Channel-forming properties of cecropins and related model compounds incorporated into planar lipid membranes.

TLDR

Cecropins and synthetic analogs form large, time‑variant, voltage‑dependent ion channels in planar lipid membranes at physiological concentrations, a property that aligns with the known insensitivity of eukaryotic cells to these peptides. A comparative study of cecropins A, B, D and six synthetic analogs was conducted to determine structural requirements for pore formation. The study found that cecropins and certain analogs form large (≈4 nm) voltage‑dependent channels with conductances up to 2.5 nS, a Cl⁻/Na⁺ selectivity of 2:1 for AD and MP3, and that a flexible linker between amphipathic and hydrophobic regions is essential; shorter peptides failed to form channels, while AD was the most effective pore‑former and antibacterial agent, with conductance reduced ≥5‑fold by positive surface charge or cholesterol, supporting the hypothesis that broad antibacterial activity arises from large pore formation.

Abstract

Cecropins, positively charged antibacterial peptides found in the cecropia moth, and synthetic peptide analogs form large time-variant and voltage-dependent ion channels in planar lipid membranes in the physiological range of concentration. Single-channel conductances of up to 2.5 nS (in 0.1 M NaCl) were observed, which suggests a channel diameter of 4 nm. Channels formed by the peptides cecropin AD and MP3 had a permeability ratio of Cl-/Na+ = 2:1 in 0.1 M NaCl. A comparative study of the three cecropins, cecropins A, B, and D, and of six synthetic analogs allowed determination of structural requirements for pore formation. Shorter amphipathic peptides did not form channels, although they adsorbed to the bilayer. A flexible segment between the N-terminal amphipathic region and the C-terminal more hydrophobic region of the peptide was required for the observation of a time-variant, voltage-dependent conductance. Cecropin AD was the most effective voltage-dependent pore-forming peptide and was also the most potent antibacterial peptide against several test organisms. A positive surface charge or cholesterol in the bilayer reduced the conductances caused by cecropin AD or MP3 by at least 5-fold. This behavior is consistent with the known insensitivity of eukaryotic cells to cecropins. Our observations suggest that the broad antibacterial activity of cecropins is due to formation of large pores in bacterial cell membranes.