Brush border membrane aminopeptidase-n in the midgut of the gypsy moth serves as the receptor for the CryIA(c) δ-endotoxin of Bacillus thuringiensis

Valaitis, Algimantas P.; Lee, Mi Kyong; Rajamohan, Francis; Dean, Donald H.

doi:10.1016/0965-1748(95)00050-x

Cited by 112 publications

(73 citation statements)

References 39 publications

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“…The first receptor to be identified was detected recently as a protein of 120 kDa identified as an aminopeptidase-N. This protein bound B. thuringiensis CrylAc in M. sexta (Knight et al, 1994), in Lymantria dispar (Valaitis et al, 1995) and in Heliothis virescens (Gill et al, 1995). In dipteran species such as A. stephensi, a 148-kDa protein was found to bind B. thuringiensis serovar israelensis CryllA (Feldmann et al, 1995).…”

Section: Discussionmentioning

confidence: 99%

Binding Kinetics of Bacillus Sphaericus Binary Toxin to Midgut Brush‐Border Membranes of Anopheles and Culex sp. Mosquito Larvae

Silva-Filha

Nielsen-LeRoux

Charles

1997

European Journal of Biochemistry

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Direct-binding assays and homologous-competition assays were used to identify specific binding between the radiolabelled toxin of Bacillus sphaericus and brush-border membrane fractions (BBMF) from Anopheles gambiae and Anopheles stephensi, obtained from whole larvae preparations. In both species, the toxin bound to a single class of receptors. BBMF of A. gunzbiae had the highest binding affinity for the toxin of the species tested, with a dissociation constant ( K J of 30 2 15 nM and a maximum receptor concentration of 5 2 1 pmol/mg. Toxin binding to A. gambiae BBMF was compared with that to BBMF from B. sphaericus-susceptible (IP) and B. sphaericus-resistant (SPHAE) Culex pipiens populations. BBMF toxin binding was slower in A. gambiae than in the C. pipiens populations. The BBMF of the B. sphaericus-resistant population of C. pipiens had an association profile that was similar to the susceptible population, despite of the lack of susceptibility in vivo. No relationship between toxicity and irreversibility of toxin binding was detected. On the contrary, toxin dissociation from BBMF was fast and almost complete in BBMF of all species studied.

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Section: Discussionmentioning

confidence: 99%

Binding Kinetics of Bacillus Sphaericus Binary Toxin to Midgut Brush‐Border Membranes of Anopheles and Culex sp. Mosquito Larvae

Silva-Filha

Nielsen-LeRoux

Charles

1997

European Journal of Biochemistry

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“…The BBMV proteins that were solubilized with CHAPS as stated above were concentrated by Amicon YM-50 ultrafiltration, and a 2-ml aliquot was applied to a HR 5/10 Mono Q (GE Healthcare) column equilibrated with 20 mM Tris-HCl pH 8.5 buffer containing 2 mM MgCl 2 , 2 mM KCl (28,30). The column was eluted with a gradient of NaCl (0.5-1 M) with a flow rate of 1 ml/min for 40 min.…”

Section: Methodsmentioning

confidence: 99%

Role of Alkaline Phosphatase from Manduca sexta in the Mechanism of Action of Bacillus thuringiensis Cry1Ab Toxin

Arenas

Bravo

Soberón

et al. 2010

Journal of Biological Chemistry

155

157

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Cry toxins produced by Bacillus thuringiensis have been recognized as pore-forming toxins whose primary action is to lyse midgut epithelial cells in their target insect. In the case of the Cry1A toxins, a prepore oligomeric intermediate is formed after interaction with cadherin receptor. The Cry1A oligomer then interacts with glycosylphosphatidylinositol-anchored receptors. Two Manduca sexta glycosylphosphatidylinositol-anchored proteins, aminopeptidase (APN) and alkaline phosphatase (ALP), have been shown to bind Cry1Ab, although their role in toxicity remains to be determined. Detection of Cry1Ab binding proteins by ligand blot assay revealed that ALP is preferentially expressed earlier during insect development, because it was found in the first larval instars, whereas APN is induced later after the third larval instar. The binding of Cry1Ab oligomer to pure preparations of APN and ALP showed that this toxin structure interacts with both receptors with high affinity (apparent K d ‫؍‬ 0.6 nM), whereas the monomer showed weaker binding (apparent K d ‫؍‬ 101.6 and 267.3 nM for APN and ALP, respectively). Several Cry1Ab nontoxic mutants located in the exposed loop 2 of domain II or in ␤-16 of domain III were affected in binding to APN and ALP, depending on their oligomeric state. In particular monomers of the nontoxic domain III, the L511A mutant did not bind ALP but retained APN binding, suggesting that initial interaction with ALP is critical for toxicity. Our data suggest that APN and ALP fulfill two roles. First APN and ALP are initial receptors promoting the localization of toxin monomers in the midgut microvilli before interaction with cadherin. Then APN and ALP function as secondary receptors mediating oligomer insertion into the membrane. However, the expression pattern of these receptors and the phenotype of L511A mutant suggest that ALP may have a predominant role in toxin action because Cry toxins are highly effective against the neonate larvae that is the target for pest control programs.The Cry toxins produced by Bacillus thuringiensis are used worldwide as effective biological control agents for many species of insects including agricultural and forest pests and several vectors of human and animal diseases. Its insecticidal property results from crystalline inclusions produced during sporulation that are formed by ␦-endotoxins known as Cry toxins (1).The Cry protein family is composed of more than 54 groups, among which the three-domain Cry family forms the major group, having members that show toxicity to different insect orders and to nematodes. The crystal structure has been solved for six three-domain Cry toxins and one protoxin that display different insect specificities (2). The activated Cry toxins are globular molecules consisting of a bundle of seven ␣-helices (domain I), a three-␤-sheet prism (domain II), and a ␤-sandwich (domain III) (3, 4). The N-terminal domain I is involved in oligomer formation, membrane insertion, and pore formation (5-8). Domain II and III are involved in the recogn...

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“…A putative receptor for the Cry1Ac toxin has been identified as aminopeptidase N (APN) in four lepidopteran insects [9][10][11][12][13]. APN was also identified as a receptor for Cry1Aa in Bombyx mori (silkworm) [14], and for Cry1C and Cry1Ab in Manduca sexta (tobacco hornworm) [15,16].…”

Section: Introductionmentioning

confidence: 99%

Bacillus thuringiensis Cry1Ac toxin interaction with Manduca sexta aminopeptidase N in a model membrane environment

COOPER¹,

CARROLL²,

TRAVIS³

et al. 1998

Biochemical Journal

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The Bacillus thuringiensis Cry1Ac δ-endotoxin was shown to bind in a biphasic manner to Manduca sexta aminopeptidase N (APN) present in a novel model membrane. Surface plasmon resonance analysis allowed the quantification of toxin binding to M. sexta APN in a supported lipid monolayer. The initial binding was rapid and reversible, with an affinity constant of 110 nM. The second phase was slower and resulted in an overall affinity constant of 3.0 nM. Reagents used to disrupt protein-protein interactions did not dissociate the toxin after highaffinity binding was attained. The initial association between Cry1Ac and APN was inhibited by the sugar GalNAc, but the

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Brush border membrane aminopeptidase-n in the midgut of the gypsy moth serves as the receptor for the CryIA(c) δ-endotoxin of Bacillus thuringiensis

Cited by 112 publications

References 39 publications

Binding Kinetics of Bacillus Sphaericus Binary Toxin to Midgut Brush‐Border Membranes of Anopheles and Culex sp. Mosquito Larvae

Binding Kinetics of Bacillus Sphaericus Binary Toxin to Midgut Brush‐Border Membranes of Anopheles and Culex sp. Mosquito Larvae

Role of Alkaline Phosphatase from Manduca sexta in the Mechanism of Action of Bacillus thuringiensis Cry1Ab Toxin

Bacillus thuringiensis Cry1Ac toxin interaction with Manduca sexta aminopeptidase N in a model membrane environment

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