Expression of membrane-bound and secreted forms of equine herpesvirus 1 glycoprotein D by recombinant baculovirus.
Analyses of the synthesis and processing of recombinant full-length glycoprotein D of equine herpesvirus type 1 (EHV-1; gD392) or recombinant truncated gD (gD352) expressed in baculovirus-infected Sf9 cells revealed the following: (1) gD polypeptides encoded by both recombinant baculoviruses react with gD-specific antibodies including peptide-specific antiserum that neutralizes EHV-1 in a plaque reduction assay, (2) both the full-length recombinant gD392 and the truncated gD352 are expressed predominantly as gD species that contain high mannose-type oligosaccharides (55 kDa and 52 kDa, respectively), (3) both the full-length recombinant gD392 and the truncated gD352 are also expressed in lesser amounts as gD species that contain complex-type oligosaccharides (58 kDa and 55 kDa, respectively) as well as the unglycosylated forms of gD (43 kDa and 37 kDa, respectively), (4) flow cytometric analyses of cells expressing gD392 revealed that gD first appears on the cell surface at 24 h post infection; by 60 h, 95% of the cells express high levels of cell surface gD, (5) cells expressing gD352, in contrast to cells expressing gD392, secrete gD into the extracellular medium. This initial demonstration that immunoreactive EHV-1 glycoprotein D can be produced as a secreted polypeptide in the baculovirus system should provide reagents to assess the potential use of gD as a subunit vaccine in an animal model.
Flowers
C C
CC
Department of Microbiology and Immunology, Louisiana State University Medical Center, Shreveport 71130-3932, USA.
Flowers
S P
SP
Sheng
Y
Y
Tarbet
E B
EB
Jennings
S R
SR
O'Callaghan
D J
DJ
D000818:Animals; D000914:Antibodies, Viral / Q000276:immunology; D002460:Cell Line; D005822:Genetic Vectors / Q000235:genetics*; D006031:Glycosylation; D004861:Herpesvirus 1, Equid / Q000235:genetics* / Q000276:immunology; D008565:Membrane Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D017924:Nucleopolyhedroviruses / Q000235:genetics*; D009844:Oligosaccharides / Q000032:analysis; D011499:Protein Processing, Post-Translational; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000276:immunology / Q000378:metabolism; D018411:Spodoptera; D014759:Viral Envelope Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology / Q000378:metabolism
D000914:Antibodies, Viral; D008565:Membrane Proteins; D009844:Oligosaccharides; D011993:Recombinant Fusion Proteins; D014759:Viral Envelope Proteins; C488988:glycoprotein D, Equid herpesvirus 1