title: Isoleucine at position 150 of Cyt2Aa toxin from Bacillus thuringiensis plays an important role during membrane binding and oligomerization.
abstract: Cyt2Aa2 is a mosquito larvicidal and cytolytic toxin produced by Bacillus thuringiensis subsp. darmstadiensis. The toxin becomes inactive when isoleucine at position 150 was replaced by alanine. To investigate the functional role of this position, Ile150 was substituted with Leu, Phe, Glu and Lys. All mutant proteins were produced at high level, solubilized in carbonate buffer and yielded protease activated product similar to those of the wild type. Intrinsic fluorescence spectra analysis suggested that these mutants retain similar folding to the wild type. However, mosquito larvicidal and hemolytic activities dramatically decreased for the I150K and were completely abolished for I150A and I150F mutants. Membrane binding and oligomerization assays demonstrated that only I150E and I150L could bind and form oligomers on lipid membrane similar to that of the wild type. Our results suggest that amino acid at position 150 plays an important role during membrane binding and oligomerization of Cyt2Aa2 toxin.
authors.affiliation: National Center for Genetic Engineering and Biotechnology, National Science and Technology Development Agency, 113 Phahonyothin Road, Khlong Nueng, Khlong Luang, Pathum Thani 12120, Thailand.
authors.forename: Wanwarang
authors.initials: W
authors.lastname: Pathaichindachote
authors.forename: Amporn
authors.initials: A
authors.lastname: Rungrod
authors.forename: Mongkon
authors.initials: M
authors.lastname: Audtho
authors.forename: Sumarin
authors.initials: S
authors.lastname: Soonsanga
authors.forename: Chartchai
authors.initials: C
authors.lastname: Krittanai
authors.forename: Boonhiang
authors.initials: B
authors.lastname: Promdonkoy
mesh_terms: D001413:Bacillus thuringiensis / Q000378:metabolism*; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism*; D001483:Base Sequence; D001704:Biopolymers / Q000737:chemistry / Q000378:metabolism*; D002151:Calorimetry; D017931:DNA Primers; D004731:Endotoxins / Q000737:chemistry / Q000378:metabolism*; D006460:Hemolysin Proteins / Q000737:chemistry / Q000378:metabolism*; D007532:Isoleucine / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding
chemical_list: D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D001704:Biopolymers; D017931:DNA Primers; D004731:Endotoxins; D006460:Hemolysin Proteins; C030956:insecticidal crystal protein, Bacillus Thuringiensis; D007532:Isoleucine