Abstract
The δ-endotoxin CytB, found in parasporal inclusions of Bacillus thuringiensissubspecies kyushuensis, is a membrane pore-forming protein which is lethal to the larvae of Dipteraninsects and broadly cytolytic in vitro. The crystal structure of CytB in the protoxin form has been determined by isomorphous replacement using heavy-atom derivatives of both the wild-type protein and an engineered cysteine mutant. The atomic model comprising residues 19 to 245 and 28 bound water molecules has been refined at 2.6 Å resolution to a crystallographic R-factor of 19.7% and a free R-factor of 26.1%. CytB has a single domain of α/β architecture but a novel connectivity, comprising two outer layers of α-helix hairpins wrapped around a mixed β-sheet. In the protoxin form, CytB is a dimer linked by the intertwined N-terminal strands in a continuous, 12-stranded β-sheet. Proteolytic processing cleaves the intertwined β-strands to release the active CytB as a monomer, as well as removing the C-terminal tail to uncover the three-layered core. The homologous toxin CytA should show the same fold. Mutations in CytA that inhibit expression map to the dimer contacts and to the tip of helix pair A-B in contact with the sheet, apparently preventing correct folding. Mutations that inhibit toxicity map to the edge of the β-sheet adjoining the helix pair C-D and to the sheet face, while mutations on the helix surfaces have no effect. Therefore segments forming the sheet, rather than the amphiphilic but short helices, are responsible for membrane binding and pore formation. A conformational change is postulated by which the helix pair C-D peels away from the sheet to lie on the membrane surface, while the sheet region rearranges to form an oligomeric trans-membrane pore.
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Disclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.