title: Non conserved residues between Cqm1 and Aam1 mosquito α-glucosidases are critical for the capacity of Cqm1 to bind the Binary toxin from Lysinibacillus sphaericus.
abstract: The Binary (Bin) toxin from the entomopathogenic bacterium Lysinibacillus sphaericus acts on larvae of the culicid Culex quinquefasciatus through its binding to Cqm1, a midgut-bound α-glucosidase. Specific binding by the BinB subunit to the Cqm1 receptor is essential for toxicity however the toxin is unable to bind to the Cqm1 ortholog from the refractory species Aedes aegypti (Aam1). Here, to investigate the molecular basis for the interaction between Cqm1 and BinB, recombinant Cqm1 and Aam1 were first expressed as soluble forms in Sf9 cells. The two proteins were found to display the same glycosilation patterns and BinB binding properties as the native α-glucosidases. Chimeric constructs were then generated through the exchange of reciprocal fragments between the corresponding cqm1 and aam1 cDNAs. Subsequent expression and binding experiments defined a Cqm1 segment encompassing residues S129 and A312 as critical for the interaction with BinB. Through site directed mutagenesis experiments, replacing specific sets of residues from Cqm1 with those of Aam1, the 159GG160 doublet was required for this interaction. Molecular modeling mapped these residues to an exposed loop within the Cqm1's structure, compatible with a target site for BinB and providing a possible explanation for its lack of binding to Aam1.
authors.affiliation: Department of Entomology, Centro de Pesquisas Aggeu Magalhães/FIOCRUZ, Recife, PE 50740-465, Brazil.
authors.forename: Lígia Maria
authors.initials: LM
authors.lastname: Ferreira
authors.affiliation: Department of Entomology, Centro de Pesquisas Aggeu Magalhães/FIOCRUZ, Recife, PE 50740-465, Brazil.
authors.forename: Tatiany Patrícia
authors.initials: TP
authors.lastname: Romão
authors.affiliation: Department of Entomology, Centro de Pesquisas Aggeu Magalhães/FIOCRUZ, Recife, PE 50740-465, Brazil.
authors.forename: Nathaly Alexandre do
authors.initials: NA
authors.lastname: Nascimento
authors.affiliation: Department of Entomology, Centro de Pesquisas Aggeu Magalhães/FIOCRUZ, Recife, PE 50740-465, Brazil.
authors.forename: Maria da Conceição Mendes Ferreira da
authors.initials: Mda C
authors.lastname: Costa
authors.affiliation: Department of Microbiology, Centro de Pesquisas Aggeu Magalhães/FIOCRUZ, Recife, PE 50670-420, Brazil.
authors.forename: Antônio Mauro
authors.initials: AM
authors.lastname: Rezende
authors.affiliation: Department of Microbiology, Centro de Pesquisas Aggeu Magalhães/FIOCRUZ, Recife, PE 50670-420, Brazil.
authors.forename: Osvaldo Pompílio
authors.initials: OP
authors.lastname: de-Melo-Neto
authors.affiliation: Department of Entomology, Centro de Pesquisas Aggeu Magalhães/FIOCRUZ, Recife, PE 50740-465, Brazil. Electronic address: mhneves@cpqam.fiocruz.br.
authors.forename: Maria Helena Neves Lobo
authors.initials: MH
authors.lastname: Silva-Filha
mesh_terms: D000330:Aedes / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D000818:Animals; D001407:Bacillus; D001427:Bacterial Toxins / Q000378:metabolism* / Q000633:toxicity; D003465:Culex / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D004064:Digestive System / Q000378:metabolism; D019476:Insect Proteins / Q000235:genetics* / Q000378:metabolism; D007814:Larva; D016297:Mutagenesis, Site-Directed; D000520:alpha-Glucosidases / Q000378:metabolism*
chemical_list: D001427:Bacterial Toxins; D019476:Insect Proteins; D000520:alpha-Glucosidases
keywords: Aedes aegypti; Binding sites; Biolarvicides; Culex quinquefasciatus; Orthologs; Receptor