Aminopeptidase N from Bombyx mori as a candidate for the receptor of Bacillus thuringiensis Cry1Aa toxin.
Cry1Aa toxin-binding proteins from the midgut brush border membrane vesicles of Bombyx mori, a toxin-susceptible silkworm, were analyzed to find candidates for the toxin receptors. Ligand blotting showed that Cry1Aa toxin bound to a 120-kDa protein. A part of the 120-kDa protein was solubilized from the membrane vesicles with phosphatidylinositol-specific phospholipase C, resulting in a 110-kDa protein which therefore may be linked to a glycosyl-phosphatidylinositol anchor. The 120-kDa and 110-kDa Cry1Aa toxin-binding proteins were solubilized with detergent or pohosphatidylinositol-specific phospholipase C, respectively, and purified using anion-exchange chromatography. Scatchard plot analysis for the specific binding of purified 110-kDa protein to Cry1Aa toxin yielded a Kd value of 7.6 nM, which was similar to that for the binding of intact brush border membrane vesicles to the toxin. N-terminal and internal amino acid sequences of the 120-kDa and 110-kDa proteins showed high degrees of similarity to those of aminopeptidase N, a putative Cry1Ac toxin receptor, reported in Manduca sexta and Heliothis virescens. On this basis, the 120-kDa Cry1Aa toxin-binding protein from B. mori was identified as a member of the aminopeptidase family.
Yaoi
K
K
Department of Applied Biological Science, Faculty of Agriculture, Tokyo University of Agriculture and Technology, Japan.
Kadotani
T
T
Kuwana
H
H
Shinkawa
A
A
Takahashi
T
T
Iwahana
H
H
Sato
R
R
D000595:Amino Acid Sequence; D000818:Animals; D001413:Bacillus thuringiensis; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism*; D001427:Bacterial Toxins; D001665:Binding Sites; D012831:Bombyx; D018826:CD13 Antigens / Q000737:chemistry / Q000378:metabolism*; D002352:Carrier Proteins / Q000737:chemistry / Q000378:metabolism; D004731:Endotoxins / Q000737:chemistry / Q000378:metabolism*; D004797:Enzyme-Linked Immunosorbent Assay; D006460:Hemolysin Proteins; D019476:Insect Proteins; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010446:Peptide Fragments / Q000737:chemistry; D010449:Peptide Mapping; D011956:Receptors, Cell Surface / Q000737:chemistry / Q000378:metabolism*; D012995:Solubility
D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D001427:Bacterial Toxins; D002352:Carrier Proteins; C095416:Cry toxin receptors; D004731:Endotoxins; D006460:Hemolysin Proteins; D019476:Insect Proteins; D010446:Peptide Fragments; D011956:Receptors, Cell Surface; C030956:insecticidal crystal protein, Bacillus Thuringiensis; D018826:CD13 Antigens