The mechanisms of action of water-soluble aminohexanoic and malonic adducts of fullerene C60 with hexamethonium on model lipid membranes

Pneumolysin is a member of the cholesterol-dependent cytolysin (CDC) family of pore-forming proteins that are produced as water-soluble monomers or dimers, bind to target membranes and oligomerize into large ring-shaped assemblies comprising approximately 40 subunits and approximately 30 nm across. This pre-pore assembly then refolds to punch a large hole in the lipid bilayer. However, in addition to forming large pores, pneumolysin and other CDCs form smaller lesions characterized by low electrical conductance. Owing to the observation of arc-like (rather than full-ring) oligomers by electron microscopy, it has been hypothesized that smaller oligomers explain smaller functional pores. To investigate whether this is the case, we performed cryo-electron tomography of pneumolysin oligomers on model lipid membranes. We then used sub-tomogram classification and averaging to determine representative membrane-bound low-resolution structures and identified pre-pores versus pores by the presence of membrane within the oligomeric curve. We found pre-pore and pore forms of both complete (ring) and incomplete (arc) oligomers and conclude that arc-shaped oligomeric assemblies of pneumolysin can form pores. As the CDCs are evolutionarily related to the membrane attack complex/perforin family of proteins, which also form variably sized pores, our findings are of relevance to that class of proteins as well.

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The Mechanisms of Action of Triindolylmethane Derivatives on Lipid Membranes

Acta Naturae ◽

10.32607/20758251-2019-11-3-38-45 ◽

2019 ◽

Vol 11 (3) ◽

pp. 38-45 ◽

Cited By ~ 2

Author(s):

S. S. Efimova ◽

T. E. Tertychnaya ◽

S. N. Lavrenov ◽

O. S. Ostroumova

Keyword(s):

Lipid Bilayers ◽

Lipid Membranes ◽

Antibacterial Agents ◽

Membrane Lipids ◽

Mechanisms Of Action ◽

Electrical Stability ◽

Membrane Breakdown ◽

Model Lipid Membranes ◽

Transmembrane Pores ◽

The Difference

The effects of new synthetic antibacterial agents - tris(1-pentyl-1H-indol-3-yl)methylium chloride (LCTA-1975) and (1-(4-(dimethylamino)-2,5-dioxo-2,5-dihydro-1H-pyrrol-3-yl)-1H-indol-3-yl)bis(1-propyl- 1H-indol-3-yl)methylium chloride (LCTA-2701 - on model lipid membranes were studied. The ability of the tested agents to form ion-conductive transmembrane pores, influence the electrical stability of lipid bilayers and the phase transition of membrane lipids, and cause the deformation and fusion of lipid vesicles was investigated. It was established that both compounds exert a strong detergent effect on model membranes. The results of differential scanning microcalorimetry and measuring of the threshold transmembrane voltage that caused membrane breakdown before and after adsorption of LCTA-1975 and LCTA-2701 indicated that both agents cause disordering of membrane lipids. Synergism of the uncoupling action of antibiotics and the alkaloid capsaicin on model lipid membranes was shown. The threshold concentration of the antibiotic that caused an increase in the ion permeability of the lipid bilayer depended on the membrane lipid composition. It was lower by an order of magnitude in the case of negatively charged lipid bilayers than for the uncharged membranes. This can be explained by the positive charge of the tested agents. At the same time, LCTA-2701 was characterized by greater efficiency than LCTA-1975. In addition to its detergent action, LCTA-2701 can induce ion-permeable transmembrane pores: step-like current fluctuations corresponding to the opening and closing of individual ion channels were observed. The difference in the mechanisms of action might be related to the structural features of the antibiotic molecules: in the LCTA-1975 molecule, all three substituents at the nitrogen atoms of the indole rings are identical and represent n-alkyl (pentyl) groups, while LCTA-2701 contains a maleimide group, along with two alkyl substituents (n-propyl). The obtained results might be relevant to our understanding of the mechanism of action of new antibacterial agents, explaining the difference in the selectivity of action of the tested agents on the target microorganisms and their toxicity to human cells. Model lipid membranes should be used in further studies of the trends in the modification and improvement of the structures of new antibacterial agents.

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