Study On The Interacting Protein With HCV Envelope Protein 2

The molecular mechanism responsible for hapetocellular damage caused by HCV infection is strongly associated with the interaction between virus proteins and hapetocellular proteins. The investigation on this possible protein-protein interaction with each peptide of HCV can partly explain the mechanism of HCV infection, and may provide some new clues for the prevention and cure of hepatitis C.To explore the possible function of HCV envelope protein 2 (HCV E2 ) after infected HCV, HCV E2 bait plasmid was constructed by ligating the gene of HCV E2 into pGBKT7, then transformed into yeast AH 109 (a type), the transformed yeast mated with yeast Y187 ( type) containing leukocyte cDNA library plasmid in 2 YPDA medium. Diploid yeast was plated on synthetic dropout nutrient medium containing X- -gal for selection and screening. After extracting and sequencing of plasmids from blue colonies, we underwent sequence analysis by bioinformatics. Some clones were obtained, and among them, a gene whose function is unknown was found, we named it HCV E2 binding protein 1 (HCV E2BP1). We investigated the biological functions of HCV E2BP1. The whole HCV E2BP1 gene sequence was amplified by PCR from human hepatocarcinoma cells. Expression vector pcDNA3.1(-) -HCV E2BP1 and pGBKT7- HCV E2BP1 was established. Advanced research was to screen proteins in hepatocytes interacting with HCV E2BP1 using yeast-two hybrid technique and suppression subtractive hybridization ( SSH) technique.In the study on the interacting with HCV Envelope protein 2, Seventy-six colonies were sequenced, among fourteen, a new gene with unknown function was identified. The second step , we investigated the biological functions of one of unknown function gene (HCV E2BP1) using yeast two-hybrid and suppression subtractive hybridization techniques. Some genes interacting with HCV Envelope protein 2 was obtained.It is the first step to investigated the biological functions of HCV E2 and HCV E2BP1, and pave a way for the following investigation ofthe more exact biological functions and the modulate of the gene expression.