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Interaction of Destruxin A with Three Silkworm Proteins: BmCRT, BmDPP3, and BmPDIA5
Xuyu Yin1, Haitao Peng1, Qunfang Weng1
1Key Laboratory of Bio-Pesticide Innovation and Application of Guangdong Province, College of Plant, Protection, South China Agricultural University, Guangzhou 510642, China.
Abstract:
Destruxin A (DA), a hexa-cyclodepsipeptidic mycotoxin produced by the entomopathogenic fungus Metarhizium anisopliae, has insecticidal activity, but its molecular mechanism of action is still not clear. Three proteins with modification-related functions, calreticulin (BmCRT), dipeptidyl peptidase Ⅲ (BmDPP3), and protein disulfide isomerase A5 (BmPDIA5), were selected to verify the interactions with DA in this study. The kinetic data of the interactions were measured by surface plasmon resonance (SPR) and bio-layer interferometry (BLI) in vitro. The KD values of DA with BmCRT, BmDPP3, and BmPDIA5 ranged from 10-4 to 10-5 mol/L, which suggested that the three proteins all had fairly strong interactions with DA. Then, it was found that DA in a dose-dependent manner affected the interactions of the three proteins with their partners in insect two-hybrid tests in SF-9 cells. Furthermore, the results of enzyme activities by ELISA indicated that DA could inhibit the activity of BmDPP3 but had no significant effect on BmPDIA5. In addition, DA induced the upregulation of BmDPP3 and the downregulation of BmCRT. The results prove that BmCRT, BmDPP3, and BmPDIA5 are all binding proteins of DA. This study might provide new insights to elucidate the molecular mechanism of DA.
Insights
Destruxin A (DA), a mycotoxin, binds strongly to three insect proteins: calreticulin (BmCRT), dipeptidyl peptidase III (BmDPP3), and protein disulfide isomerase A5 (BmPDIA5), revealing potential insecticidal mechanisms.
Area of Science:
- Biochemistry
- Molecular Biology
- Mycotoxicology
Background:
- Destruxin A (DA) is a mycotoxin from *Metarhizium anisopliae* with insecticidal properties.
- The precise molecular mechanism underlying DA's insecticidal activity remains largely unelucidated.
- Understanding DA's interactions with host proteins is crucial for deciphering its mode of action.
Purpose of the Study:
- To investigate the interaction between Destruxin A (DA) and three specific insect proteins: calreticulin (BmCRT), dipeptidyl peptidase III (BmDPP3), and protein disulfide isomerase A5 (BmPDIA5).
- To elucidate the functional consequences of these interactions on protein activity and expression.
- To provide insights into the molecular basis of DA's insecticidal effects.
Main Methods:
- Surface Plasmon Resonance (SPR) and Bio-Layer Interferometry (BLI) were employed to measure the kinetic binding parameters (KD) between DA and the target proteins.
- Insect two-hybrid assays in SF-9 cells were conducted to assess the impact of DA on protein-protein interactions in a cellular context.
- Enzyme-linked immunosorbent assays (ELISA) were used to evaluate the effects of DA on the enzymatic activities of BmDPP3 and BmPDIA5.
- Quantitative analysis of protein expression levels (upregulation/downregulation) was performed.
Main Results:
- DA exhibited strong binding affinities (KD values in the 10-4 to 10-5 mol/L range) with BmCRT, BmDPP3, and BmPDIA5.
- DA modulated protein-protein interactions involving BmCRT, BmDPP3, and BmPDIA5 in a dose-dependent manner.
- DA inhibited the enzymatic activity of BmDPP3 but did not significantly affect BmPDIA5.
- DA induced the upregulation of BmDPP3 and the downregulation of BmCRT expression.
Conclusions:
- BmCRT, BmDPP3, and BmPDIA5 are identified as direct binding partners of Destruxin A.
- DA's interaction with these proteins influences their activity and expression levels, contributing to its insecticidal effects.
- This study provides novel molecular insights into the mechanism of action of Destruxin A, potentially paving the way for targeted pest control strategies.
