Hepatitis C virus core, NS3, NS4B and NS5A are the major immunogenic proteins in humoral immunity in chronic HCV infection

Maarit Sillanpaa,Maija Lappalainen,Paivi Porkka,Krister Melen,Kaisu Nevalainen,Ilkka Julkunen,Riku Fagerlund

doi:10.1186/1743-422x-6-84

Abstract

BackgroundThe viral genome of hepatitis C virus constitutes a 9.6-kb single-stranded positive-sense RNA which encodes altogether 11 viral proteins. In order to study the humoral immune responses against different HCV proteins in patients suffering from chronic HCV infection, we produced three structural (core, E1 and E2) and six nonstructural proteins (NS2, NS3, NS4A, NS4B, NS5A and NS5B) in Sf9 insect cells by using the baculovirus expression system.ResultsThe recombinant HCV core, E1, E2, NS2, NS3, NS4A, NS4B, NS5A and NS5B proteins were purified and used in Western blot analysis to determine antibody responses against individual HCV protein in 68 HCV RNA and antibody positive human sera that were obtained from patients suffering from genotype 1, 2, 3 or 4 infection. These sera were also analysed with INNO-LIA Score test for HCV antibodies against core, NS3, NS4AB and NS5A, and the results were similar to the ones obtained by Western blot method. Based on our Western blot analyses we found that the major immunogenic HCV antigens were the core, NS4B, NS3 and NS5A proteins which were recognized in 97%, 86%, 68% and 53% of patient sera, respectively. There were no major genotype specific differences in antibody responses to individual HCV proteins. A common feature within the studied sera was that all except two sera recognized the core protein in high titers, whereas none of the sera recognized NS2 protein and only three sera (from genotype 3) recognised NS5B.ConclusionThe data shows significant variation in the specificity in humoral immunity in chronic HCV patients.

Highlights

The viral genome of hepatitis C virus constitutes a 9.6-kb single-stranded positivesense RNA which encodes altogether 11 viral proteins
We have described the expression and purification of nine different recombinant Hepatitis C virus (HCV) proteins in insect cells and analyzed humoral immune response against each viral protein using Western blotting in patients suffering from chronic HCV infection of genotypes 1, 2, 3 or 4
The PCR fragments encoding for different HCV proteins were subcloned into the pAcYM1 baculovirus transfer vector, and baculoviruses expressing the recombinant core, E1, E2, NS2, NS3, NS4A, NS4B, NS5A, and NS5B proteins were constructed

Summary

Introduction

The viral genome of hepatitis C virus constitutes a 9.6-kb single-stranded positivesense RNA which encodes altogether 11 viral proteins. The viral genome constitutes a 9.6-kb single-stranded positive-sense RNA with 5' and 3' noncoding regions and a long open reading frame encoding a polyprotein precursor of about 3,000 amino acids in length. The core region encodes for an alternative open reading frame protein (ARFP) or F protein whose function is presently not known [1,3]. The region between the structural and nonstructural genes encodes for an integral membrane cation channel protein p7 [4] which is essential for virus production [5]. There is little information of the function of NS4B protein, but it participates in the formation of a membranous web where HCV RNA replication is suggested take place [6,7]. The NS5B protein encodes for an RNA-dependent RNA polymerase (RdRp), which is the central catalytic enzyme of the HCV replicase [9,10]

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