The production of human parvovirus capsid proteins in Escherichia coli and their potential as diagnostic antigens.
We have expressed a number of polypeptides derived from the capsid proteins of the human parvovirus B19 in Escherichia coli. These include native VP1 (84K) and VP2 (58K) proteins and also fusions to beta-galactosidase containing differing amounts of the amino terminus of the VP1/2 polypeptide. Although each of these was expressed at high levels and the majority were produced as full-length proteins, only one was soluble. This soluble polypeptide, p132, is a beta-galactosidase fusion protein that includes 145 amino acids from B19 which are entirely derived from the region unique to VP1. Despite containing such a small portion of VP1, which itself constitutes only 4% of total capsid protein, p132 reacted with all our known anti-B19 IgM-positive human serum samples. We conclude that this region contains epitopes which must be prominently exposed on the intact virus. We have demonstrated the use of this recombinant antigen in a simple diagnostic assay for B19-specific antibodies which can be used for initial screening of human serum samples. In a survey of 103 serum specimens, our ELISA positively identified all samples (19/19) which were positive by IgM antibody capture radioimmunoassay. The recombinant p132 antigen is efficiently produced and readily purified from E. coli, and its use as a diagnostic antigen should increase the availability of routine clinical testing for human parvovirus infection.
Department of Molecular Biology, Wellcome Biotech, Beckenham, Kent, U.K.
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Rayment
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Crosdale
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Morris
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Pattison
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Talbot
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D000956:Antigens, Viral / Q000276:immunology*; D001483:Base Sequence; D002213:Capsid / Q000235:genetics* / Q000276:immunology; D036022:Capsid Proteins; D003001:Cloning, Molecular; D003429:Cross Reactions; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes; D004926:Escherichia coli / Q000235:genetics*; D006801:Humans; D007075:Immunoglobulin M / Q000276:immunology; D007763:Lac Operon; D008969:Molecular Sequence Data; D010321:Parvoviridae / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D010957:Plasmids; D011994:Recombinant Proteins / Q000276:immunology; D001616:beta-Galactosidase / Q000235:genetics
D000956:Antigens, Viral; D036022:Capsid Proteins; D000939:Epitopes; D007075:Immunoglobulin M; D011994:Recombinant Proteins; D001616:beta-Galactosidase