Hepatitis C Virus Core Protein: Carboxy-Terminal Boundaries of Two Processed Species Suggest Cleavage by a Signal Peptide Peptidase

The expression and processing of hepatitis C virus core protein was analyzed. Two protein bands, 21 kDa (P21), corresponding to the full-length core, and 19 kDa (P19), were detected as major products when core protein was expressed in the standard rabbit reticulocyte lysate system or inSf9insect cel...

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Bibliographic Details
Published in:Virology (New York, N.Y.) N.Y.), 1996-10, Vol.224 (1), p.93-104
Main Authors: Hüssy, Peter, Langen, Hanno, Mous, Jan, Jacobsen, Helmut
Format: Article
Language:English
Subjects:
Amino Acid Sequence
Animals
Binding Sites
Cell Line
Chromosome Mapping
Hepacivirus - genetics
Hepacivirus - metabolism
Membrane Proteins
Molecular Sequence Data
Mutation
Protein Biosynthesis
Protein Precursors - genetics
Protein Precursors - metabolism
Protein Processing, Post-Translational
Protein Sorting Signals - metabolism
Rabbits
Recombinant Fusion Proteins
Serine Endopeptidases - metabolism
Spodoptera - cytology
Tetrahydrofolate Dehydrogenase - genetics
Tetrahydrofolate Dehydrogenase - metabolism
Viral Core Proteins - genetics
Viral Core Proteins - metabolism
Viral Envelope Proteins - genetics
Viral Envelope Proteins - metabolism
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Summary:The expression and processing of hepatitis C virus core protein was analyzed. Two protein bands, 21 kDa (P21), corresponding to the full-length core, and 19 kDa (P19), were detected as major products when core protein was expressed in the standard rabbit reticulocyte lysate system or inSf9insect cells. Core proteins with amino-terminal hexa-histidine tags were expressed which allowed the purification of the hexa-histidine-P19 core with Ni2+-NTA columns. With the help of mass spectrometry, the molecular weight of hexa-histidine-P19 was analyzed and its carboxy-terminus could be calculated. Fusion proteins of truncated core/core-E1 species fused to mouse dihydrofolate reductase (mDHFR) showed cleavage in the expected region. Cleavage sites could be determined by amino-terminal protein sequencing of the DHFR-fusion partner. Our data show that there are not one but two core products with an apparent molecular weight of about 19 kDa, ending either at amino acid leucine 179 or leucine 182, respectively. These cleavages in the hydrophobic, carboxy-terminal region of HCV core suggest processing by (a) recently proposed eucaryotic signal peptide peptidase(s) (F. Lykoet al.(1995)J. Biol. Chem.270, 19873–19878). Furthermore, our results demonstrate that cleavage at these sites and the formation of the P19 species does not require previous processing at the signalase site (position 191/192) of the HCV-polyprotein.
ISSN:0042-6822
1096-0341
DOI:10.1006/viro.1996.0510