Structural analysis of the protein/lipid complexes associated with pore formation by the bacterial toxin pneumolysin.
Pneumolysin, a major virulence factor of the human pathogen Streptococcus pneumoniae, is a soluble protein that disrupts cholesterol-containing membranes of cells by forming ring-shaped oligomers. Magic angle spinning and wideline static (31)P NMR have been used in combination with freeze-fracture electron microscopy to investigate the effect of pneumolysin on fully hydrated model membranes containing cholesterol and phosphatidylcholine and dicetyl phosphate (10:10:1 molar ratio). NMR spectra show that the interaction of pneumolysin with cholesterol-containing liposomes results in the formation of a nonbilayer phospholipid phase and vesicle aggregation. The amount of the nonbilayer phase increases with increasing protein concentration. Freeze-fracture electron microscopy indicates the coexistence of aggregated vesicles and free ring-shaped structures in the presence of pneumolysin. On the basis of their size and analysis of the NMR spectra it is concluded that the rings are pneumolysin oligomers (containing 30-50 monomers) complexed with lipid (each with 840-1400 lipids). The lifetime of the phospholipid in either bilayer-associated complexes or free pneumolysin-lipid complexes is > 15 ms. It is further concluded that the effect of pneumolysin on lipid membranes is a complex combination of pore formation within the bilayer, extraction of lipid into free oligomeric complexes, aggregation and fusion of liposomes, and the destabilization of membranes leading to formation of small vesicles.
Bonev
B B
BB
Biomembrane Structure Unit, Department of Biochemistry, University of Oxford, South Parks Road, Oxford OX1 3QU, United Kingdom.
Gilbert
R J
RJ
Andrew
P W
PW
Byron
O
O
Watts
A
A
D001426:Bacterial Proteins; D002784:Cholesterol; D003931:Diacetyl / Q000031:analogs & derivatives*; D008081:Liposomes; D008566:Membranes / Q000187:drug effects*; D008961:Models, Structural; D019906:Nuclear Magnetic Resonance, Biomolecular; D009943:Organophosphorus Compounds; D010713:Phosphatidylcholines; D010759:Phosphorus Isotopes; D011510:Proteolipids / Q000648:ultrastructure*; D013296:Streptococcus pneumoniae / Q000472:pathogenicity; D013301:Streptolysins / Q000494:pharmacology*
D001426:Bacterial Proteins; D008081:Liposomes; D009943:Organophosphorus Compounds; D010713:Phosphatidylcholines; D010759:Phosphorus Isotopes; D011510:Proteolipids; D013301:Streptolysins; C032290:plY protein, Streptococcus pneumoniae; C005146:diacetyl phosphate; D002784:Cholesterol; D003931:Diacetyl