Amphipathic α-helical peptide, HP (2–20), and its analogues derived from Helicobacter pylori: Pore formation mechanism in various lipid compositions

Abstract

In a previous study, we determined that HP(2-20) (residues 2-20 of parental HP derived from the N-terminus of Helicobacter pylori Ribosomal Protein L1) and its analogue, HPA3, exhibit broad-spectrum antimicrobial activity. The primary objective of the present study was to gain insight into the relevant mechanisms of action using analogues of HP(2-20) together with model liposomes of various lipid compositions and electron microscopy. We determined that these analogues, HPA3 and HPA3NT3, exert potent antibacterial effects in low-salt buffer and antifungal activity against chitin-containing fungi, while having little or no hemolytic activity or cytotoxicity against mammalian cell lines. Our examination of the interaction of HP(2-20) and its analogues with liposomes showed that the peptides disturb both neutral and negatively-charged membranes, as demonstrated by the release of encapsulated fluorescent markers. The release of fluorescent markers induced by HP(2-20) and its analogues was inversely related to marker size. The pore created by HP(2-20) shows that the radius is approximately 1.8 nm, whereas HPA3, HPA3NT3, and melittin have apparent radii between 3.3 and 4.8 nm. Finally, as shown by electron microscopy, the liposomes and various microbial cells treated with HPA3 and HPA3NT3 showed oligomerization and blebbing similar to that seen with melittin, while HP(2-20) exhibited flabbiness. These results suggest that HP(2-20) may exert its antibiotic effects through a small pore (about 1.8 nm), whereas HPA3 and HPA3NT3 formed pores of a size consistent with those formed by melittin.

Comparative functional properties of engineered cationic antimicrobial peptides consisting exclusively of tryptophan and either lysine or arginine

“…A common antimicrobial mechanism of helical AMPs is the ability to recognize their bacterial targets by electrostatic interactions and to permeabilize the cell membrane (Jelokhani-Niaraki et al, 1998;Jean-François et al, 2008;Park et al, 2008). Based on this understanding, we sought to compare the shortest eCAPs with optimal activity, WK12 and WR12, for interactions with live P. aeruginosa using SPR spectroscopy and flow cytometry (Lequin et al, 2006).…”

Novel chimeric peptide with enhanced cell specificity and anti-inflammatory activity

“…The hemolytic activity against fresh rat red blood cells (rRBCs) and the cytotoxic activity against HaCaT (human keratinocyte) cells were examined using the method described previously [27,40].…”

“…To design a novel model AMP, the N-terminus of HPA3NT3 (1e7) [27] and the C-terminus of melittin (17e26) were combined as a chimera form (Hn-Mc, Fig. 1A).…”

“…The supernatant filters were then treated with 1% (w/v) osmium tetroxide, washed in 5% (w/v) sucrose in cacodylate buffer, and then dehydrated with a graded series of ethanol. After lyophilization and gold coating, the samples were observed by SEM (JSM-7100F, JEOL, Tokyo, Japan)[27].…”

The Functional Role of the Tachykinin Consensus Region of Urechistachykinin Peptide Family for Its Antimicrobial Activity

“…The supernatant was recovered, suspended in potassium phosphate buffer and fluorescence intensity recorded (excitation 495 nm, emission 520 nm). 10) RESULTS AND DISCUSSION Design of Analogues and Their Structures, Antimicrobial and Hemolytic Activity U I and II are very short peptides with only 10-residue and showed significant antimicrobial activities. 7) U II has the conserved amino acids Phe-6, Gly-8 and Arg-10 in the sequence, with other urechistachykinin peptide families (U III-VII) (Fig.…”