One of these clones gave germline transfer of the mutated gene and was used to create PME-1 mice on an outbred background

1 nm, respectively, in lipid membranes. However, Delta toxin channels might have a pore structure similar to that of Staphylococcus alpha hemolysin, which is considered as the basic model of b-barrel pore-forming toxins. The latter includes the aerolysin family which encompasses aerolysin, C. septicum alpha toxin, C. perfringens epsilon toxin, and probably Beta toxin. The large channels formed by Delta toxin could account for the broad spectrum of conductance and a detergent-like effect. b-Barrel pore-forming toxins contain amphipatic b-hairpin forming sequences that associate to form a b-barrel when the toxin is oligomerized, which inserts itself into the lipid bilayer, resulting in pore formation. Two stretches of alternating hydrophilic and hydrophobic residues were identified in the N-terminal region of Delta toxin and one in Beta toxin using the program http://psfs.cbrc.jp/tmbeta-net/. These sequences are IPI 145 likely two amphipatic b-strands involved in b-barrel formation. Only one of the two putative amphipatic b-hairpins in Delta toxin can be involved in pore formation as this was shown for S. aureus alpha toxin which requires one b-hairpin from each monomer to form the bbarrel. Alternatively both putative amphipatic b-hairpins can be involved in pore formation as this is the case with the two transmembrane hairpins of Perfringolysin O. In contrast to aerolysin and probably C. perfringens epsilon toxin, where the bhairpin forming the pore is located in domain 3 from the central region of the toxin, the 20171952 putative b-hairpins are found in the N-terminal part of Delta toxin. A similar location has been observed in C. perfringens enterotoxin, where residues from 81 to 106 predicted to form an amphipatic loop are important in pore formation. However, further investigations are required to precisely define the pore domain in Delta and Beta toxins. The role of Delta toxin in pathogenesis is not well understood. C. perfringens type B and C of which some strains can produce Delta toxin as an additional toxin, are involved in necrotic enteritis in various animal species, mainly piglets, and also in human. Beta toxin is considered as the main virulence factor of 10980276 these strains. Recently, NetB was found to be responsible for necrotic enteritis in chicken using a C. perfringens netB mutant. Based on the relatedness of Delta toxin with Beta and NetB toxins, Delta toxin might represent a potent virulence factor, which can induce intestinal diseases. The genetic characterization of Delta toxin will be useful for further epidemiological studies and mutant analysis to address the involvement of this toxin in pathology. In conclusion, C. perfringens Delta toxin has been characterized at the amino acid level and is highly related to C. perfringens Beta toxin and to a lesser extent to C. perfringens NetB as well as to Staphylococcus alpha hemolysin and leukotoxins. As wild type toxin, recombinant Delta recognizes GM2 and is cytotoxic for cells enriched in GM2 in their membrane. The C-terminal part of Delta is involved in the recognition of the cell membrane receptor. Delta toxin forms channels in artificial lipid bilayers, which are anion selective and larger than those induced by Staphylococcus alpha hemolysin and toxins from the aerolysin family characterized by a heptameric pore structure. Delta toxin probably retains a common structure organization with that of b-pore forming toxins, but its exact mode of action remains to be determined. Since Delta tox

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