Cry4Aa and Cry4Ba Mosquito-Active Toxins Utilize Different Domains in Binding to a Particular Culex ALP Isoform: A Functional Toxin Receptor Implicating Differential Actions on Target Larvae
Issued Date
2022-10-01
Resource Type
eISSN
20726651
Scopus ID
2-s2.0-85140651955
Pubmed ID
36287921
Journal Title
Toxins
Volume
14
Issue
10
Rights Holder(s)
SCOPUS
Bibliographic Citation
Toxins Vol.14 No.10 (2022)
Suggested Citation
Dechkla M., Charoenjotivadhanakul S., Imtong C., Visitsattapongse S., Li H.C., Angsuthanasombat C. Cry4Aa and Cry4Ba Mosquito-Active Toxins Utilize Different Domains in Binding to a Particular Culex ALP Isoform: A Functional Toxin Receptor Implicating Differential Actions on Target Larvae. Toxins Vol.14 No.10 (2022). doi:10.3390/toxins14100652 Retrieved from: https://repository.li.mahidol.ac.th/handle/20.500.14594/86830
Title
Cry4Aa and Cry4Ba Mosquito-Active Toxins Utilize Different Domains in Binding to a Particular Culex ALP Isoform: A Functional Toxin Receptor Implicating Differential Actions on Target Larvae
Other Contributor(s)
Abstract
The three-domain Cry4Aa toxin produced from Bacillus thuringiensis subsp. israelensis was previously shown to be much more toxic to Culex mosquito larvae than its closely related toxin—Cry4Ba. The interaction of these two individual toxins with target receptors on susceptible larval midgut cells is likely to be the critical determinant in their differential toxicity. Here, two full-length membrane-bound alkaline phosphatase (mALP) isoforms from Culex quinquefasciatus larvae, Cq-mALP1263and Cq-mALP1264, predicted to be GPI-linked was cloned and functionally expressed in Spodoptera frugiperda (Sf9) cells as 57- and 61-kDa membrane-bound proteins, respectively. Bioinformatics analysis disclosed that both Cq-mALP isoforms share significant sequence similarity to Aedes aegypti-mALP—a Cry4Ba toxin receptor. In cytotoxicity assays, Sf9 cells expressing Cq-mALP1264, but not Cq-mALP1263, showed remarkably greater susceptibility to Cry4Aa than Cry4Ba, while immunolocalization studies revealed that both toxins were capable of binding to each Cq-mALP expressed on the cell membrane surface. Molecular docking of the Cq-mALP1264-modeled structure with individual Cry4 toxins revealed that Cry4Aa could bind to Cq-mALP1264 primarily through particular residues on three surface-exposed loops in the receptor-binding domain—DII, including Thr512, Tyr513 and Lys514 in the β10-β11loop. Dissimilarly, Cry4Ba appeared to utilize only certain residues in its C-terminal domain—DIII to interact with such a Culex counterpart receptor. Ala-substitutions of selected β10-β11loop residues (T512A, Y513A and K514A) revealed that only the K514A mutant displayed a drastic decrease in biotoxicity against C. quinquefasciatus larvae. Further substitution of Lys514 with Asp (K514D) revealed a further decrease in larval toxicity. Furthermore, in silico calculation of the binding affinity change (ΔΔGbind) in Cry4Aa-Cq-mALP1264 interactions upon these single-substitutions revealed that the K514D mutation displayed the largest ΔΔGbind value as compared to three other mutations, signifying an adverse impact of a negative charge at this critical receptor-binding position. Altogether, our present study has disclosed that these two related-Cry4 mosquito-active toxins conceivably exploited different domains in functional binding to the same Culex membrane-bound ALP isoform—Cq-mALP1264 for mediating differential toxicity against Culex target larvae.