Study Of Bioinformation Analysis Of EP0 Gene And Homology Model Establishment Of TK Protein

There are 12 pseudorabies virus strain,SS,SQ and FA,783strain,Bartha,(purchased from China Institue of Veterinary Drug Control) and SA215(purchased from China Animal Husbandry Industry CO.LTD)and SN,SL,SCZ(Isolated by Xu Zhiwen associate professor in Sichuan Agriculture University Animal biotechnology center in2000) and BJ,DG,HS (gived by Lou Gaoming professor in Shaoguan University College of Yingdong Biorngineering).Designing a pair of primer according to PRV EP0 gene,we received 12 EP0 complete gene by PCR.Using pseudorabies virus strain FA,We received a TK gene by PCR.The TK gene was transformed into E.coli DH5αwith TaKaRa's kit to retrieve gel and to ligate.After identified by COL-PCR and RE,they were sequenced by TaKaRa and GeneCore.12 sequenced EP0 genes of different PRV strains,as well as 3 EP0 genes which downloaded from GenBank were analyzed by bioinformatics software,the analysis and prediction of these genes including nucleotide homology,codons bias,location of mutation area,phylogenetic tree,amino acids homology,protein hydrophilicity and epitope. Meanwhile,the nucleotide homology,amino acids homology and the protein domain of EP0 genes of pseudorabies virus strain FA,Herpesvirus I,BHV I,EHV I,MDV I and varicella-herpes zoster virus were analysed.The results showed that the ORF length of PRV-EP0 gene is 1230-1233nt,amino acids length is 409-410,nucleotide homology is 97.6%-99.9%,and amino acids homology is 96.6%-100%.One hypermutation replicated plot is located in 684～688nt.Different strains were prefer to GC basi-.The results of different strains of enzyme site,protein hydrophilicity and epitope were very similar.The nucleotide and amino acids homology of Herpesvirus was 1.4%～41.4%and 5.9%～42% respectively. With PRV FA as subject,TK gene has been cloned and sequence analysed.We searched for homology protein through FASTA procedure on Brookhaven protein data bank(PDB) and then established its 3D model by the method of homology model establishment and molecular dynamic analysis.Validity of such model was verified by Ramachandran and Profile-3D procedure.The acive site of TK had been accurately located with InsightⅡ/Binding site and Delphi procedures.Based on this 3D profile,we designed inhibitor(N-phenyl-N'- methylurea) of TK,and clarified the interactive mode between the active site and its inhibitor.The objective of this study was to study the background of PRV-EP0 gene based on molecular level and bioinformation analysis and provide a base for further study of effects of TK gene on latent infection of PRV.With 3D model,we have predicted the active domain of TK protein and designed its ligand and discussed the inhibitory effect between the ligand and protein TK.It is possible that a new therapeutic procedure for Pseudorabies could be presented.