Incomplete pneumolysin oligomers form membrane pores.
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.
Division of Structural Biology, Wellcome Trust Centre for Human Genetics, University of Oxford, Roosevelt Drive, Oxford OX3 7BN, UK.
Andreas F-P
AF
Sonnen
Jürgen M
JM
Plitzko
Robert J C
RJ
Gilbert
D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D001427:Bacterial Toxins / Q000737:chemistry / Q000378:metabolism; D002784:Cholesterol / Q000737:chemistry; D008051:Lipid Bilayers / Q000737:chemistry / Q000378:metabolism*; D008081:Liposomes / Q000737:chemistry / Q000378:metabolism; D046529:Microscopy, Electron, Transmission; D010755:Organophosphates / Q000737:chemistry; D010713:Phosphatidylcholines / Q000737:chemistry; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D013301:Streptolysins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
D001426:Bacterial Proteins; D001427:Bacterial Toxins; D008051:Lipid Bilayers; D008081:Liposomes; D010755:Organophosphates; D010713:Phosphatidylcholines; D011994:Recombinant Proteins; D013301:Streptolysins; C032290:plY protein, Streptococcus pneumoniae; C002290:dicetylphosphate; D002784:Cholesterol
cryo-electron tomography; pneumolysin; pore formation; proteolipid pore; toroidal pore