title: Structural changes of the Cry1Ac oligomeric pre-pore from bacillus thuringiensis induced by N-acetylgalactosamine facilitates toxin membrane insertion.
abstract: The primary action of Cry toxins produced by Bacillus thuringiensis is to lyse midgut epithelial cells in their target insect by forming lytic pores. The toxin-receptor interaction is a complex process, involving multiple interactions with different receptor and carbohydrate molecules. It has been proposed that Cry1A toxins sequentially interact with a cadherin receptor, leading to the formation of a pre-pore oligomer structure, and that the oligomeric structure binds to glycosylphosphatidyl-inositol-anchored aminopeptidase-N (APN) receptor. The Cry1Ac toxin specifically recognizes the N-acetylgalactosamine (GalNAc) carbohydrate present in the APN receptor from Manduca sexta larvae. In this work, we show that the Cry1Ac pre-pore oligomer has a higher binding affinity with APN than the monomeric toxin. The effects of GalNAc binding on the toxin structure were studied in the monomeric Cry1Ac, in the soluble pre-pore oligomeric structure, and in its membrane inserted state by recording the fluorescence status of the tryptophan (W) residues. Our results indicate that the W residues of Cry1Ac have a different exposure to the solvent when compared with that of the closely related Cry1Ab toxin. GalNAc binding specifically affects the exposure of W545 in the pre-pore oligomer in contrast to the monomer where GalNAc binding did not affect the fluorescence of the toxin. These results indicate a subtle conformational change in the GalNAc binding pocket in the pre-pore oligomer that could explain the increased binding affinity of the Cry1Ac pre-pore to APN. Although our analysis did not reveal major structural changes in the pore-forming domain I upon GalNAc binding, it showed that sugar interaction enhanced membrane insertion of soluble pre-pore oligomeric structure. Therefore, the data presented here permits to propose a model in which the interaction of Cry1Ac pre-pore oligomer with APN receptor facilitates membrane insertion and pore formation.
authors.affiliation: Instituto de Biotecnologia, Universidad Nacional Autónoma de México, Apdo. Postal 510-3, Cuernavaca 62250, Morelos, Mexico.
authors.forename: Liliana
authors.initials: L
authors.lastname: Pardo-López
authors.forename: Isabel
authors.initials: I
authors.lastname: Gómez
authors.forename: Carolina
authors.initials: C
authors.lastname: Rausell
authors.forename: Jorge
authors.initials: J
authors.lastname: Sanchez
authors.forename: Mario
authors.initials: M
authors.lastname: Soberón
authors.forename: Alejandra
authors.initials: A
authors.lastname: Bravo
mesh_terms: D000117:Acetylglucosamine / Q000737:chemistry* / Q000378:metabolism; D000818:Animals; D001413:Bacillus thuringiensis / Q000737:chemistry* / Q000378:metabolism; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000737:chemistry* / Q000494:pharmacology; D001427:Bacterial Toxins / Q000737:chemistry* / Q000494:pharmacology; D018826:CD13 Antigens / Q000737:chemistry / Q000378:metabolism; D004731:Endotoxins / Q000737:chemistry* / Q000494:pharmacology; D006460:Hemolysin Proteins; D019476:Insect Proteins / Q000737:chemistry / Q000378:metabolism; D007306:Insecticides / Q000737:chemistry / Q000494:pharmacology; D007814:Larva / Q000378:metabolism / Q000382:microbiology; D018402:Manduca / Q000378:metabolism / Q000382:microbiology; D008567:Membranes, Artificial; D011485:Protein Binding; D020836:Protein Structure, Quaternary / Q000187:drug effects
chemical_list: D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D001427:Bacterial Toxins; D004731:Endotoxins; D006460:Hemolysin Proteins; D019476:Insect Proteins; D007306:Insecticides; D008567:Membranes, Artificial; C030956:insecticidal crystal protein, Bacillus Thuringiensis; D018826:CD13 Antigens; D000117:Acetylglucosamine