Screening Of Interaction Proteins Of Hepatitis G Virus And Human T Lymphocyte

Hepatitis G virus(GB Virus C/Hepatitis G Virus,GBV-C/HGV)was found in 1995～1996,which is a positive strand RNA virus,and whole gene of 9.4,belonging to the Flaviviridae family Pegivirus genus.Route of transmission of GBV-C and HIV are very similar,GBV-C/HIV coin-infection rates is as high as 26.3%～33%.After clinical studies of many years,it found that GBV-C is not pathogenic,but co-infection GBV-C and HIV can delay the disease process of the HIV patients.The main Performance include decreased in patients with HIV viral load,increased the number of CD4+ cells in patients with HIV,extended the lives of HIV patients.In view of the fact that GBV-C itself is not pathogenic,but with HIV coin-infection is beneficial to slow down the progression of the disease in patients with HIV,it is necessary to clarify the mechanism of GBV-C inhibition of HIV,and it will be the key scientific problems GBV-C applied to HIV disease treatment.To determine protein of virus and shared host and its function is the key to the mechanism of interaction between the two viruses.Research on the mechanism of action that GBV-C can inhibit the proliferation of HIV-1 is mainly concentrated on the view which GBV-C block HIV-1 endocytosis process and change the T cell activity in two aspects.Although it is reported that the envelope glycoprotein E2 and NS5A peptide literature GBV-C can inhibit HIV-1 replication,it’s not clear that what the target protein of shared host have direct interaction with GBV-C,and how block HIV-1 endocytosis and affect T cell activity and effect,and what function in target proteins.In view of this,this paper uses the Gold yeast two hybrid technology for search proteins of T cell CDNA Library.First of all,this subject uses the Yunnan province main epidemic of type 7 GBV-C strain,and constructing the eight bait vector of GBV-C by PCR,restriction enzyme,connection,transformation technique.Restriction enzyme analysis and DNA sequence analysis showed that these vector was constructed successfully.Secondly,this topic through auxotrophic medium for culture,we show that successful construction of the eight bait vector are not self activation in yeast Y2H,in addition to pGBKT7-NS5A 85aa,and does not produce toxic to yeast Y2H,suitable for the fourth generation of Gold by yeast two hybrid system.Finally,taking the bait vector self activation and toxicity test successful as bait,by various of SD defects of medium culture screening T cell cDNA library.And positive clones were obtained with GBV-C-E1/E2/NS2 interacting protein.At the same time,The return of verification and trans activation of validation was successful.The results of sequencing by NCBI Blast analysis found that the 10 clones containing three cDNA sequences.And it has one kind of GBV-C E1 interacting protein-ATP6-4;two kinds of GBV-C E2 interacting protein,LT-α and PRKAR1;one kind of GBV-C NS2 interacting protein,PRKAR1A.GBV-C NS3/NS4/NS5 is not screened positive clones.In this study,screening to interaction protein of GBV-C,E1,E2,NS2 protein by yeast two hybrid experiment preliminary.Three proteins have not been reported.This is the mechanism of GBV-C and HIV on the next interaction laid solid foundation in T lymphocyte level.