Molecular Simulations Of The Interactions Between ACCase And Herbicides And Computational Design Of Novel Herbicides

Since the 1940s,herbicide has been used for control of weeds in the farmland.But along with the long-term and big area use,the weed's resistance appeared,especially for herbicide only having the single target.And then,it did not easy to control weed by using the general method.At the same time, the service live of many herbicides was reduced.For example,A.myosuroides,which is resistant to diclofop in the Australian wheat field,shows cross-resistance to many herbicides,damages the area to reach 0.5×108hm²,and damages other crops.Therefore,studying resistant mechanism becomes very important.And then designing anti-resistance inhibitor would be an arduous task for researchers.ACCase is the key enzyme in the plant metabolism process catalyzing the plant fatty acid biosynthesis.It contains three subunits:BC,CT and BCCP.The CT subunit is our research category, whose biological function is mainly to catalyze acetyl-CoA to form malonyl-CoA,and then forms the fatty acid under the Fattyacidsynthetase.APPs and CHDs are the two kind of mainly inhibitor type for CT.As early as in 1980s,they had been widely used for control gramineous weed,and they had caused the resistant weed's appearance finally.So far,already discovered that some 35 kind of gramineous weed,mainly distributing in the US,Canada,Australia and so on 14 countries,have showed resistance to APPs and CHDs.The resistant mechanism mainly contains two types:target enzyme mutation and metabolism acceleration.And the former is the main resistant mechanism. Many resistant sites had been reported in the literatures.As to higher plant,the 3D-structure of CT and its binding model with inhibitor were unknown,and the resistant mechanism research in molecular level is unable to carry out.Until 2003,Tong.L et al reported for the first time the crystal structure of CT from yeast in the form of free enzyme and complex with inhibitors,which has laid the foundation for the research in the higher plant.This dissertation may be summarized as follows:Firstly,through the literature consult and the network search for CT's primary sequence,we select the sensitive and resistant CT sequence from foxtail millet to carry on the homology modeling. The two sable conformations were obtained followed by the dimer construction and the molecular dynamics simulation.Comparing the conformation of these two models,we found that the spatial orientation of residue 695 is different from each other.In foxACC-2S,the Ile-695 extends its side chain outside the active site,while Leu-695 in foxACC-2R extended its side chain toward the cavity of the active site,inhibiting the entrance of ligand.It is the first time we elaborated from the molecular level the resistant mechanism of I695L.Simultaneously,using molecular docking,we study the interaction between the sensitive CT and APPs inhibitors,and discover two important amino acid residues(Ser-698 and Tyr-728),which would laid the foundation for the reasonable medicine design.Secondly,according to Délye et al reports in 2005,selects the sequence of CT from A. myosuroides to be used for homology modeling.The two models(AJ_free and AJ_com) were obtained alter molecular dynamics simulation.The difference between AJ_free and AJ_com in conformation is consistent with the difference between the crystal structures.The binding model in AJ_com is similar to the crystal complex structure.These results had proven the models we obtained are credible.And then,based on manual docking and molecular dynamics methods,the binding model between CT and four APPs inhibitors were obtained.We carried on the MM/PBSA and entropy computations for every system.The tendency obtained from computing binding free energy is consistent with the experiment value.So,from another point of view,these results had proven the model accuracy and the binding model's accuracy,simultaneously,also showed the method of manual docking for similar skeleton compound was feasible.And the binding models of four APPs inhibitors were used in the following resistance mechanism research.Thirdly,combining with the amino acid mutation directly and molecular dynamics simulation, the resistant mechanism of four resistant sites(W374C,I388N,D425G and the G443A) for each compound were studied.There are all sixteen mutated systems.We carried on the MM/PBSA and entropy computation for each system.The results show that the binging free energy was reduced by enthalpy and entropy for each mutated system.The enthalpy mainly manifests for VDW interaction or the Hbond interaction between acceptor and ligand,the entropy mainly represents by the conformation entropy.Meanwhile,as to D425G and G443A,belonging to the non-active site,they firstly changed or broken the Hbond network of 388-446,then led the residues in the active site to have the conformation change.Therefore,the binding free energy was reduced by VDW or Hbond interaction between receptor and ligand.The resistant mechanism provides the theory instruction for anti-resistant design.Finally,we randomly selected two mutated system for re-designing anti-resistant compound. Based on not changing the inhibitor's skeleton,we substituted different groups in order to increasing the binding free energy between receptor and ligand.The results show that the enthalpy would increase by substituting o-Cl to-CN for the haloxyfop,and the entropy would increase by substituting o-Cl to -CH₃ for diclofop.When the binding free energy increased,the resistance would be overcome. All these results would be helpful for synthesis anti-resistant herbicide.