Shortened derivatives from native antimicrobial peptide LyeTx I: In vitro and in vivo biological activity assessment

Abstract

In the continuing search for novel antibiotics, antimicrobial peptides are promising molecules, due to different mechanisms of action compared to classic antibiotics and to their selectivity for interaction with microorganism cells rather than with mammalian cells. Previously, our research group has isolated the antimicrobial peptide LyeTx I from the venom of the spider Lycosa erythrognatha. Here, we proposed to synthesize three novel shortened derivatives from LyeTx I (LyeTx I mn; LyeTx I mnΔK; LyeTx I mnΔKAc) and to evaluate their toxicity and biological activity as potential antimicrobial agents. Peptides were synthetized by Fmoc strategy and circular dichroism analysis was performed, showing that the three novel shortened derivatives may present membranolytic activity, like the original LyeTx I, once they folded as an alpha helix in 2.2.2-trifluorethanol and sodium dodecyl sulfate. In vitro assays revealed that the shortened derivative LyeTx I mnΔK presents the best score between antimicrobial (↓ MIC) and hemolytic (↑ EC50) activities among the synthetized shortened derivatives, and LUHMES cell-based NeuriTox test showed that it is less neurotoxic than the original LyeTx I (EC50 [LyeTx I mnΔK] ⋙ EC50 [LyeTx I]). In vivo data, obtained in a mouse model of septic arthritis induced by Staphylococcus aureus, showed that LyeTx I mnΔK is able to reduce infection, as demonstrated by bacterial recovery assay (∼10-fold reduction) and scintigraphic imaging (less technetium-99m labeled-Ceftizoxime uptake by infectious site). Infection reduction led to inflammatory process and pain decreases, as shown by immune cells recruitment reduction and threshold nociception increment, when compared to positive control group. Therefore, among the three shortened peptide derivatives, LyeTx I mnΔK is the best candidate as antimicrobial agent, due to its smaller amino acid sequence and toxicity, and its greater biological activity.

Antimicrobial Peptide Analogs From Scorpions: Modifications and Structure-Activity

“…Furthermore, a study using artificial intelligence 100,000 peptides of which 200 were synthetized and tested; the results showed that a peptide shorter than usual, containing nine amino-acid residues, presented minimum inhibitory concentration (MIC) lower than 1 µM ( Cherkasov et al, 2009 ). Other studies using antimicrobial peptides derived from other animals also showed that shortened peptides produced better antimicrobial activity ( Solstad et al, 2020 ; Fuscaldi et al, 2021 ).…”

Non‐canonical amino acid bioincorporation into antimicrobial peptides and its challenges

“…This strategy has been useful in synthesising short derivatives of proteins. For instance, it has been used to produce short portions of spider venom as potential AMPs 177,178 . It has also aided in examining the effect of AA substitution on the efficacy and stability of small AMPs 179,180 .…”

Spider-Venom Peptides: Structure, Bioactivity, Strategy, and Research Applications

“…Nevertheless, the major problems of these conventional therapies were the relatively low success rate, serious side effects, and drug resistance [48][49][50]. Consequently, Lycotoxin I Lycosa carolinensis IWLTALKFLGKHAAKHLAKQQLSKL-NH 2 [24] Lycotoxin II Lycosa carolinensis KIKWFKTMKSIAKFIAKEQMKKHLGGE-OH [24] Lycocitin 1 Lycosa singoriensis GKLQAFLAKMKEIAAQTL-NH 2 [37] Lycocitin 2 Lycosa singoriensis GRLQAFLAKMKEIAAQTL-NH 2 [37] Ltc 1 Lachesana tarabaevi SMWSGMWRRKLKKLRNALKKKLKGE-OH [38] Ltc 2a Lachesana tarabaevi GLFGKLIKKFGRKAISYAVKKARGKH-OH [38] Ltc 3a Lachesana tarabaevi SWKSMAKKLKEYMEKLKQRA-NH 2 [38] Ltc 3b Lachesana tarabaevi SWASMAKKLKEYMEKLKQRA-NH 2 [38] Ltc 4a Lachesana tarabaevi GLKDKFKSMGEKLKQYIQTWKAKF-NH 2 [38] Ltc 4b Lachesana tarabaevi SLKDKVKSMGEKLKQYIQTWKAKF-NH 2 [38] Ltc 5 Lachesana tarabaevi GFFGKMKEYFKKFGASFKRRFANLKKRL-NH 2 [38] CIT 1a Lachesana tarabaevi GFFGNTWKKIKGKADKIMLKKAVKIMVKKEGISKEEAQAKVD AMSKKQIRLYLLKYYGKKALQKASEKL-OH [38] LyeTx I Lycosa erythrognatha IWLTALKFLGKNLGKHLAKQQLAKL-OH [39] Cupiennin 1a Cupiennius salei GFGALFKFLAKKVAKTVAKQAAKQGAKYVVNKQME-NH 2 [40] LyeTx I Lycosa erythrognatha IWLTALKFLGKNLGKHLAKQQLAKL-NH 2 [41] U1-SCRTX-Lg1a Loxosceles gaucho VGTDFSGNDDISDVQK-NH 2 [42] Lycotoxin-Pa4a Pardosa astrigera…”