A membrane associated metalloprotease cleaves Cry3Aa Bacillus thuringiensis toxin reducing pore formation in Colorado potato beetle brush border membrane vesicles.
Insect proteases are implicated in Bacillus thuringiensis insecticidal proteins mode of action determining toxin specificity and sensitivity. Few data are available on the involvement of proteases in the later steps of toxicity such as protease interaction with toxin-receptor complexes and the pore formation process. In this study, a Colorado potato beetle (CPB) midgut membrane metalloprotease was found to be involved in the proteolytic processing of Cry3Aa. Interaction of Cry3Aa with BBMV membrane proteases resulted in a distinct pattern of proteolysis. Cleavage was demonstrated to occur in protease accessible regions of domain III and was specifically inhibited by the metalloprotease inhibitors 1,10-phenanthroline and acetohydroxamic acid. Proteolytic inhibition by a peptide representing a segment of proteolysis in domain III and the metalloprotease inhibitor acetohydroxamic acid correlated with increased pore formation, evidencing that Cry3Aa is a specific target of a CPB membrane metalloprotease that degrades potentially active toxin.
Rausell
C
C
Departamento de Genética, Facultad de Ciencias Biológicas, Universidad de Valencia, Dr. Moliner 50, Burjassot 46100, Valencia, Spain.
Ochoa-Campuzano
C
C
Martínez-Ramírez
A C
AC
Bravo
A
A
Real
M D
MD
D000818:Animals; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism* / Q000648:ultrastructure; D001427:Bacterial Toxins / Q000737:chemistry* / Q000378:metabolism*; D001665:Binding Sites; D002463:Cell Membrane Permeability / Q000502:physiology*; D001517:Coleoptera / Q000737:chemistry / Q000502:physiology*; D004731:Endotoxins / Q000737:chemistry* / Q000378:metabolism*; D006460:Hemolysin Proteins / Q000737:chemistry* / Q000378:metabolism* / Q000648:ultrastructure; D008871:Microvilli / Q000737:chemistry / Q000378:metabolism*; D008956:Models, Chemical; D016062:Porosity; D011485:Protein Binding; D022142:Secretory Vesicles / Q000737:chemistry / Q000378:metabolism*
D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D001427:Bacterial Toxins; D004731:Endotoxins; D006460:Hemolysin Proteins; C030956:insecticidal crystal protein, Bacillus Thuringiensis