Article

Structures of Lysenin Reveal a Shared Evolutionary Origin for Pore-Forming Proteins And Its Mode of Sphingomyelin Recognition

Division of Structural Biology, Wellcome Trust Centre for Human Genetics, University of Oxford, Roosevelt Drive, Oxford OX3 7BN, UK.
Structure (Impact Factor: 5.62). 07/2012; 20(9):1498-507. DOI: 10.1016/j.str.2012.06.011
Source: PubMed

ABSTRACT

Pore-forming proteins insert from solution into membranes to create lesions, undergoing a structural rearrangement often accompanied by oligomerization. Lysenin, a pore-forming toxin from the earthworm Eisenia fetida, specifically interacts with sphingomyelin (SM) and may confer innate immunity against parasites by attacking their membranes to form pores. SM has important roles in cell membranes and lysenin is a popular SM-labeling reagent. The structure of lysenin suggests common ancestry with other pore-forming proteins from a diverse set of eukaryotes and prokaryotes. The complex with SM shows the mode of its recognition by a protein in which both the phosphocholine headgroup and one acyl tail are specifically bound. Lipid interaction studies and assays using viable target cells confirm the functional reliance of lysenin on this form of SM recognition.

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    • "Recently, the mushroomderived , structurally homologous proteins of the aegerolysin family [105] were employed to visualize CPE, independently of the lipid envi- ronment [47] . Crystallographic analysis combined with point mutagenesis studies has facilitated the precise mapping of SL-binding domains and the development of non-toxic derivatives suitable for live-cell im- aging [106,107]. Antibodies and toxins have also been used to visualize SM and CPE distribution through EM. Based on rapid freezing and freeze fracture techniques, the intracellular distribution and asymmetry of the lipids between the two leaflets of the plasma membrane have been revealed [47,91]. "
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    ABSTRACT: Pore-forming toxins (PFTs) represent a unique class of highly specific lipid-binding proteins. The cytotoxicity of these compounds has been overcome through crystallographic structure and mutation studies, facilitating the development of non-toxic lipid probes. As a consequence, non-toxic PFTs have been utilized as highly specific probes to visualize the diversity and dynamics of lipid nanostructures in living and fixed cells. This review is focused on the application of PFTs and their non-toxic analogs as tools to visualize sphingomyelin and ceramide phosphoethanolamine, two major phosphosphingolipids in mammalian and insect cells, respectively.
    No preview · Article · Oct 2015 · Biochimica et Biophysica Acta
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    • "Eisenia fetida expels its coelomic fluid when attacked, and the fluid is known to be toxic to vertebrates, probably as a result of the presence of Lysenin (Kobayashi et al. 2001). Lysenin is hemolytic and can lyse cells by inserting into cell membranes, an ability which probably also allows it to play a role in innate immunity as it is able to attack the cell membranes of parasites (De Colibus et al. 2012). "
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    • "This experiment provided supplementary evidence for the channels not being pulled out from the supportive BLM by the cationic polymers. Given the particular structure of the lysenin monomer interacting with SM in a BLM [49], the asymmetric shape, and the hydrophilic C-terminus, it is unlikely that either of the charged polymers could successfully pull the lysenin channels in either direction through the BLM. Nonetheless, this experiment led us to conclude that the hypothesis of a unique binding site closer to the trans side may be no longer valid. "
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    ABSTRACT: The pore-forming toxin lysenin self-assembles large and stable conductance channels in natural and artificial lipid membranes. The lysenin channels exhibit unique regulation capabilities, which open unexplored possibilities to control the transport of ions and molecules through artificial and natural lipid membranes. Our investigations demonstrate that the positively charged polymers polyethyleneimine and chitosan inhibit the conducting properties of lysenin channels inserted into planar lipid membranes. The preservation of the inhibitory effect following addition of charged polymers on either side of the supporting membrane suggests the presence of multiple binding sites within the channel's structure and a multistep inhibition mechanism that involves binding and trapping. Complete blockage of the binding sites with divalent cations prevents further inhibition in conductance induced by the addition of cationic polymers and supports the hypothesis that the binding sites are identical for both multivalent metal cations and charged polymers. The investigation at the single-channel level has shown distinct complete blockages of each of the inserted channels. These findings reveal key structural characteristics which may provide insight into lysenin's functionality while opening innovative approaches for the development of applications such as transient cell permeabilization and advanced drug delivery systems.
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