GB virus C E2 glycoprotein: expression in CHO cells, purification and characterization

• Published in: Journal of general virology Vol. 78; no. 8; pp. 1851 - 1859
• Main Authors: Surowy, TK, Leary, TP, Carrick, RJ, Knigge, MF, Pilot-Matias, TJ, Heynen, C, Gutierrez, RA, Desai, SM, Dawson, GJ, Mushahwar, IK
• Format: Journal Article
• Language: English
• Published: England Soc General Microbiol 01-08-1997
• Subjects: Amino Acid Sequence; Animals; CHO Cells; Chromatography, Affinity; Cricetinae; Enzyme-Linked Immunosorbent Assay; Flaviviridae - genetics; Flaviviridae - metabolism; Glycosylation; Hepatitis Antibodies - blood; Hepatitis, Viral, Human - diagnosis; Hepatitis, Viral, Human - epidemiology; Hepatitis, Viral, Human - immunology; Humans; Molecular Sequence Data; Molecular Weight; Recombinant Proteins - biosynthesis; Recombinant Proteins - chemistry; Recombinant Proteins - isolation & purification; Transfection; Viral Envelope Proteins - biosynthesis; Viral Envelope Proteins - chemistry; Viral Envelope Proteins - isolation & purification
• ISSN: 0022-1317; 1465-2099
• DOI: 10.1099/0022-1317-78-8-1851

TK Surowy, TP Leary, RJ Carrick, MF Knigge, TJ Pilot-Matias, C Heynen, RA Gutierrez, SM Desai, GJ Dawson and IK Mushahwar
Experimental Biology Research, Abbott Laboratories, Dept. 90D, North Chicago, IL 60064, USA. [e-mail]
A 315 amino acid recombinant segment of the GB virus C (GBV-C) E2 envelope
glycoprotein (E2-315) was expressed and secreted from CHO cells. E2-315 was
purified by affinity chromatography using a monoclonal antibody directed to
a FLAG sequence genetically engineered onto the C terminus of the
recombinant protein. The secreted protein had a molecular mass of 48-56 kDa
and was shown to be N-glycosylated. Amino acid sequencing confirmed the
expected N-terminal sequence. Purified E2-315 was used to develop an ELISA
for detection of E2 antibodies in human sera. Antibodies to GBV-C E2
appeared to be directed toward conformational epitopes since human sera
reactivity was detected in ELISA using native E2-315, but it was extremely
weak or non- existent with denatured E2 protein. The use of an ELISA which
can detect human GBV-C E2 antibodies will be important in further
understanding of the clinical significance and epidemiology of GBV-C.