Structural and Mechanistic Features of ClyA-Like α-Pore-Forming Toxins

Bräuning, Bastian; Groll, M.

doi:10.3390/toxins10090343

Cited by 12 publications

(7 citation statements)

References 38 publications

(76 reference statements)

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“…In contrast, saturated phospholipids behave oppositely resulting in less availability of cholesterol in the membrane for the toxin to interact. The preferential partitioning of the two PFTs to the different membrane is due to the difference in binding motif and the conformational changes of ClyA and LLO [56,57]. Our diffusivity measurements (figure 2) from the spatially distinct cholesterol-rich and poor regimes of DOPC:Chl bilayers in confocal mode suggest that a higher reduction of lipid mobility for LLO incubated membranes than ClyA.…”

Section: Discussionmentioning

confidence: 73%

Preferential binding and re-organization of nanoscale domains on model lipid membranes by pore-forming toxins: insight from STED-FCS

Sarangi¹,

Basu²

2019

J. Phys. D: Appl. Phys.

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Potential nanodomains in cellular membranes are widely believed to be targeted by proteins and other biomolecules to enable execution of critical cellular functions. However, characterization of these nanodomains remains elusive, primarily due to the diffraction limit of a conventional optical microscope. Herein using super-resolution STED microscopy coupled with fluorescence correlation spectroscopy (STED-FCS), we provide experimental evidence of lipid nanodomains present in model membranes comprised of phosphocholine-cholesterol binary mixture, and its reorganization induced by pore-forming toxins (PFTs). In this study, we used two different types of PFTs, namely α-PFT (cytolysin A) and β-PFT (listeriolysin O), that preferentially associate as well as reorganize cholesterol-containing saturated and unsaturated phosphocholine membranes. The emergence of nanodomains due to the lipidlipid and lipid-PFT interactions was quantified at a length scales of ~50-150 nm using FCS diffusion law enabled by variation of spot sizes in the super-resolution STED microscopy. Our results shed light on the usefulness of super-resolution microscopy to quantify the underlying nanoscale domains in model membranes with implications for a wide variety of membranemediated cellular events observed in real cell membranes.

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Section: Discussionmentioning

confidence: 73%

Preferential binding and re-organization of nanoscale domains on model lipid membranes by pore-forming toxins: insight from STED-FCS

Sarangi¹,

Basu²

2019

J. Phys. D: Appl. Phys.

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“…Understanding the pore formation mechanism of ClyA has broader implications in our appreciation of the mechanisms of pore formation across the wider family of ClyA toxins which include the Cry6Aa toxin, bipartite XaxAB, YaxAB, as well as tripartite Nhe and Hbl toxins where several topological similarities are observed with ClyA. With advances in cryo-EM and single-molecule and super-resolution techniques , molecular insights into assembly pathways on PFTs will see an impetus in the future.…”

Section: Discussionmentioning

confidence: 99%

Mechanistic Insights into Pore Formation by an α-Pore Forming Toxin: Protein and Lipid Bilayer Interactions of Cytolysin A

2020

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Metrics & MoreArticle Recommendations CONSPECTUS: Pore forming toxins (PFTs) are the largest class of bacterial toxins playing a central role in bacterial pathogenesis.They are proteins specifically designed to form nanochannels in the membranes of target cells, ultimately resulting in cell death and establishing infection. PFTs are broadly classified as αand β-PFTs, depending on secondary structures that form the transmembrane channel. A unique feature about this class of proteins is the drastic conformational changes and complex oligomerization pathways that occur upon exposure to the plasma membrane. A molecular understanding of pore formation has implications in designing novel intervention strategies to combat rising antimicrobial resistance, targeted-cancer therapy, as well as designing nanopores for specialized technologies. Central to unraveling the pore formation pathway is the availability of high resolution crystal structures. In this regard, β-toxins are better understood, when compared with α-toxins whose pore forming mechanisms are complicated by an incomplete knowledge of the driving forces for amphiphatic membrane-inserted helices to organize into functional pores. With the publication of the first crystal structure for an α-toxin, cytolysin A (ClyA), in 2009 we embarked on an extensive multiscale study to unravel its pore forming mechanism. This Account represents the collective mechanistic knowledge gained in our laboratories using a variety of experimental and theoretical techniques which include large scale molecular dynamics (MD) simulations, kinetic modeling studies, single-molecule fluorescence imaging, and super-resolution spectroscopy. We reported MD simulations of the ClyA protomer, oligomeric intermediates, and full pore complex in a lipid bilayer and mapped the conformational transitions that accompany membrane binding. Using single-molecule fluorescence imaging, the conformational transition was experimentally verified by analysis of various diffusion states of membrane bound ClyA. Importantly, we have uncovered a hitherto unknown putative cholesterol binding motif in the membrane-inserted helix of ClyA. Distinct binding pockets for cholesterol formed by adjacent membrane-inserted helices are revealed in MD simulations. Cholesterol appears to play a dual role by stabilizing both the membrane-inserted protomer as well as oligomeric intermediates. Molecular dynamics simulations and kinetic modeling studies suggest that the membrane-inserted arcs oligomerize reversibly to form the predominant transmembrane oligomeric intermediates during pore formation. We posit that this mechanistic understanding of the complex action of α-PFTs has implications in unraveling pore assembly across the wider family of bacterial toxins. With emerging antimicrobial resistance, alternate therapies may rely on disrupting pore functionality or oligomerization of these pathogenic determinants utilized by bacteria, and our study includes assessing the potential for dendrimers as pore blockers.

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“…E. coli ClyA contains a single component, and YaxAB presents two-component. YaxAB from Yesinia enterolitica and XaxAB from Xenorhabdus nematophila have been determined to provide similar structure with ClyA but demonstrate low sequence identity [ 20 , 72 ]. In mature YaxAB pore, two-component proteins (bipartite PFTs) are arranged as ten symmetrical heterodimers showing a spoked rim from the top [ 19 ], while XaxAB possesses 12–15 heterodimers.…”

Section: Structural Classification and Characterization Of The Pormentioning

confidence: 99%

Structural Basis of the Pore-Forming Toxin/Membrane Interaction

Mengist

et al. 2021

Toxins

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With the rapid growth of antibiotic-resistant bacteria, it is urgent to develop alternative therapeutic strategies. Pore-forming toxins (PFTs) belong to the largest family of virulence factors of many pathogenic bacteria and constitute the most characterized classes of pore-forming proteins (PFPs). Recent studies revealed the structural basis of several PFTs, both as soluble monomers, and transmembrane oligomers. Upon interacting with host cells, the soluble monomer of bacterial PFTs assembles into transmembrane oligomeric complexes that insert into membranes and affect target cell-membrane permeability, leading to diverse cellular responses and outcomes. Herein we have reviewed the structural basis of pore formation and interaction of PFTs with the host cell membrane, which could add valuable contributions in comprehensive understanding of PFTs and searching for novel therapeutic strategies targeting PFTs and interaction with host receptors in the fight of bacterial antibiotic-resistance.

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Structural and Mechanistic Features of ClyA-Like α-Pore-Forming Toxins

Cited by 12 publications

References 38 publications

Preferential binding and re-organization of nanoscale domains on model lipid membranes by pore-forming toxins: insight from STED-FCS

Preferential binding and re-organization of nanoscale domains on model lipid membranes by pore-forming toxins: insight from STED-FCS

Mechanistic Insights into Pore Formation by an α-Pore Forming Toxin: Protein and Lipid Bilayer Interactions of Cytolysin A

Structural Basis of the Pore-Forming Toxin/Membrane Interaction

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