Role of two arginine residues in domain II, loop 2 of Cry1Ab and Cry1Ac Bacillus thuringiensisδ‐endotoxin in toxicity and binding to Manduca sexta and Lymantria dispar aminopeptidase N

Lee, Mi Kyong; Rajamohan, Francis; Jenkins, Jeremy L.; Curtiss, April; Dean, Donald H.

doi:10.1046/j.1365-2958.2000.02109.x

Cited by 32 publications

(40 citation statements)

References 43 publications

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“…1) The dissociation constants obtained during the course of this study are the maiden observations for a heterologously expressed and purified aminopeptidase receptor from any insect against Cry1Ac. Earlier studies have reported overall binding affinities using the surface plasmon resonance protocol against insect aminopeptidases purified from brush border membrane vesicles of the midgut (28 -30 (31,32). The off rates for dissociation calculated in this study for HaAPN1 with Cry1Ac works out to 2.08 ϫ 10 Ϫ6 s Ϫ1 , which is the lowest recorded for any APN/Cry protein interaction.…”

Section: Discussionmentioning

confidence: 71%

“…Also, the preliminary data of catalytic and Cry1Ac-interfering domain mapping suggests that the two domains do not overlap and may reside at different positions on the APN polypeptide. The lower values of dissociation obtained might be due to the fact that the APN employed in this study has been heterologously expressed and purified, whereas earlier studies relied on APN purified from brush border membrane vesicles of the host insect midgut (32).…”

Section: Discussionmentioning

confidence: 93%

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Knockdown of Aminopeptidase-N from Helicoverpa armigera Larvae and in Transfected Sf21 Cells by RNA Interference Reveals Its Functional Interaction with Bacillus thuringiensis Insecticidal Protein Cry1Ac

Swaminathan¹,

Rajagopal²,

Venkatesh

et al. 2007

Journal of Biological Chemistry

110

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Section: Discussionmentioning

confidence: 71%

Section: Discussionmentioning

confidence: 93%

Knockdown of Aminopeptidase-N from Helicoverpa armigera Larvae and in Transfected Sf21 Cells by RNA Interference Reveals Its Functional Interaction with Bacillus thuringiensis Insecticidal Protein Cry1Ac

Swaminathan¹,

Rajagopal²,

Venkatesh

et al. 2007

Journal of Biological Chemistry

110

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“…Also, we analyzed the binding of Cry1Ab mutants located in regions of domain II and domain III previously characterized as APN-binding epitopes (27,28). Our data suggest that both APN and ALP are functional receptor molecules that bind the oligomeric structure of Cry1Ab toxin with high affinity, facilitating membrane insertion and pore formation, and we also found that ALP may a have a predominant and more important role in Cry1Ab toxicity at earlier stages of larval development.…”

mentioning

confidence: 58%

“…Cry1Ab-binding Regions Involved in APN and ALP Interaction-Domain II loop 2 and 3 mutants were previously shown to be affected in APN binding (27,37,38). Also, a scFv antibody that bound the domain III ␤16-␤22 region competed the binding of Cry1Ab to APN (10).…”

Section: Analysis Of Binding Of Cry1abmentioning

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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“…In another study, additional Cry1Ac derivatives with mutations at loop 2, loop 3, and loop ␣8 were tested for their abilities to bind to purified L. dispar APN, and each showed decreased binding to only one of the two proposed APN binding sites (82). A subsequent study showed that loop ␣8 mutants also decreased binding to purified M. sexta APN (103).…”

Section: Other Receptorsmentioning

confidence: 99%

Role of Receptors in Bacillus thuringiensis Crystal Toxin Activity

Pigott

Ellar

2007

Microbiol Mol Biol Rev

600

598

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SUMMARY Bacillus thuringiensis produces crystalline protein inclusions with insecticidal or nematocidal properties. These crystal (Cry) proteins determine a particular strain's toxicity profile. Transgenic crops expressing one or more recombinant Cry toxins have become agriculturally important. Individual Cry toxins are usually toxic to only a few species within an order, and receptors on midgut epithelial cells have been shown to be critical determinants of Cry specificity. The best characterized of these receptors have been identified for lepidopterans, and two major receptor classes have emerged: the aminopeptidase N (APN) receptors and the cadherin-like receptors. Currently, 38 different APNs have been reported for 12 different lepidopterans. Each APN belongs to one of five groups that have unique structural features and Cry-binding properties. While 17 different APNs have been reported to bind to Cry toxins, only 2 have been shown to mediate toxin susceptibly in vivo. In contrast, several cadherin-like proteins bind to Cry toxins and confer toxin susceptibility in vitro, and disruption of the cadherin gene has been associated with toxin resistance. Nonetheless, only a small subset of the lepidopteran-specific Cry toxins has been shown to interact with cadherin-like proteins. This review analyzes the interactions between Cry toxins and their receptors, focusing on the identification and validation of receptors, the molecular basis for receptor recognition, the role of the receptor in resistant insects, and proposed models to explain the sequence of events at the cell surface by which receptor binding leads to cell death.

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Role of two arginine residues in domain II, loop 2 of Cry1Ab and Cry1Ac Bacillus thuringiensisδ‐endotoxin in toxicity and binding to Manduca sexta and Lymantria dispar aminopeptidase N

Cited by 32 publications

References 43 publications

Knockdown of Aminopeptidase-N from Helicoverpa armigera Larvae and in Transfected Sf21 Cells by RNA Interference Reveals Its Functional Interaction with Bacillus thuringiensis Insecticidal Protein Cry1Ac

Knockdown of Aminopeptidase-N from Helicoverpa armigera Larvae and in Transfected Sf21 Cells by RNA Interference Reveals Its Functional Interaction with Bacillus thuringiensis Insecticidal Protein Cry1Ac

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

Role of Receptors in Bacillus thuringiensis Crystal Toxin Activity

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