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  • Specific involvement of two amino acid residues in cis-nerolidol binding to odorant-binding protein 5 AlinOBP5 in the alfalfa plant bug, Adelphocoris lineolatus (Goeze).

Specific involvement of two amino acid residues in cis-nerolidol binding to odorant-binding protein 5 AlinOBP5 in the alfalfa plant bug, Adelphocoris lineolatus (Goeze).

Insect molecular biology (2013-01-09)
S-Y Wang, S-H Gu, L Han, Y-Y Guo, J-J Zhou, Y-J Zhang
ABSTRACT

Olfaction plays an important role in insects' survival and reproduction. Odorant-binding proteins (OBPs) are considered to be one of the crucial proteins in the insect olfactory pathway. In this study, an antenna-specific OBP of the alfalfa plant bug, Adelphocoris lineolatus AlinOBP5, was expressed and purified in vitro. The binding affinities of AlinOBP5 with sex pheromone analogues of the Miridae and cotton volatiles were investigated by fluorescence competitive binding assays. The binding sites of AlinOBP5 were predicted by three-dimensional structure modelling and molecular docking, and site-directed mutagenesis. AlinOBP5 could not effectively bind with sex pheromone analogues of Miridae but showed high binding abilities with specific cotton volatiles, such as cis-nerolidol, ethyl laurate, β-ionone, β-caryophyllene, 2,3-dimethylbenzoic acid and (E)-farnesol. The strongest binding affinity was to cis-nerolidol, suggesting a role of AlinOBP5 in general odorant chemoreception. Based on the relatively strong binding affinity and the reported physiological activity of cis-nerolidol in other insects, we chose cis-nerolidol for further homology modelling and ligand docking. The results of molecular simulation and site-directed mutagenesis indicated that two amino acids, Lys74 and Pro121, in the protein binding pocket are the key amino acids involved in the binding of cis-nerolidol. The Lys74 residue may participate in specific recognition of ligands, and the Pro121 residue plays a crucial role in ligand binding and release by changing the binding pocket environment and stabilizing the conformation of the C-terminus of AlinOBP5.

MATERIALS
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Supelco
trans-Nerolidol, analytical standard
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Nerolidol, mixture of cis and trans, ≥97%, stabilized, FG
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Nerolidol, 98%, Mixture of cis and trans
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Sigma-Aldrich
2,3-Dimethylbenzoic acid, 98%
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