Hepatitis C virus core protein is a dimeric alpha-helical protein exhibiting membrane protein features.
The building block of hepatitis C virus (HCV) nucleocapsid, the core protein, together with viral RNA, is composed of different domains involved in RNA binding and homo-oligomerization. The HCV core protein 1-169 (C(HCV)169) and its N-terminal region from positions 1 to 117 (C(HCV)117) were expressed in Escherichia coli and purified to homogeneity suitable for biochemical and biophysical characterizations. The overall conformation and the oligomeric properties of the resulting proteins C(HCV)169 and C(HCV)117 were investigated by using analytical centrifugation, circular dichroism, intrinsic fluorescence measurements, and limited proteolysis. Altogether, our results show that core protein (C(HCV)169) behaves as a membranous protein and forms heterogeneous soluble micelle-like aggregates of high molecular weight in the absence of detergent. In contrast, it behaves, in the presence of mild detergent, as a soluble, well-folded, noncovalent dimer. Similar to findings observed for core proteins of HCV-related flaviviruses, the HCV core protein is essentially composed of alpha-helices (50%). In contrast, C(HCV)117 is soluble and monodispersed in the absence of detergent but is unfolded. It appears that the folding of the highly basic domain from positions 2 to 117 (2-117 domain) depends on the presence of the 117-169 hydrophobic domain, which contains the structural determinants ensuring the binding of core with cellular membranes. Finally, our findings provide valuable information for further investigations on isolated core protein, as well as for attempts to reconstitute nucleocapsid particles in vitro.
Boulant
Steeve
S
Institut de Biologie et Chimie des Protéines, UMR5086 CNRS-Université Claude Bernard Lyon I, France.
Vanbelle
Christophe
C
Ebel
Christine
C
Penin
François
F
Lavergne
Jean-Pierre
JP
D000595:Amino Acid Sequence; D003902:Detergents / Q000494:pharmacology; D004926:Escherichia coli / Q000378:metabolism; D016174:Hepacivirus / Q000737:chemistry*; D008565:Membrane Proteins / Q000737:chemistry; D008956:Models, Chemical; D008969:Molecular Sequence Data; D016415:Sequence Alignment; D014758:Viral Core Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism
D003902:Detergents; D008565:Membrane Proteins; D014758:Viral Core Proteins; C069130:nucleocapsid protein, Hepatitis C virus