SELF-ASSOCIATION OF THE C-TERMINAL DOMAIN OF THE HEPATITIS-C VIRUS CORE PROTEIN

The N-terminal region of the hepatitis-C virus (HCV) core protein is r ich in basic residues, while the C-terminal end of the protein compris es of a stretch of hydrophobic amino acids. Between these two extremes is an amphipathic region with two predicted a-helical segments. This region embodies Leu or hydrophobic residues in positions of heptad rep eats and is possibly capable of self-association. To investigate this possibility, the core sequence was divided into two fragments and expr essed separately as recombinant proteins. Recombinant proteins with th e N-terminal fragment remained as monomers even at high concentrations in SDS/PAGE. Recombinant protein with the C-terminal fragment appeare d largely monomeric on denaturing gels but some oligomers were also de tected. Furthermore, proline mutations in either one of the predicted a helices adversely affected the observed oligomerization. The self-as sociation capacity of the core protein C-terminal region was further s upported by results from a yeast two-hybrid system. To affirm our conc lusion, a peptide covering the heptad repeats and the predicted a heli ces was synthesized. Data from mass spectrometry and gel-filtration ch romatography concluded that this peptide readily self-associated into the homodimer. Therefore, our results suggest that the oligomerization motifs of the HCV core protein may not be limited to the previously s uggested N-terminal region.