Autor(es): Zhang, Rui; Hua, Gang; Andacht, Tracy M.; Adang, Michael J.

Resumo: Bacillus thuringiensis (Bt) insecticidal toxins bind to receptors on midgut epithelial cells of susceptible insects, and binding triggers biochemical events that lead to insect mortality. Recently, a 100-kDa aminopeptidase N (APN) was isolated from brush border membrane vesicles (BBMV) of Anopheles quadrimaculatus and shown to bind Cry11Ba toxin with surface plasmon resonance (SPR) detection [Abdullah et al. (2006) BMC Biochem. 7, 16]. In our study, a 106-kDa APN, called AgAPN2, released by phosphatidylinositol-specific phospholipase C (PI-PLC) from Anopheles gambiae BBMV was extracted by Cry11Ba bound to beads. The AgAPN2 cDNA was cloned, and analysis of the predicted AgAPN2 protein revealed a zinc-binding motif (HEIAH), three potential N-glycosylation sites, and a predicted glycosylphosphatidylinositol (GPI) anchor site. Immunohistochemistry localized AgAPN2 to the microvilli of the posterior midgut. A 70-kDa fragment of the 106-kDa APN was expressed in Escherichia coli. When purified, it competitively displaced I-125-Cry11Ba binding to An. gambiae BBMV and bound Cry11Ba on dot blot and microtiter plate binding assays with a calculated K-d of 6.4 nM. Notably, this truncated peptide inhibited Cry11Ba toxicity to An. gambiae larvae. These results are evidence that the 106-kDa GPI-anchored APN is a specific binding protein, and a putative midgut receptor, for Bt Cry11Ba toxin.

Palavras-Chave: Aedes - Aegypti larvae ;ÿDelta - Endotoxin ;ÿBinding - Properties ; Culex - Pipiens ;ÿManduca -Sexta ;ÿMidgut ceca ;ÿBombyx - Mori ;ÿMembranes ;ÿProteins ; Culicidae

Imprenta: Biochemistry, v. 47, n. 43, p. 11263-11272, 2008

Identificador do objeto digital: 10.1021/bi801181g

Descritores: Aedes aegypti - Cell ; Aedes aegypti - Proteins

Data de publicação: 2008

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