Antimicrobial peptides in toroidal and cylindrical pores

Abstract

Antimicrobial peptides (AMPs) are small, usually cationic peptides, which permeabilize biological membranes. Their mechanism of action is still not well understood. Here we investigate the preference of alamethicin and melittin for pores of different shapes, using molecular dynamics (MD) simulations of the peptides in pre-formed toroidal and cylindrical pores. When an alamethicin hexamer is initially embedded in a cylindrical pore, at the end of the simulation the pore remains cylindrical or closes if glutamines in the N-termini are not located within the pore. On the other hand, when a melittin tetramer is embedded in toroidal pore or in a cylindrical pore, at the end of the simulation the pore is lined both with peptides and lipid headgroups, and, thus, can be classified as a toroidal pore. These observations agree with the prevailing views that alamethicin forms barrel-stave pores whereas melittin forms toroidal pores. Both alamethicin and melittin form amphiphilic helices in the presence of membranes, but their net charge differs; at pH approximately 7, the net charge of alamethicin is -1 whereas that of melittin is +5. This gives rise to stronger electrostatic interactions of melittin with membranes than those of alamethicin. The melittin tetramer interacts more strongly with lipids in the toroidal pore than in the cylindrical one, due to more favorable electrostatic interactions.

Figure 1

Snapshots from MD simulations of a melittin monomer started from a toroidal pore (A), a melittin monomer started from a cylindrical pore (B), an alamethicin monomer started from a toroidal pore (C), and an alamethicin monomer started from a cylindrical pore (D). The green balls represent the phosphocholines; the red balls represent ions; the blue balls represent water; the nonpolar residues are shown in black, the polar residues are yellow and the charged residues are magenta; the lipid tails have been removed for clarity.

Figure 2

Snapshots from MD simulations of a melittin tetramer started from a toroidal pore (A) and started from a cylindrical pore (B). The green balls represent the phosphocholines; the red balls represent Cl⁻ ions; the gray lines represent lipid tails; the blue balls represent water.

Figure 3

The interaction energy between a melittin tetramer and Cl⁻ ions versus time. ‘Cylinder’ refers to the MD simulation of melittin started from a cylindrical pore; ‘torus’ refers to the simulation started from a toroidal pore. The energies are averaged over 1 ns.

Figure 4

Snapshots from MD simulations of an alamethicin tetramer started from a toroidal pore (A) and an alamethicin hexamer started from a cylindrical pore: closed pore (B) and wet pore (C). The green balls represent the phosphocholines; the red balls represent K⁺ ions; the gray lines represent lipid tails; the blue balls represent water; in (C), the peptide are shown as ribbons and the lipid tails are removed for clarity.

Figure 5

The total, electrostatic (elec) or van der Waals (vdw) interaction energy between a melittin tetramer and the lipid headgroups (hg) or tails (lt) in toroidal pores (torus) or in constrained cylindrical pores (cyl). Error bars are the standard deviation. The Welch two sample t-test was used to determine significant differences between the peptide-lipid interaction energies in the constrained cylindrical pore and in the toroidal pore; in all the cases p value is less than 0.05.

Figure 6

The electrostatic interaction energy between polar residues of a melittin tetramer and lipid headgroups. ‘N-polar’ refers to K7, T10 and T11; ‘C-polar’ refers to S18, W19, K21, R22, K23, R24, Q25 and Q26; ‘cyl_cons’ refers to the MD simulation in a constrained cylindrical pore; ‘torus’ refers to the MD simulation started from a toroidal pore; ‘cyl_nocons’ refers to the MD simulation started from a cylindrical pore. The Welch two sample t-test was used to determine significant differences between the polar residue-lipid headgroup interaction energies in the constrained cylindrical pore and in the toroidal or the initially cylindrical pore; in all the cases p value is less than 0.05.

Figure 7

Polar and charged residues of a melittin tetramer interacting with lipid headgroups and Cl⁻ ions in a constrained cylindrical pore (A) and toroidal pores (B and C). Snapshots (B) and (C) are from the MD simulations started from a cylindrical pore and a toroidal pore, respectively. Water molecules are removed for clarity. Polar residues are shown as yellow licorice, charged residues as blue licorice, W19 as black licorice, phosphocholines as green balls, ions as red balls, lipid tails as gray lines, and peptides as blue, orange, yellow and purple ribbons.

Figure 8

The peptide-peptide electrostatic interactions versus time. The interaction energies are calculated for melittin tetramer embedded into a cylindrical pore and averaged over 1ns; mlt1, mlt2, mlt3 and mlt4 denote monomers 1 through 4.