof of concept, we started with the antimicrobial peptide temporin-SHa (FLSGIVGMLGKLFamide). Temporin-SHa was isolated from the skin of the Sahara frog Pelophylax saharicus and reported as an antimicrobial agent active against bacteria, yeasts and fungi [22,23]. Temporin-SHa, the analogs temporin-[S3K]SHa and the L-alanine…”

The role of d - allo -isoleucine in the deposition of the anti- Leishmania peptide bombinin H4 as revealed by 31 P solid-state NMR, VCD spectroscopy, and MD simulation

“…Because temporin [K 3 ]SHa is possible to initially stronger interact with membrane due to presence of third lysine residue, it displays more efficient and faster killing ability than SHa [64]. Moreover, H4 showed slightly faster reached the plateau of depolarization compared to H2 on Leishmania promastigotes [20].…”

Rational Design of Cyclic Antimicrobial Peptides Based on BPC194 and BPC198

“…They are short, cationic sequences that contain up to 50% of hydrophobic residues and are able to adopt an amphipathic structure. These peptides display a broad spectrum of activity, selectivity towards microbial targets, and a low frequency in developing microbial resistance [ 9 , 10 , 11 , 12 , 13 , 14 , 15 , 16 ]. Their unique mechanism of action is the basis for these properties.…”