Computational and Experimental Approaches to Decipher the Binding Mechanism of General Odorant-Binding Protein 2 from Athetis lepigone to Chlorpyrifos and Phoxim

Abstract

Insect resistance to insecticides is an increasingly serious problem, and the resistant mechanisms are complicated. The resistance research based on the chemosensory pathway is one of the hot problems at present, but the specific binding mechanism of chemosensory genes and insecticides remains elusive. The binding mechanism of AlepGOBP2 (belong to insect chemosensory gene) with two insecticides was investigated by computational and experimental approaches. Our calculation results indicated that four key residues (Phe12, Ile52, Ile94, and Phe118) could steadily interact with these two insecticides and be assigned as hotspot sites responsible for their binding affinities. The significant alkyl−π and hydrophobic interactions involved by these four hotspot residues were found to be the driving forces for their binding affinities, especially for two residues (Phe12 and Ile94) that significantly contribute to the binding of chlorpyrifos, which were also validated by our binding assay results. Furthermore, we also found that the AlepGOBP2–chlorpyrifos/phoxim complexes can be more efficiently converged in the residue-specific force field-(RSFF2C) and its higher accuracy and repeatability in protein dynamics simulation, per-residue free energy decomposition, and computational alanine scanning calculations have also been achieved in this paper. These findings provided useful insights for efficient and reliable calculation of the binding mechanism of relevant AlepGOBPs with other insecticides, facilitating to develop new and efficient insecticides targeting the key sites of AlepGOBP2.

Ligand‐binding properties of odorant‐binding protein 6 in Athetis lepigone to sex pheromones and maize volatiles

“…In accordance with previous methods, 21,37–39 purified PCR products of AlepOBP6 were first subcloned into the pEASY ‐ Blunt3 cloning vector (TransGen). The expression vectors pET‐30a(+) (Novagen) and pEASY‐Blunt3‐AlepOBP6 plasmid were then digested using FastDigest® restriction enzymes BamHI and XhoI (Fermentas), respectively.…”

“…A Spectra Max M2 fluorescence spectrophotometer (Molecular Devices) was used for the fluorescence competitive binding assays, as previously described 37,38,40 . At the beginning, the fluorescent probe N ‐phenyl‐1‐naphthylamine (1‐NPN) and ligand (sex pheromones and maize volatiles) were dissolved in chromatography‐grade methanol to give a 1 m m solution.…”

Genome-Wide Analysis of Odorant-Binding Proteins in Papilio xuthus with Focus on the Perception of Two PxutGOBPs to Host Odorants and Insecticides

“…53 In several previous studies, the two odorants have been found to strongly bind olfactory-related proteins including GOBPs. 18,19,54,55 In Athetis lepigone, AlepGOBP2 exhibited a high affinity to chlorpyrifos, of which its two sites (Phe12 and Ile94) were of particular importance in the binding of this insecticide, but not including Ile52 and Phe118. 19 Our analysis on the PxutGOBP1 mutation from Phe118 to Ala showed a similar result with the mutated AlepGOBP2 Phe118Ala , i.e., they exhibited a slightly reduced binding affinity to chlorpyrifos.…”

Structure Based Affinity Maturation and Characterizing of SARS-CoV Antibody CR3022 against SARS-CoV-2 by Computational and Experimental Approaches

“…The optimized structure was fully dissolved in a rectangular water box filled with TIP3P (size 10.0 Å × 14.5 Å × 10.0 Å) [35,36]. The RSFF2C force field was applied for MD calculation [37]. All systems were equilibrated by NVT and NPT for 500 ps [29,38].…”