The 41-amino-acid C-terminal region of the hepatitis E virus ORF3 protein interacts with bikunin, a kunitz-type serine protease inhibitor.
Hepatitis E virus (HEV), a human plus-stranded RNA virus, contains three open reading frames (ORF). Of these, ORF1 encodes the viral nonstructural polyprotein, ORF2 encodes the major capsid protein, and ORF3 codes for a phosphoprotein of undefined function. Recently, using the yeast two-hybrid system to screen a human cDNA liver library, we have isolated and characterized AMBP (alpha1-microglobulin/bikunin precursor), which specifically interacts with the ORF3 protein of HEV. The ORF3 protein expedites the processing and secretion of alpha1-microglobulin. When checked individually for interaction, the second processed protein from AMBP, bikunin, strongly interacted with the full-length ORF3 protein. This protein-protein interaction has been validated by immunoprecipitation in both COS-1 and Huh7 cells and by His6 pull-down assays. In dual-labeling immunofluorescent staining, followed by fluorescence microscopy of transfected human liver cells, ORF3 colocalized with endogenously expressed bikunin. Finally, a 41-amino-acid C-terminal region of ORF3 has been found to be responsible for interacting with bikunin. The importance of this virus-host protein-protein interaction, with reference to the viral life cycle, has been discussed.
Tyagi
Shweta
S
Virology Group, International Centre for Genetic Engineering & Biotechnology, P. O. Box 10504, Aruna Asaf Ali Road, New Delhi 10067, India.
Surjit
Milan
M
Lal
Sunil K
SK
D000510:Alpha-Globulins / Q000235:genetics / Q000378:metabolism; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D019556:COS Cells; D002460:Cell Line; D002522:Chlorocebus aethiops; D004279:DNA, Viral / Q000235:genetics; D016752:Hepatitis E virus / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D066298:In Vitro Techniques; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D011499:Protein Processing, Post-Translational; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D015842:Serine Proteinase Inhibitors / Q000235:genetics / Q000378:metabolism*; D014360:Trypsin Inhibitor, Kunitz Soybean / Q000235:genetics / Q000378:metabolism*; D020798:Two-Hybrid System Techniques; D014764:Viral Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
D000510:Alpha-Globulins; D004279:DNA, Viral; D008562:Membrane Glycoproteins; C400973:ORF3 protein, Hepatitis E virus; D011994:Recombinant Proteins; C109148:SPINT2 protein, human; D015842:Serine Proteinase Inhibitors; D014764:Viral Proteins; C016001:alpha-1-microglobulin; D014360:Trypsin Inhibitor, Kunitz Soybean