Expression of env sequences of the bovine leukemia virus (BLV) in the yeast Saccharomyces cerevisiae.
DNA sequences of the envelope (env) gene of the bovine leukemia virus (BLV) were expressed in the yeast Saccharomyces cerevisiae. Two yeast promoters, the repressible PHO5 promoter and the constitutive PGK promoter, were used to construct four expression plasmids comprising either a sequence of the surface antigen gp51 or a (gp51 + gp30) sequence. The expressed heterologous gene products were characterized by Western blot analysis and competitive radioimmunoassay. By means of Northern blot analysis the steady-state level of env-specific mRNA was analysed. The highest expression rate was obtained from recombinant plasmid YEpSG 94 comprising a gp51 sequence--a 630 base pair fragment containing 70% of the gp51 but lacking the N terminus--as well as the PHO5 promoter including PHO5 signal sequence and the PHO5 terminator. The recombinant gp51 was partially glycosylated but the PHO5 signal peptide did not seem to be cleaved off. No immunoreactive material could be found in the periplasm or in the culture medium. By means of monoclonal antibodies directed against eight different epitopes of viral gp51, all four sequential antigenic determinants were detected in the AH 216 (YEpSG 94) expression product.
Brantl
S
S
Central Institute of Microbiology and Experimental Therapy, Academy of Sciences of the GDR, DDR, Jena.
Eldarov
M A
MA
Rössler
H
H
Drescher
B
B
Lang
H
H
Rosenthal
S
S
Skryabin
K G
KG
D000595:Amino Acid Sequence; D000954:Antigens, Surface / Q000235:genetics; D000956:Antigens, Viral / Q000235:genetics; D001483:Base Sequence; D015152:Blotting, Northern; D015153:Blotting, Western; D003001:Cloning, Molecular; D004279:DNA, Viral / Q000235:genetics*; D005786:Gene Expression Regulation; D005822:Genetic Vectors; D007124:Immunoenzyme Techniques; D001909:Leukemia Virus, Bovine / Q000235:genetics*; D008565:Membrane Proteins / Q000235:genetics; D008938:Mitosis; D008969:Molecular Sequence Data; D010957:Plasmids; D011401:Promoter Regions, Genetic; D021382:Protein Sorting Signals / Q000235:genetics; D012333:RNA, Messenger / Q000032:analysis; D011863:Radioimmunoassay; D015183:Restriction Mapping; D012190:Retroviridae / Q000235:genetics*; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D014764:Viral Proteins / Q000235:genetics
D000954:Antigens, Surface; D000956:Antigens, Viral; D004279:DNA, Viral; D008565:Membrane Proteins; D021382:Protein Sorting Signals; D012333:RNA, Messenger; D014764:Viral Proteins