Identification of the Functional Site in the Mosquito Larvicidal Binary Toxin of Bacillus sphaericus 1593M by Site-Directed Mutagenesis

Elangovan, G.; Shanmugavelu, M.; Rajamohan, Francis; Dean, Donald H.; Jayaraman, Kunthala

doi:10.1006/bbrc.2000.3575

Cited by 21 publications

(30 citation statements)

References 24 publications

(37 reference statements)

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“…Since the interaction of the binary toxin with target lipid membrane is one of the key steps in eliciting cytopathological effects on mosquito larvae, molecular insights into the interaction of the binary toxin with lipid membranes is required to gain a better understanding of the mechanism of toxicity. Due to the lack of three dimensional structure of the binary toxin, functional characterization has been based mainly on its amino acid sequence analysis and secondary structure prediction (10)(11)(12)(13)(14)(15). Aromatic residues, especially tyrosine and tryptophan, conceivably play a key role in membrane anchoring of many membrane proteins and are mainly found at the membrane-water interface (16).…”

Section: Introductionmentioning

confidence: 99%

Essential role of tryptophan residues in toxicity of binary toxin from Bacillus sphaericus

Kunthic

Promdonkoy

Srikhirin³

et al. 2011

BMB Reports

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Bacillus sphaericus produces mosquito-larvicidal binary toxin composed of BinA and BinB. While BinB is expected to bind to a specific receptor on the cell membrane, BinA interacts to BinB or BinB receptor complex and translocates into the cytosol to exert its activity via unknown mechanism. To investigate functional roles of aromatic cluster in BinA, amino acids at positions Y213, Y214, Y215, W222 and W226 were substituted by leucine. All mutant proteins were highly produced and their secondary structures were not affected by these substitutions. All mutants are able to insert into lipid monolayers as observed by Langmuir-Blodgett trough and could permeabilize the liposomes in a similar manner as the wild type. However, mosquito-larvicidal activity was abolished for W222L and W226L mutants suggesting that tryptophan residues at both positions play an important role in the toxicity of BinA, possibly involved in the cytopathological process after toxin entry into the cells. [BMB reports 2011; 44(10): 674-679]

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Section: Introductionmentioning

confidence: 99%

Essential role of tryptophan residues in toxicity of binary toxin from Bacillus sphaericus

Kunthic

Promdonkoy

Srikhirin³

et al. 2011

BMB Reports

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“…In the absence of its 3D structure, directed-mutagenesis is one of useful techniques for studying the mode of action of binary toxin. IQ 393 → AA) totally lost toxicity towards C. quinquefasciatus larvae (7,23). Although the exact roles of these amino acids had not been elucidated, ability for two nontoxic derivatives, one with mutation at the N-terminus and the other with mutation at the C-terminus, to functionally complement each other suggests that binary toxin functions as oligomers (7,23).…”

Section: C67 and C161 Are Required For Toxicity Of Binary Toxinmentioning

confidence: 99%

“…IQ 393 → AA) totally lost toxicity towards C. quinquefasciatus larvae (7,23). Although the exact roles of these amino acids had not been elucidated, ability for two nontoxic derivatives, one with mutation at the N-terminus and the other with mutation at the C-terminus, to functionally complement each other suggests that binary toxin functions as oligomers (7,23). This result is in agreement with the finding that in solution binary toxin exists as tetramer (13).…”

Section: C67 and C161 Are Required For Toxicity Of Binary Toxinmentioning

confidence: 99%

“…Once ingested by susceptible larvae, the protein crystal is solubilized and proteolytically activated in the alkaline condition of the larvae gut (1,6). BinA protein is responsible for toxicity via unknown mechanism, while BinB is a receptor binding component which binds to a single class of receptor on brush border membrane of the larvae (7,8). The binary toxin receptor in Culex pipiens larvae is a 60 kDa protein identified as an α-glucosidase anchored on the membrane via a glycosyl-phosphatidylinositol (GPI) moiety (9,10).…”

Section: Introductionmentioning

confidence: 99%

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Role of cysteine at positions 67, 161 and 241 of a Bacillus sphaericus binary toxin BinB

Boonyos¹,

Soonsanga²,

Boonserm³

et al. 2010

BMB Reports

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Binary toxin consisting of BinA and BinB from Bacillus sphaericus is toxic to mosquito larvae. BinB is responsible for specific binding to the larval gut cell membrane while BinA is crucial for toxicity. To investigate functional role of cysteine in BinB, three cysteine residues at positions 67, 161, and 241 were replaced by alanine or serine. Mutations at these positions did not affect protein production and overall structure of BinB. These cysteine residues are not involved in disulfide bond formation between BinB molecules. Mosquito-larvicidal assays revealed that C67 and C161 are essential for toxicity, whereas C241 is not. Mutations at C67 and C161 resulted in weaker BinA-BinB interaction. The loss of toxicity may be due to the reduction of interactions between BinA and BinB or BinB and its receptor. C67 and C161 could also play a part during conformational changes or internalization of the binary toxin into the target cell. [BMB reports 2010; 43(1): 23-28]

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“…This is followed by internalization of both components, resulting in cell death mediated by an unknown mechanism (Oei et al, 1992). Previous studies have suggested that BinB is a two-domain protein in which the N-terminal half would play a role in receptor binding, whereas the C-terminal half would be responsible for interaction with BinA (Elangovan et al, 2000;Oei et al, 1992).…”

Section: Introductionmentioning

confidence: 99%

Functional characterization of truncated fragments of Bacillus sphaericus binary toxin BinB

Tangsongcharoen¹,

Boonserm²,

Promdonkoy³

2011

Journal of Invertebrate Pathology

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Identification of the Functional Site in the Mosquito Larvicidal Binary Toxin of Bacillus sphaericus 1593M by Site-Directed Mutagenesis

Cited by 21 publications

References 24 publications

Essential role of tryptophan residues in toxicity of binary toxin from Bacillus sphaericus

Essential role of tryptophan residues in toxicity of binary toxin from Bacillus sphaericus

Role of cysteine at positions 67, 161 and 241 of a Bacillus sphaericus binary toxin BinB

Functional characterization of truncated fragments of Bacillus sphaericus binary toxin BinB

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