Arch Virol (1997) 142: 2211±2223
Expression and interaction of the hepatitis C virus structural
proteins and the 5
in baculovirus infected cells
Y.-H. Wang, R. Trowbridge, and E. J. Gowans
Sir Albert Sakzewski Virus Research Centre, Royal Children's Hospital,
Accepted July, 1, 1997
Summary. The structural proteins, core, E1 and a C-terminal truncated E2
(C-E1-E2p) as well as the 5
UTR linked to the core gene (5
UTR-C) of the
hepatitis C virus were expressed in Sf9 cells. Expression of the C-E1-E2p
polyprotein from a single vector resulted in expression of multiple forms
suggesting that the cleavage of the polyprotein in the insect cells is incomplete.
The structural proteins were expressed as insoluble forms and were solubilized
by freeze/thaw cycles or detergent treatment. Analysis of the co-expressed
UTR-C and C-E1-E2p) proteins through a 20±60% sucrose gradient revealed
that the core protein migrated rapidly into the gradient, in common with the
sedimentation pro®le for core protein expressed from the core gene only.
Neither E1 nor E2 proteins were involved in the core-core protein interaction.
Expression of the 5
UTR-C in insect cells resulted in a high level of mRNA
transcription but no translation of core protein, presumably due to a failure to
initiate HCV translation by an internal ribosome entry mechanism, since the
mRNA transcript was translated in vitro, suggesting that the 5
in¯uence HCV tropism.
Hepatitis C virus (HCV) is the major cause of non-A, non-B hepatitis. The viral
genome comprises a positive sense single strand RNA molecule that contains a
single open reading frame which encodes a polyprotein of about 3010 amino
acids . The polyprotein is cleaved to produce three structural proteins (core
and envelope proteins E1 and E2) and six nonstructural proteins (NS2 to NS5B)
. The 5
untranslated region (5
UTR) of the genome is 341 nt and forms a
stable structure which is similar to that of the 5
UTR region of pesitiviruses .
Recently, it was shown that the 5
UTR functions as an internal ribosomal entry
site (IRES) for HCV translation [12, 35±37] that extends into the core protein
coding region .