Structure-Based Analysis of the Ligand-Binding Mechanism for DhelOBP21, a C-minus Odorant Binding Protein, from Dastarcus helophoroides (Fairmaire; Coleoptera: Bothrideridae)

Abstract

Odorant binding proteins (OBPs) transport hydrophobic odor molecules across the sensillar lymph to trigger a neuronal response. Herein, the Minus-C OBP (DhelOBP21) was characterized from Dastarcus helophoroides, the most important natural parasitic enemy insect that targets Monochamus alternatus. Homology modeling and molecular docking were conducted on the interaction between DhelOBP21 and 17 volatile molecules (including volatiles from pine bark, the larva of M. alternatus, and the faeces of the larva). The predicted three-dimensional structure showed only two disulfide bridges and a hydrophobic binding cavity with a short C-terminus. Ligand-binding experiments using N-phenylnaphthylamine (1-NPN) as a fluorescent probe showed that DhelOBP21 exhibited better binding affinities against those ligands with a molecular volume between 100 and 125 ų compared with ligands with a molecular volume between 160 and 185 ų. Molecules that are too big or too small are not conducive for binding. We mutated the amino acid residues of the binding cavity to increase either hydrophobicity or hydrophilia. Ligand-binding experiments and cyber molecular docking assays indicated that hydrophobic interactions are more significant than hydrogen-bonding interactions. Although hydrogen-bond interactions could be predicted for some binding complexes, the hydrophobic interactions had more influence on binding following hydrophobic changes that affected the cavity. The orientation of ligands affects binding by influencing hydrophobic interactions. The binding process is controlled by multiple factors. This study provides a basis to explore the ligand-binding mechanisms of Minus-C OBP.

Optimization of reverse chemical ecology method: false positive binding of Aenasius bambawalei odorant binding protein 1 caused by uncertain binding mechanism

“…However, the n value of (−)‐limonene at pH 7.4 as well as those of 2, 4, 4‐trimethyl‐2‐pentene and 1‐octen‐3‐ol under all conditions were much lower than 1 and close to 0.5, indicating a high possibility that two proteins bind to one ligand. In addition, for 1‐octen‐3‐one, the n value was not close to either 0.5 or 1 except for the condition of pH 7.4 at 288 K. Then, thermodynamic equations were used to calculate the thermodynamic parameters and determine the binding force (Li et al ., ): , where Δ G , Δ H and Δ S are the free energy change, enthalpy change and entropy change, respectively T, temperature; R, thermodynamic constant; K, equilibrium constant. If the temperature was slightly changed, the enthalpy change was always regarded as a constant.…”

“…Fluorescence quenching assays are considered as a reliable method to measure the intrinsic fluorescence of proteins, which is mainly produced by Trp, Tyr and Phe (Van de Weert & Stella, 2011). Additional calculations, such as of the Stern-Volmer equation, double logarithm equation and thermodynamic parameters, can contribute to revealing the binding mechanism of OBPs with ligands (Li et al, 2015).…”

“…After centrifugation, the protein presented as inclusion bodies, which were solubilized by the addition of 10 ml 8 M urea in 50 mM Tris buffer (pH 7.4), incubated in 1 mM dithiothreitol, followed by renaturation and extensive dialysis against 30 mM Tris buffer at pH 7.4. The protein was purified by combinations of chromatographic steps on anionexchange resins as described previously (Li et al, 2015). All purification steps were monitored by SDS-PAGE.…”

“…Studies of the binding affinities and structural characteristics of OBPs and their ligands could provide substantial information compared with sequence studies. Recently, molecular docking simulation involving OBP homology models was proved to be a reliable and easy approach for studying OBP structures (Venthur et al, 2014;Li et al, 2015). Additionally, fluorescence quenching has become an important tool to investigate various aspects of ligand binding (Weert & Stella, 2011).…”

Molecular characterization and functional analysis of pheromone binding protein 1 from Cydia pomonella (L.)

“…By reference to former studies, it is reasonable to state that the length of ligand backbones plays a key role in determining the affinity between ligands and lepidopteran PBPs. A ligand with a shorter chain usually results in a smaller hydrophobic interaction which is vital to the interaction between PBPs and ligands, whereas a ligand with too many carbons in the chain may go beyond the capacity of binding pockets (Li et al, 2015;Tian et al, 2016). Codlemone, 1-dodecanol and E,E-2,4-dodecadienal are characterized by having the hydroxyl or carbonyl group at one end of each molecule as well; previous studies have revealed that such functional groups tend to form hydrogen bonds with the sidechains of PBP residues (Sandler et al, 2000;Tian et al, 2016).…”