PMID: 9725668Sep 2, 1998Paper

Secretion and purification of HCV E1 protein forms as glutathione-S-transferase fusion in the baculovirus insect cell system

Virus Research
A R CiccaglioneM Rapicetta

Abstract

We have expressed the E1 protein of Hepatitis C Virus (HCV) in a new recombinant form by using a baculovirus transfer vector directing the expression of proteins fused to the carboxy-terminus of glutathione-S-transferase (GST). The E1 domain was expressed varying at its carboxy terminus in order to retain (GST-E1) or delete (GST-E1b) the C-terminal hydrophobic region that may be involved in membrane association. Following infection with the recombinant virus, GST-E1b was efficiently secreted into the culture media and could be purified in a single step with the minimum of denaturation by glutathione affinity chromatography. The purified product was specifically immunoprecipitated by HCV positive human sera suggesting the maintenance of an immuno-relevant tertiary structure despite removal of the hydrophobic anchor. By contrast, cells infected with a recombinant baculovirus expressing GST-E1 gave a fusion protein with an appropriate molecular weight but also a series of polypeptides of lower molecular weight consistent with cleavage at the C-terminus of E1. GST-E1 was not secreted into the medium and was associated predominantly with the membrane fraction following cell disruption; the lower molecular weight forms were soluble a...Continue Reading

References

Jul 1, 1991·Proceedings of the National Academy of Sciences of the United States of America·M HijikataK Shimotohno
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May 1, 1996·The Journal of General Virology·A Fournillier JacobC Wychowski

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Citations

Aug 30, 2008·PloS One·José Manuel CuevasFernando González-Candelas
Nov 28, 2001·Protein Expression and Purification·T MiyasakaY Miyamoto
Feb 6, 2004·Biochimica Et Biophysica Acta·Cédric MontignyPierre Falson
Jan 1, 2012·Asian Pacific Journal of Cancer Prevention : APJCP·Hossein KeyvaniHamid Reza Mollaie
Aug 22, 2001·The Journal of General Virology·A R CiccaglioneM Rapicetta

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