Biosynthesis of the secretory core protein of duck hepatitis B virus: intracellular transport, proteolytic processing, and membrane expression of the precore protein.
The biosynthesis of the secretory core gene product of the duck hepatitis B virus (DHBe protein) was examined. Recombinant vaccinia viruses were constructed encoding either the full-length or C-terminally truncated forms of the DHBe precursor protein (precore protein) and used to express these proteins in the human hepatoma cell line HepG2. Western immunoblot analysis of core gene products isolated from cells producing the full-length precore protein revealed the presence of DHBe precursor proteins containing the strongly basic C-terminal sequence which is lacking in the mature DHBe protein. These proteins were not secreted, suggesting that C-terminal proteolytic processing of the precore protein represents an obligatory step for DHBe biosynthesis. Pulse-chase experiments showed that this cleavage reaction occurs late during DHBe synthesis. Interestingly, when mutated precore proteins were expressed which lacked the basic C-terminal domain, proteins were produced which were glycosylated but not secreted. This shows that the transient presence of this region is essential for intracellular transport of the precore protein. Cell sorter analyses revealed that production of a cell surface-expressed variant of the secretory core protein is a feature conserved between the duck and the human hepatitis B viruses. Surprisingly, the C terminus of the membrane-expressed DHBe protein was accessible from the outside, showing that the topology of this interesting protein is more complicated than expected.
Schlicht
H J
HJ
Department of Virology, University of Ulm, Germany.
D000595:Amino Acid Sequence; D001692:Biological Transport; D015153:Blotting, Western; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D003001:Cloning, Molecular; D005434:Flow Cytometry; D006512:Hepatitis B Core Antigens / Q000378:metabolism*; D015611:Hepatitis B Virus, Duck / Q000378:metabolism*; D006513:Hepatitis B e Antigens / Q000378:metabolism*; D006801:Humans; D066298:In Vitro Techniques; D008969:Molecular Sequence Data; D011498:Protein Precursors / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D014758:Viral Core Proteins / Q000378:metabolism*
D006512:Hepatitis B Core Antigens; D006513:Hepatitis B e Antigens; D011498:Protein Precursors; D014758:Viral Core Proteins