Characterization of hepatitis C virus structural proteins with a recombinant baculovirus expression system.
We cloned and expressed the sequences encoding the structural proteins of the hepatitis C virus in a baculovirus eukaryotic expression system. Four recombinant constructs expressed sufficient hepatitis C virus-specific proteins in insect cell culture to allow analysis of protein cleavage, glycosylation and immunoreactivity. Using immunoblot analysis, we detected a 22-kD protein corresponding to the hepatitis C virus capsid protein cleaved from a larger precursor. Recombinant constructs encoding the presumptive envelope (E1) protein produced products ranging from 30 to 35 kD, whereas constructs encoding the presumptive E2/NS1 protein expressed products ranging in size from 68 to 73 kD. The recombinant envelope proteins were glycosylated, as shown by sensitivity to endoglycosidase F digestion, whereas the capsid was not. We examined the immunoreactivity of these recombinant proteins using sera from 50 patients chronically infected with HCV. Forty-seven of 50 of these sera contained antibodies against the capsid, 14 (28%) also had antibodies against E1 and at least 5 (10%) had antibody against E2/NS1. Forty-seven of 50 sera (94%) were viremic, as determined on hepatitis C virus polymerase chain reaction. The three sera that were hepatitis C virus polymerase chain reaction negative did not have envelope antibodies, whereas all sera that had envelope antibodies were also hepatitis C virus polymerase chain reaction positive. Thus antibodies to baculovirus-expressed hepatitis C virus structural proteins, including E1 and E2/NS1, are found in the presence of viremia.
Department of Medicine, Veterans Affairs Palo Alto Medical Center, California 94304.
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Hsu
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Donets
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Greenberg
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Feinstone
D016367:Baculoviridae / Q000235:genetics*; D001483:Base Sequence; D004797:Enzyme-Linked Immunosorbent Assay; D006031:Glycosylation; D016174:Hepacivirus / Q000276:immunology / Q000378:metabolism*; D006526:Hepatitis C / Q000276:immunology; D006801:Humans; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000096:biosynthesis*; D015678:Viral Structural Proteins / Q000096:biosynthesis* / Q000276:immunology / Q000302:isolation & purification
D011994:Recombinant Proteins; D015678:Viral Structural Proteins