Igand pose program and ranked by Consensus score program. Finally, the optimal 3D binding conformations of complexes were selected and shown in Figure 7. The interaction energies between key residues and the ligand are listed in the Table 1. All of these residues are located in the cavity formed by six helices. Several residues including Ile45, Leu49, Thr64, Leu68 and Leu82 identified by current docking simulations have been shown to play an important role in the binding of CSPSlit and rhodojaponin III. Figure 8 shows the time series of potential energy and RMSD of backbone for 1000 ps MD simulation of CSPSlit structure. The potential energy of the complex was stabilized at 200 ps production after 800 ps equilibration, and the RMSD of backbone compared to the ?starting coordinate remained at 0.14 A up and down fluctuations. These two properties converged at production, indicating that the complex is stable.Table 1. VdW Energy (EvdW) and Electrostatic Energy (Eele) between rhodojaponin III and CSPSlit.Residue Leu41 Ile45 Ala48 Leu49 Cys53 Cys56 Gln60 Thr64 Val67 Leu68 Leu71 Trp79 Leu82 Cys83 Tyr86 Asp87 TyrEvdW (kcal/mol) 20.830793 23.700930 21.080190 22.234000 20.358571 20.318546 21.109890 22.165880 22.477020 22.773280 20.885637 0.174320 22.684090 21.524360 21.640490 20.697000 20.Eele (kcal/mol) 0.017039 20.346153 0.739731 20.098858 0.154808 20.240461 0.331317 20.800064 0.549604 20.523853 0.179657 0.187195 0.219521 0.297607 0.447274 20.607238 0.3.5 Binding Assay of Rhodojaponin III and CSPSlitPurified CSPSlit protein solution contained 5.0 mg/ml was used to analyse the binding property of CSPSlit with rhodojaponin III. When excited at 295 nm, the fluorescence emission spectra showed maximally relative fluorescence intensity at 383.6 nm for CSPSlit. Following, with the increasing concentration of rhodojaponin III, CSPSlit peak underwent a blue shift, but no peak intensity increaseing was observed. When the final concentration of rhodojaponin III was 600 mM and 300 mM, the fluorescence intensity decreased to 29.43 and 18.47 , respectively (Fig. 9). These results showed that CSPSlit could be intensely bound with rhodojaponin III.DiscussionThe olfactory system of insects is essential for Lepidoptera as well as in other insect orders to initiate behavioral responses, suchdoi:10.1371/journal.pone.0047611.tCharacterisation Binding Properties of CSPSlitFigure 8. Potential energy (A) and root-mean-square deviation (B) with respect to simulation time for 1000 ps MedChemExpress NT-157 molecular dynamics simulation on the CSPSlit- rhodojaponin III complex model. doi:10.1371/journal.pone.0047611.gas searching for food sources, mating, oviposition and feeding [52?53]. Chemosensory proteins (CSPs), known as another class of soluble protein, share no sequence homology with either PBPs or general OBPs of many insects [11?2,18]. The CSPs are smaller proteins which contain four cysteines instead of six with 57773-63-4 site conserved interval spacing involved in two disulfide bonds [35,54]. In the present study, a cDNA sequence encoding the CSP of S. litura was cloned. The CSPSlit has 4 typical conservative cysteines in the sequence (Fig.1). It is consistent with previous report. The CSPSlit was expressed in antennae, legs, wings and female abdomens (Fig.3), these results is similar with the research in other insect [9,21]. In M. brassicae, the CSPs has a abundant expression in proboscis [15,30], but in this study, there is no hybridization signal was detected in the de-antennated head (Fig.Igand pose program and ranked by Consensus score program. Finally, the optimal 3D binding conformations of complexes were selected and shown in Figure 7. The interaction energies between key residues and the ligand are listed in the Table 1. All of these residues are located in the cavity formed by six helices. Several residues including Ile45, Leu49, Thr64, Leu68 and Leu82 identified by current docking simulations have been shown to play an important role in the binding of CSPSlit and rhodojaponin III. Figure 8 shows the time series of potential energy and RMSD of backbone for 1000 ps MD simulation of CSPSlit structure. The potential energy of the complex was stabilized at 200 ps production after 800 ps equilibration, and the RMSD of backbone compared to the ?starting coordinate remained at 0.14 A up and down fluctuations. These two properties converged at production, indicating that the complex is stable.Table 1. VdW Energy (EvdW) and Electrostatic Energy (Eele) between rhodojaponin III and CSPSlit.Residue Leu41 Ile45 Ala48 Leu49 Cys53 Cys56 Gln60 Thr64 Val67 Leu68 Leu71 Trp79 Leu82 Cys83 Tyr86 Asp87 TyrEvdW (kcal/mol) 20.830793 23.700930 21.080190 22.234000 20.358571 20.318546 21.109890 22.165880 22.477020 22.773280 20.885637 0.174320 22.684090 21.524360 21.640490 20.697000 20.Eele (kcal/mol) 0.017039 20.346153 0.739731 20.098858 0.154808 20.240461 0.331317 20.800064 0.549604 20.523853 0.179657 0.187195 0.219521 0.297607 0.447274 20.607238 0.3.5 Binding Assay of Rhodojaponin III and CSPSlitPurified CSPSlit protein solution contained 5.0 mg/ml was used to analyse the binding property of CSPSlit with rhodojaponin III. When excited at 295 nm, the fluorescence emission spectra showed maximally relative fluorescence intensity at 383.6 nm for CSPSlit. Following, with the increasing concentration of rhodojaponin III, CSPSlit peak underwent a blue shift, but no peak intensity increaseing was observed. When the final concentration of rhodojaponin III was 600 mM and 300 mM, the fluorescence intensity decreased to 29.43 and 18.47 , respectively (Fig. 9). These results showed that CSPSlit could be intensely bound with rhodojaponin III.DiscussionThe olfactory system of insects is essential for Lepidoptera as well as in other insect orders to initiate behavioral responses, suchdoi:10.1371/journal.pone.0047611.tCharacterisation Binding Properties of CSPSlitFigure 8. Potential energy (A) and root-mean-square deviation (B) with respect to simulation time for 1000 ps molecular dynamics simulation on the CSPSlit- rhodojaponin III complex model. doi:10.1371/journal.pone.0047611.gas searching for food sources, mating, oviposition and feeding [52?53]. Chemosensory proteins (CSPs), known as another class of soluble protein, share no sequence homology with either PBPs or general OBPs of many insects [11?2,18]. The CSPs are smaller proteins which contain four cysteines instead of six with conserved interval spacing involved in two disulfide bonds [35,54]. In the present study, a cDNA sequence encoding the CSP of S. litura was cloned. The CSPSlit has 4 typical conservative cysteines in the sequence (Fig.1). It is consistent with previous report. The CSPSlit was expressed in antennae, legs, wings and female abdomens (Fig.3), these results is similar with the research in other insect [9,21]. In M. brassicae, the CSPs has a abundant expression in proboscis [15,30], but in this study, there is no hybridization signal was detected in the de-antennated head (Fig.

Igand pose program and ranked by Consensus score program. Finally, the

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