Research On The Resistance And Fish Toxicity Mechanism Of Phenylpyrazole Insecticides Based On Structural Modeling Study Of GABA_A Receptors

In this thesis,a series of computational methods such as the homology modeling,docking and molecular dynamics simulation were integrated to study the different resistance mechanism between the wild type and mutant(A2'S and R0'Q)Nilaparvata lugens Rdl-GABA receptor with fipronil,and to explore the key roles of the mutant residues.Secondly,the structures of zebrafish GABAA receptors were built by homology method and to explore the different mechanism of fipronil and flufiprole with zebrafish GABAA receptors.In chapter one,the research development of phenylpyrazole insecticide,as well as the computational methods referred in this thesis were reviewed briefly.In chapter two,homology modeling,docking and molecular dynamics simulations were combined to explore the different resistance mechanism of fipronil with Nilaparvata lugens Rdl-GABA receptors,caused by the different mutant residue in the binding pocket.It was found that,in the WT system,the trifluoromethyl group of the benzene ring faced to the intracellular domain,with a H-bond between 6'Thr and fipronil exerting key effects on the recognition of fipronil.The A2'S mutation displayed weak influence on fipronil binding pocket,the pore diameter and the binding mode were similar to the WT system,though the corresponding H-bond interaction was replaced and weakened by the 2'Ser.However,the R0'Q mutation deeply influenced on the fipronil binding pocket,the pore diameter and the binding mode were different from the WT and A2'S systems.It showed that the trifluoromethyl group of the benzene ring faced to the extracellular domain.Moreover,the H-bond interaction was absent.It indicated that the A2'S might be considered a risk signal and RO'Q should be a direct evidence for high fipronil resistance.In chapter three,the zebrafish GABAA receptors were constructed by homology modeling based on 3RHW and 4COF,respectively.Then,the two homology models were optimized by molecular dynamics simulation,achieving an open state zebrafish GABAA receptor based on 4COF.Finally,the molecular docking and MM-GBSA was applied to explore the different mechanism of fish toxicity between fipronil and flufiprole with zebrafish GABAA receptor based on 4COF.It indicated that the average Glide Score(-7.60 and-6.57)and MM-GBSA(-72.74Kcal/mol and-53.93Kcal/mol)for fipronil and flufiprole,respectively,well indicated the weak binding affinity of flufiprole,which might result in the low fish toxicity.Meanwhile,the key residues 2'Ala,6'Thr and 9'Leu,which had been reported in previous studies,displayed key effects on the recognition of fipronil and flufiprole.Also,our study revealed that residue-2'Ala might be responsible for the interaction between the fipronil and flufiprole with zebrafish GABAA receptor.Moreover,the-1'Ala and 5'Ile in the TM2 domain played an important role in the fipronil binding but was absent in the flufiprole binding,which might be responsible to the different fish toxicity caused by fipronil and flufiprole.These findings might be helpful in designing selective insecticides against pests versus fishes.In chapter four,the whole thesis was briefly summarized.