X-ray crystallography shines a light on pore-forming toxins

Johnstone, Bronte A.; Christie, Michelle P.; Morton, Craig J.; Parker, Michael W.

doi:10.1016/bs.mie.2021.01.001

Cited by 11 publications

(12 citation statements)

References 214 publications

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“…Further, the moderate throughput enables statistical power to quantify individual processes within the full assembly pathway. In conjunction with kinetic modelling, this analysis represents a useful method to predict and measure the effects of modification to specific steps in the pathway, with applications in drug development or biotechnology ( Johnstone et al, 2021 ).…”

Section: Discussionmentioning

confidence: 99%

“…Pore-forming proteins (PFPs) possess an ancient and ubiquitous mechanism for forming aqueous channels in the membranes surrounding cells and organelles ( Dal Peraro and van der Goot, 2016 ; Johnstone et al, 2021 ). The largest and most sequence diverse class of PFPs is the membrane attack complex-perforin (MACPF)/cholesterol dependent cytolysin (CDC) superfamily with thousands of members now identified ( Christie et al, 2018 ; Dunstone and Tweten, 2012 ; Rosado et al, 2008 ).…”

Section: Introductionmentioning

confidence: 99%

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Single-molecule analysis of the entire perfringolysin O pore formation pathway

Guinness

Walsh

Bayly-Jones

et al. 2022

eLife

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The cholesterol-dependent cytolysin perfringolysin O (PFO) is secreted by Clostridium perfringens as a bacterial virulence factor able to form giant ring-shaped pores that perforate and ultimately lyse mammalian cell membranes. To resolve the kinetics of all steps in the assembly pathway, we have used single-molecule fluorescence imaging to follow the dynamics of PFO on dye-loaded liposomes that lead to opening of a pore and release of the encapsulated dye. Formation of a long-lived membrane-bound PFO dimer nucleates the growth of an irreversible oligomer. The growing oligomer can insert into the membrane and open a pore at stoichiometries ranging from tetramers to full rings (~35-mers), whereby the rate of insertion increases linearly with the number of subunits. Oligomers that insert before the ring is complete continue to grow by monomer addition post insertion. Overall, our observations suggest that PFO membrane insertion is kinetically controlled.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Single-molecule analysis of the entire perfringolysin O pore formation pathway

Guinness

Walsh

Bayly-Jones

et al. 2022

eLife

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“…The cryo-EM analysis of CPB clearly showed that like other members of the α-hemolysin, the CPB protomer is composed of a cap, a rim, and a stem domain (Johnstone et al , 2021). In addition to the prototypical features of the family members, CPB possesses a second β-barrel on top of the cap domain.…”

Section: Resultsmentioning

confidence: 99%

“…Based on sequence homology between CPB and other bacterial toxins (Supplementary Table 1 and Supplementary Fig. 1), the toxin is a member of the S. aureus αhemolysin family of β-PFTs 4 . Here we describe the cryo-EM structure of CPB in styrene maleic anhydride (SMA) discs, which represents the membrane-inserted pore form, at near atomic resolution.…”

mentioning

confidence: 99%

Cryo-EM structure of the octameric pore of Clostridium perfringens β-toxin

Bruggisser

Iacovache

Musson

et al. 2021

Preprint

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We describe the cryo-EM structure of Clostridium perfringens β-toxin (CBP) in styrene maleic acid (SMA) discs, which represents the membrane-inserted pore form, at near atomic resolution. We show that CPB forms an octamer, which though having a similar conformation to the hetero-oligomeric pores of bicomponent leukocidins, features a different receptor binding region and a novel N-terminal β-barrel. The latter contains an additional selectivity filter and creates a bipolar pore. We propose that the N-terminal β-barrel domain may regulate oligomerization and solubility of the complex and influence channel conductance and monomer stability. In addition, we show that the β-barrel protrusion domain can be modified or exchanged without affecting the pore forming ability, thus making the pore particularly attractive for macromolecule sensing and nanotechnology. The cryo-EM structure of the octameric pore of CPB will facilitate future developments in both nanotechnology and basic research.

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“…Our understanding of the structure of secreted, monomeric CDCs is comprehensive, owing primarily to the contribution of X‐ray crystallographic studies. The first CDC crystal structure was perfringolysin O (PFO), 8 now considered to be the archetypical CDC as subsequent CDC structures revealed high structural similarity 9 . PFO contains a four domain architecture (D1–D4) with D1, D2, and D4 arranged in a linear manner to reveal an elongated molecule that is rich in β‐sheet (Figure 1).…”

Section: Structural Features Of Secreted Cdcsmentioning

confidence: 99%

Cholesterol‐dependent cytolysins: The outstanding questions

et al. 2022

Self Cite

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The cholesterol-dependent cytolysins (CDCs) are a major family of bacterial pore-forming proteins secreted as virulence factors by Gram-positive bacterial species. CDCs are produced as soluble, monomeric proteins that bind specifically to cholesterol-rich membranes, where they oligomerize into ring-shaped pores of more than 30 monomers. Understanding the details of the steps the toxin undergoes in converting from monomer to a membrane-spanning pore is a continuing challenge. In this review we summarize what we know about CDCs and highlight the remaining outstanding questions that require answers to obtain a complete picture of how these toxins kill cells. K E Y W O R D S CD59, cholesterol-binding protein, cholesterol-dependent cytolysin, intermedilysin, MACPF, membrane-protein interactions, perfringolysin O, pneumolysin, pore-forming toxin 1 | INTRODUCTION Pore-forming toxins (PFTs), bacterial toxins that act via membrane permeabilization, are typically produced to aid bacterial virulence or survival. Cholesterol-dependent cytolysins (CDCs) are a large family of PFTs observed in numerous Gram-positive bacterial genera including Streptococcus, Clostridium, Listeria, and Gardnerella.

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X-ray crystallography shines a light on pore-forming toxins

Cited by 11 publications

References 214 publications

Single-molecule analysis of the entire perfringolysin O pore formation pathway

Single-molecule analysis of the entire perfringolysin O pore formation pathway

Cryo-EM structure of the octameric pore of Clostridium perfringens β-toxin

Cholesterol‐dependent cytolysins: The outstanding questions

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