Bacterial pore-forming toxins (Özkan Yildiz)


Figure 19. Membrane insertion of the pore-forming toxin pneumolysin from Streptococcus pneumoniae.

Gram-positive pathogens, such as Pneumococcus, Listeria and Clostridium, secrete cholesterol-dependent cytolysins (CDCs) as soluble monomers. The monomers attach to cholesterol-containing target membranes, where they assemble into large, circular pores that perforate the target cells. The structure and mechanism of pore-forming toxins are studied in the project group of Dr. Özkan Yildiz in the Department, who determined the x-ray structures of two CDCs in the soluble form: listeriolysin (LLO) from Listeria monocytogenes (Köster et al., Nat Comm 2014), and pneumolysin (PLY) from Streptococcus pneumoniae (van Pee et al., Nano Letters 2016). To understand the mechanism of pore formation and membrane perforation indetail (Figure 19 above), we determined the structure of the ~2.2 MDa pore complex of PLY by single-particle cryoEM at a resolution of 4.5 Å (Figure 20A). The PLY pore complex is a 400-Å ring of 42 membrane-inserted monomers. During membrane insertion, one of the four protein domains in the soluble monomer refolds into two ~85 Å β‑hairpins that traverse the lipid bilayer. In the membrane, the hairpins assemble into a cylindrical 168-stranded β‑barrel (Figure 20A,B insets). Large sidechains are resolved on the apolar outer barrel surface, while the inner barrel surface is highly charged (Figure 20B). CryoET shows that the soluble monomers assemble on chole­sterol-containing liposomes into multi-subunit, circular prepores, with residues that confer lipid specificity on the membrane surface. Tomographic volumes show that the lipid bilayer in the prepore is intact (Figure 20C,D), while it has disappeared in the mature pore (Figure 20E,F). Presumably the highly charged inner pore surface repels and helps to disperse any lipid trapped in the pore (van Pee et al., eLife 2017).

Figure 20. The pore-forming toxin pneumolysin from Streptococcus pneumoniae. (A) Single-particle cryo-EM map of PLY at 4.5 Å resolution with fitted atomic model based on the PLY x-ray structure (van Pee et al., 2016). Inset: refolded β-hairpins fitted to the 4.5 Å map. (B) A cross section through the pore complex structure reveals that the inner surface of the β-barrel is highly charged (nega­tive charges, red; positive charges, blue). Inset: section of membrane-inserted PLY at 4.5 Å resolution. (C,D) Subtomo­gram average of PLY prepores and (E,F) pores at 22 and 27 Å resolution, respectively. PLY domains are red (D1), yellow (D2), green/cyan (D3) and blue (D4). The lipid bilayer (gray) is intact in the prepore complex, but absent in the pore complex.

Contact:

Max Planck Institute of Biophysics

Prof. Dr. Werner Kühlbrandt, Director
Department of Structural Biology
Secretary: Monika Hobrack

Phone: +49 (0) 69 6303-3001
Fax: +49 (0) 69 6303-3002
E-mail: monika.hobrack(at)biophys.mpg.de