Investigating the effects of L- to D-amino acid substitution and deamidation on the activity and membrane interactions of antimicrobial peptide anoplin

Abstract

Isolated from the venom sac of solitary spider wasp, Anoplius samariensis, anoplin is the smallest linear α-helical antimicrobial peptide found naturally with broad spectrum activity against both Gram-positive and Gram-negative bacteria, and little hemolytic activity toward human erythrocytes. Deamidation was found to decrease the peptide's antibacterial properties. In the present work, interactions of amidated (Ano-NH2) and deamidated (Ano-OH) forms of anoplin as well as Ano-NH2 composed of all D-amino acids (D-Ano-NH2) with model cell membranes were investigated by means of Langmuir Blodgett (LB) technique, atomic force microscopy (AFM), X-ray photoemission electron microscopy (X-PEEM) and carboxyfluorescein leakage assay in order to gain a better understanding of the effect of these peptide modifications on membrane binding and lytic properties. According to LB, all three peptides form stable monolayers at the air/water interface with Ano-NH2 occupying a slightly greater area per molecule than Ano-OH. All three forms of the peptide interact preferentially with anionic 1,2-dipalmitoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] (DPPG), rather than zwitterionic 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) lipid monolayer. Peptides form nanoscale clusters in zwitterionic but not in anionic monolayers. Finally, membrane lytic activity of all derivatives was found to depend strongly on membrane composition and lipid/peptide ratio. The results suggest that amidated forms of peptides are likely to possess higher membrane binding affinity due to the increased charge.

Design of novel analogues of short antimicrobial peptide anoplin with improved antimicrobial activity

“…Understanding how to control the interrelationships between these parameters will contribute to designing antimicrobial peptides with high therapeutic indices. Recently, structure and activity relationship studies indicated that anoplin is very sensitive to small modifications in primary structure [21][22][23]. Ifrah and co-worker reported that Ano-1K5V8K (replacement of glycine by lysine, arginine by valine, and threonine by lysine at positions 1, 5, and 8, respectively) exhibited high antimicrobial activity and low hemolytic activity.…”

Latarcins: versatile spider venom peptides

“…The balance between antimicrobial and hemolytic effects has been discussed in numerous works [116,[140][141][142][143][144][145][146][147][148][149][150][151]. The hemolytic effect is especially pronounced for Ltc 2a (Table 4).…”

Antimicrobial Activity of Purified Toxins from <i>Crossopriza lyoni</i> (Spider) against Certain Bacteria and Fungi

“…A single glycine to alanine substitution in antimicrobial peptide latarcin 2a increases interactions with lipid membrane [20]. Similarly, L-to D-amino acid substitution affects deamidation activity and membrane interactions [21]. From the inhibition zone diameter assays the IZDs obtained were larger in case of spider venom than antibiotics Similar action of spider toxins was observed in agar disc diffusion bioassay in which diameter of inhibition zones was E. coli 17 Table 2).…”