Expression, Interactive Protein Screening And Identification Of Hepatitis E Virus ORF3

Hepatitis E virus encodes three open reading frames, ORF1, ORF2and ORF3. ORF1encodes viral non-structural proteins, which are very important for viral replication.ORF2encodes viral capsid protein and ORF3encodes an around120amino acids ofphosphorylation protein that associates with cytoskeleton, while its precise biologicalfunction is still unknown.In this study, to understand the function of Hepatitis E virus, we firstly expressed theORF3protein of hepatitis E virus via baculovirus and mammalian expression systems.In order to further understand its functions, mammalian expression system was used toclone and express the RPS11, HPN, ALB, HRG and PIK3CD, basing on our previousyeast two-hybrid screening of hepatitis E virus ORF3interacting proteins in hepatocytes.Five ORF3interacting protein candidates were tested by co-immunoprecipitation. Wefound that, HPN, a transmembrane serine protease that was widely reported to functionas the inhibitor of several tumors, interacts with ORF3. We inferred that the ORF3protein of Hepatitis E virus may promote the tumorigenesis via interacting with hepsin.Also, we made use of the cloned and expressed of ORF3protein to screen and identifyseven ORF3candidate interacting proteins by combining co-immunoprecipitation andliquid chromatography-mass spectrometry methods. Our results provided us a new clueand richer perspective to understand the function of the ORF3of Hepatitis E virus.