Abstract
Lysenin from the coelomic fluid of the earthworm Eisenia fetida belongs to the aerolysin family of small β-pore-forming toxins (β-PFTs), some members of which are pathogenic to humans and animals. Despite efforts, a high-resolution structure of a channel for this family of proteins has been elusive and therefore the mechanism of activation and membrane insertion remains unclear. Here we determine the pore structure of lysenin by single particle cryo-EM, to 3.1 Å resolution. The nonameric assembly reveals a long β-barrel channel spanning the length of the complex that, unexpectedly, includes the two pre-insertion strands flanking the hypothetical membrane-insertion loop. Examination of other members of the aerolysin family reveals high structural preservation in this region, indicating that the membrane-insertion pathway in this family is conserved. For some toxins, proteolytic activation and pro-peptide removal will facilitate unfolding of the pre-insertion strands, allowing them to form the β-barrel of the channel.
Highlights
Lysenin from the coelomic fluid of the earthworm Eisenia fetida belongs to the aerolysin family of small b-pore-forming toxins (b-PFTs), some members of which are pathogenic to humans and animals
The defence protein of the earthworm Eisenia fetida, is produced by coelomocytes in the body fluid that form part of the earthworm’s immune system[1]. It is a member of the aerolysin family of small b-pore-forming toxins (b-PFTs) that include key virulence factors of a large number of bacterial pathogens, such as aerolysin produced by Aeromonas spp[2], Clostridium perfringens epsilon toxin[3] and Clostridium septicum a-toxin[4], while Bacillus thuringiensis parasporin-2 has cytocidal activity against human cancer cells[5]
Image analysis and two-dimensional (2D) classification of 53,779 particles led to a data set of 42,830 particles, which were used for an initial 3D reconstruction at 3.4 Å resolution with C9 symmetry or 4.2 Å
Summary
Lysenin from the coelomic fluid of the earthworm Eisenia fetida belongs to the aerolysin family of small b-pore-forming toxins (b-PFTs), some members of which are pathogenic to humans and animals. The defence protein of the earthworm Eisenia fetida, is produced by coelomocytes in the body fluid that form part of the earthworm’s immune system[1] It is a member of the aerolysin family of small b-pore-forming toxins (b-PFTs) that include key virulence factors of a large number of bacterial pathogens, such as aerolysin produced by Aeromonas spp[2], Clostridium perfringens epsilon toxin[3] and Clostridium septicum a-toxin[4], while Bacillus thuringiensis parasporin-2 has cytocidal activity against human cancer cells[5]. The structure provides the first atomic resolution view into the membrane-inserted state of a member of the aerolysin family and gives insights into how other members of this family may be activated and assemble into their respective pore forms
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