Expression of Epstein-Barr Virus Membrane Antigen gp340/220 in Mouse Fibroblasts Using a Bovine Papillomavirus Vector

Abstract

SUMMARYEpstein-Barr virus (EBV) membrane antigen glycoproteins gp340 and gp220 are encoded by a single gene. We have inserted this gene into a bovine papillomavirus (BPV) vector and expressed gp340/220 in mammalian cells under the control of the mouse metallothionein promoter. The proteins produced were of similar Mr, showed similar antigenic specificity and were transported to the same subcellular location as the authentic gp340/220. The inclusion of heavy metal ions in the medium had no effect on the levels of gp340/220, which were approximately the same as those found in standard EBV-transformed lymphoblastoid cell lines, e.g. B95-8. Cells that expressed gp340/220 were selected by several rounds of fluorescence-activated cell sorting, but on passage they rapidly lost the ability to express this glycoprotein. In contrast to this we found that BPV-transformed cells expressing a truncated version of gp340/220 still produced it at significant levels after extended passage.

The Effect of Bovine Herpesvirus Type 1 Glycoproteins gI and gIII on Herpesvirus Infections

“…Since the submission of our manuscript, Conway et al (1989) have reported on the use of a BPV vector to express gp340/220 of Epstein-Barr virus in murine cells with an MMT promoter. Unlike our system, they did not increase expression of this protein following induction with heavy metals.…”

Purification and quantification of recombinant Epstein-Barr viral glycoproteins gp350/220 from Chinese hamster ovary cells

“…Although cell-specific post-translational modifications critically influence the antigenic presentation of EBV-MA, all mammalian cell-derived versions of EBV-MA were found capable of inducing EBV-specific neutralizing antibodies [2]. As expressing levels of recombinant EBV-MA were low, suggesting that the protein is toxic for eukaryotic cells, truncated forms of EBV-MA lacking the membrane anchor have been expressed [2,27,28]. Truncated EBV-MA was secreted into the medium of transformed cells in culture.…”

Expression and immunogenic characterization of recombinant gp350 for developing a subunit vaccine against Epstein-Barr virus

“…In CHO and GH3 (rat pituitary cells) expression systems, the yield of recombinant gp350 was only 5 mg/L of culture medium (Jackman et al 1999;Whang et al 1987). About 50 μg gp350 was obtained from 1×10 6 mouse fibroblasts (Conway et al 1989). As a result, the yield of purified gp350 1-443 in P. pastoris (120.34 mg/L) was at least 24-fold higher than that obtained in mammalian cell systems.…”