Cholesterol microcrystals and cochleate cylinders: Attachment of pyolysin oligomers and domain 4

Harris, J. Robin; Lewis, Richard J.; Baik, Clara; Pokrajac, Lisa A.; Billington, Stephen J.; Palmer, Michaël

doi:10.1016/j.jsb.2010.07.010

Cited by 12 publications

(10 citation statements)

References 59 publications

(75 reference statements)

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“…D4 yield rings and arcs as expected for PLO-WT alone, as well as linear stripes as expected for PLO-D4 alone (Harris et al, 2011). However, occasionally, the two shapes are seen to combine into walkingcane like formations ( Figure 5C and D).…”

Section: Morphology Of Hybrid Oligomers On Crystalline Cholesterol Ssupporting

confidence: 61%

“…Cholesterol has been shown to induce oligomerization of streptolysin O (Duncan and Schlegel, 1975), and cholesterol crystals have been established as a useful model system to study the oligomerization of CDCs by EM Harris et al, 2011). On such crystals, intact wild type PLO forms the usual ring and arc structures ( Figure 5A) that have also been observed with many other CDCs.…”

Section: Morphology Of Plo-d4 Oligomersmentioning

confidence: 99%

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Oligomerization and hemolytic properties of the C-terminal domain of pyolysin, a cholesterol-dependent cytolysin

Pokrajac

Harris

Sarraf

et al. 2013

Biochem. Cell Biol.

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Pyolysin (PLO) belongs to the homologous family of the cholesterol-dependent cytolysins (CDCs), which bind to cell membranes containing cholesterol to form oligomeric pores of large size. The CDC monomer structure consists of four domains. Among these, the C-terminal domain 4 has been implicated in membrane binding of the monomer, while the subsequent processes of oligomerization and membrane insertion have primarily been assigned to other domains of the molecule.Recombinantly expressed or proteolytic fragments that span the domains 4 of the CDCs streptolysin O (SLO) and perfringolysin O (PFO) bind to membranes but fail to oligomerize, and they inhibit the activity of the respective wild type toxins. We here report that the isolated domain 4 of pyolysin (PLO-D4) not only binds to membranes but also forms oligomers with itself, as well as hybrid oligomers with the full-length toxin. As expected, the pure PLO-D4 oligomers are devoid of pore-forming activity.Surprisingly, however, within hybrid oligomers, PLO-D4 not only fails to inhibit, but even amplifies the hemolytic activity of the full-length toxin, to an extent similar to that of doubling the amount of the full-length toxin alone. We propose that this amplification may be related to the kinetics of the oligomerization reaction. Overall, our findings indicate a greater role of domain 4 in the oligomerization of CDCs than previously demonstrated.

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Section: Morphology Of Hybrid Oligomers On Crystalline Cholesterol Ssupporting

confidence: 61%

Section: Morphology Of Plo-d4 Oligomersmentioning

confidence: 99%

Oligomerization and hemolytic properties of the C-terminal domain of pyolysin, a cholesterol-dependent cytolysin

Pokrajac

Harris

Sarraf

et al. 2013

Biochem. Cell Biol.

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“…In contrast to the intertwined domains D1-D3, domain D4 is a separate folding unit with a compact b-sandwich structure. Refolded D4 of SLO and pyolysin (PLO) have been found to bind to membranes 41 and to form linear arrays on cholesterol crystals 42 . To examine whether D4 binds to and oligomerizes on the membrane in a similar way, we expressed and purified LLO domain D4 and incubated ghosts with the purified protein.…”

Section: Resultsmentioning

confidence: 99%

“…PLO D4 also is able to bind to membranes 43 and to form straight arrays on cholesterol crystals 42 . In contrast to SLO D4, addition of PLO D4 to the full-length protein amplifies its haemolytic activity, whereas added LLO D4 does not, indicating that this domain probably does not integrate into the oligomers.…”

Section: Discussionmentioning

confidence: 99%

Crystal structure of listeriolysin O reveals molecular details of oligomerization and pore formation

et al. 2014

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Listeriolysin O (LLO) is an essential virulence factor of Listeria monocytogenes that causes listeriosis. Listeria monocytogenes owes its ability to live within cells to the pH-and temperature-dependent pore-forming activity of LLO, which is unique among cholesteroldependent cytolysins. LLO enables the bacteria to cross the phagosomal membrane and is also involved in activation of cellular processes, including the modulation of gene expression or intracellular Ca 2 þ oscillations. Neither the pore-forming mechanism nor the mechanisms triggering the signalling processes in the host cell are known in detail. Here, we report the crystal structure of LLO, in which we identified regions important for oligomerization and pore formation. Mutants were characterized by determining their haemolytic and Ca 2 þ uptake activity. We analysed the pore formation of LLO and its variants on erythrocyte ghosts by electron microscopy and show that pore formation requires precise interface interactions during toxin oligomerization on the membrane.

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“…It has however been shown that D4 of streptolysin, pyolysin and LLO can oligomerise by itself on cholesterol crystals (Harris et al, 2011; Weis and Palmer, 2001) or erythrocyte ghosts (Köster et al, 2014). Our cryoEM structure confirms that D4 can indeed play a role in oligomer formation through intra- and intermolecular interactions of its loops (Figure 2—figure supplements 2 and 3).…”

Section: Discussionmentioning

confidence: 99%

CryoEM structures of membrane pore and prepore complex reveal cytolytic mechanism of Pneumolysin

Pee

Neuhaus

D’Imprima

et al. 2017

eLife

128

168

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Many pathogenic bacteria produce pore-forming toxins to attack and kill human cells. We have determined the 4.5 Å structure of the ~2.2 MDa pore complex of pneumolysin, the main virulence factor of Streptococcus pneumoniae, by cryoEM. The pneumolysin pore is a 400 Å ring of 42 membrane-inserted monomers. Domain 3 of the soluble toxin refolds into two ~85 Å β-hairpins that traverse the lipid bilayer and assemble into a 168-strand β-barrel. The pore complex is stabilized by salt bridges between β-hairpins of adjacent subunits and an internal α-barrel. The apolar outer barrel surface with large sidechains is immersed in the lipid bilayer, while the inner barrel surface is highly charged. Comparison of the cryoEM pore complex to the prepore structure obtained by electron cryo-tomography and the x-ray structure of the soluble form reveals the detailed mechanisms by which the toxin monomers insert into the lipid bilayer to perforate the target membrane.DOI: http://dx.doi.org/10.7554/eLife.23644.001

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Cholesterol microcrystals and cochleate cylinders: Attachment of pyolysin oligomers and domain 4

Cited by 12 publications

References 59 publications

Oligomerization and hemolytic properties of the C-terminal domain of pyolysin, a cholesterol-dependent cytolysin

Oligomerization and hemolytic properties of the C-terminal domain of pyolysin, a cholesterol-dependent cytolysin

Crystal structure of listeriolysin O reveals molecular details of oligomerization and pore formation

CryoEM structures of membrane pore and prepore complex reveal cytolytic mechanism of Pneumolysin

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