The Virtual Screening And Activity Vertification Of Novel Inhibitors Targeting Ap? 3C Protease

Antheraea pernyi iflavirus(ApIV)is an important cause of antheraea pernyi vomiting disease(AVD)which is seriously harming the production of Antheraea pernyi(A.pernyi)industry in China.This disease usually occurred in Liaoning,Jilin,Heilongjiang and other provinces which the climate is cold during the autumn.Usually,incidence rate of AVD is 30%,and even more than 70%-80% in serious areas.ApIV is a single-stranded,positive-sense RNA virus,belong to the family Picornavirida.Due to its significant role in virus replication,conservative sequence and no homologous sites in the host cell,3C protease is considered to be an attractive drug target for developing antiviral therapeutic agents.Although great efforts have been made to develop 3C protease inhibitors,no effective anti-viral therapy for the prevention or treatment of diseases caused by ApIV infection is available.In this study,homology modeling and molecular dynamics(MD)simulation were carried out to generate the 3D structure of ApIV 3C protease.15 compounds were identified through virtual screening using molecular docking against natural product library.Then an effective inhibitor of ApIV 3C protease-Orobol was identified through cell-based assay and in vivo assay.The cell-based assay displayed that the inhibition ratio of virus replication was stronger with the increase of the concentration of Orobol.And when the concentration was 10?g/ml,the inhibition ratio reached 80.5%.When the concentration of orobol was 10?g/ml,the cell cytotoxicity caused by orobol was only 3.42%.In vivo assay demonstrated that the facial window of the positive control pupae became black and the the facial window of the negetive control pupae became black,while the facial window of pupae infected with different concentration Orobol gradually turn white with the increase of the dose of the compound.The cell-based assay and in vivo assays demonstrated that Orobol has anti-ApIV viral activity.Thus a lead compound of ApIV 3C protease inhibitors was discoveried.After the shake flask fermentation of streptomyces avermitilis NRRL8165 were studied,Orobol was obtained by biotransformation.And the binding mode of Orobol with ApIV 3C protease was stable through molecular docking and MD analysis.Those results proposed that Orobol was a novel inhibitor for the disease of antheraea pernyi caused by ApIV infections.Orobol was modified by Grow Scaffold in Dscovery Studio(DS)in order to improve its activity against anti-ApIV.Six modified compounds were obtained,the results showed that the binding energy of the modified compounds were significantly reduced.