Unfolding events in the water-soluble monomeric Cry1Ab toxin during transition to oligomeric pre-pore and membrane-inserted pore channel.

摘要: The insecticidal crystal (Cry) proteins produced by Bacillus thuringiensis undergo several conformational changes from inclusion protoxins to membrane-inserted channels in the midgut epithelial cells of target insect. Here we analyzed stability different forms Cry1Ab toxin, monomeric pre-pore complex, and channel, after urea thermal denaturation monitoring intrinsic tryptophan fluorescence protein 1-anilinonaphthalene-8-sulfonic acid binding partially unfolded proteins. Our results showed that flexibility toxin was dramatically enhanced upon oligomerization even further increased insertion into membrane as shown lower concentration chaotropic agents needed achieve unfolding oligomeric species. structures is alkaline pH. We found monomer-monomer interaction highly stable because promotes oligomer without disassembly. Partial limited proteolysis studies demonstrated domains II III were less unfold first, followed most domain I, also I involved interaction. thermal-induced analysis energy transfer Trp residues bound dye pore are particularly sensitive heat denaturation, contrast suggesting only may be inserted membrane. Finally, structure not affected However, a looser conformation induced neutral or pH correlates with active channel formation. suggest for first time more flexible Cry could necessary insertion, this oligomerization. Finally lumen lepidopteran insects increase